LITHUANIAN UNIVERSITY OF HEALTH SCIENCES VETERINARY ACADEMY

Erika Mozūrienė

APPLICATION OF SOLID-STATE FERMENTATION FOR DEVELOPMENT OF

HIGHER VALUE AND SAFETY FOOD PRODUCTS

Doctoral Dissertation Agricultural Sciences,

Zootechnics (03A)

Kaunas, 2016

Dissertation has been prepared at the Department of Food Safety and Quality of Veterinary Academy of Lithuanian University of Health Sciences during the period of 2012–2016.

Scientific Supervisor Prof. Dr. Elena Bartkienė (Lithuanian University of Health Sciences,

Agricultural Sciences, Zootechnics – 03A) Dissertation is defended at the Zootechnical Research Council of the

Veterinary Academy of Lithuanian University of Health Sciences. Chairperson Prof. Habil. Dr. Romas Gružauskas (Lithuanian University of Health

Sciences, Agricultural Sciences, Zootechnics – 03A). Members Prof. Dr. Vaidas Oberauskas (Lithuanian University of Health Sciences,

Agricultural Sciences, Veterinary – 02A); Dr. Violeta Razmaitė (Lithuanian University of Health Sciences,

Agricultural Sciences, Zootechnics – 03A); Prof. Dr. Pranas Viškelis (Kaunas University of Technology, Physical

Sciences, Chemistry – 03P); Ass. Prof. Dipl.-Ing. Dr. Gerhard Schleining (BOKU – University of

Natural Resources and Life Sciences, Vienna, Austria, Technological Sciences, Chemical Engineering – 05T).

Dissertation will be defended at the open session of the Lithuanian University of Health Sciences on the 22nd of December, at 10:00 am in Dr. S. Jankauskas Auditorium of the Veterinary Academy.

Address: Tilžės 18, LT-47181 Kaunas, Lithuania.

LIETUVOS SVEIKATOS MOKSLŲ UNIVERSITETAS VETERINARIJOS AKADEMIJA

Erika Mozūrienė

VERTINGESNIŲ IR SAUGESNIŲ MAISTO PRODUKTŲ KŪRIMAS TAIKANT

AUGALINĖS ŽALIAVOS KIETAFAZĘ FERMENTACIJĄ

Daktaro disertacija Žemės ūkio mokslai, zootechnika (03A)

Kaunas, 2016

Disertacija rengta 2012–2016 metais Lietuvos sveikatos mokslų universitete Veterinarijos akademijos Maisto saugos ir kokybės katedroje.

Mokslinė vadovė Prof. dr. Elena Bartkienė (Lietuvos sveikatos mokslų universitetas, žemės

ūkio mokslai, zootechnika – 03A) Disertacija ginama Lietuvos sveikatos mokslų universiteto

Veterinarijos akademijos Zootechnikos mokslo krypties taryboje: Pirmininkas Prof. habil. dr. Romas Gružauskas (Lietuvos sveikatos mokslų

universitetas, žemės ūkio mokslai, zootechnika – 03A). Nariai: Prof. dr. Vaidas Oberauskas (Lietuvos sveikatos mokslų universitetas,

žemės ūkio mokslai, veterinarija – 02A); Dr. Violeta Razmaitė (Lietuvos sveikatos mokslų universitetas, žemės

ūkio mokslai, zootechnika – 03A); Prof. dr. Pranas Viškelis (Kauno technologijos universitetas, fiziniai

mokslai, chemija – 03P); Doc. dr. inž. Gerhard Schleining (BOKU – Gamtos išteklių ir gyvosios

gamtos mokslų universitetas, Viena, Austrija, technologijos mokslai, chemijos inžinerija – 05T).

Disertacija bus ginama viešame Zootechnikos mokslo krypties tarybos

posėdyje 2016 m. gruodžio 22 d. 10 val. Lietuvos sveikatos mokslų universiteto Veterinarijos akademijos dr. S. Jankausko auditorijoje.

Adresas: Tilžės g. 18, LT-47181 Kaunas, Lietuva.

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TABLE OF CONTENTS

ABREVIATIONS ............................................................................... 7 INTRODUCTION .............................................................................. 8

The aim of the study ........................................................................ 10 The objectives of the study .............................................................. 10 The scientific novelty and practical usefulness ................................. 10

1. LITERATURE REVIEW .............................................................. 12 1.1. Lactic acid bacteria and their regulation for food uses .......... 12 1.2. Design of the fermentation processes – solid state and submerged fermentation .............................................................. 16 1.3. Fermented plant products for the food of animal origin nutritional, technological and safety parameters improving ......... 17 1.4. Changes of the meat during lactic acid fermentation process and the influence of the lactic acid bacteria on smoked meat products safety parameters .................................................. 24 1.5. Lactic acid fermentation in unripened cheese production ...... 27

2. MATERIALS AND METHODS ................................................... 29 2.1. Investigation venue .............................................................. 29 2.2. Materials .............................................................................. 29 2.3. Lactic acid bacteria cultivation and plant fermentation ......... 31 2.4. Methods of evaluating plant bioproducts microbiological, physical chemical, enzymatical and antimicrobial activity parameters .................................................................................. 32 2.5. Technology of food products ................................................ 35

2.5.1. The use of fermented S. montana and S. hortensis bioproducts for beef and pork loin marination .................... 35 2.5.2. The use of lactic acid bacteria – potato juice marinade for meat treatment............................................... 35 2.5.3. Production of cold smoked pork sausages ................. 36 2.5.4. Ready-to-cook minced pork meat products technology ......................................................................... 37 2.5.5. Unripened curd cheese technology ........................... 38

2.6. Methods of evaluating food products microbiological, physical chemical, sensory and technological parameters ............ 38 2.7. Statistical analysis ................................................................ 42

3.RESULTS ........................................................................................ 43 3.1. Microbiological and physical chemical parameters of plants and their bioproducts ........................................................ 43

3.1.1. Parameters of fermented and nonfermented savory plants ................................................................................. 43

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3.1.2. Parameters of fermented defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke .............................. 46

3.2. Technological, microbiological and physical chemical parameters of meat products ....................................................... 56

3.2.1. Parameters of marinated meat products .................... 56 3.2.2. Parameters of cold smoked pork sausages ................ 89 3.2.3. Parameters of ready-to-cook minced pork meat products ............................................................................. 93

3.2.3.1. Parameters of ready-to-cook minced pork meat products produced with biotreated Satureja montana .... 93 3.2.3.2. Parameters of ready-to-cook minced pork meat products produced with biotreated Satureja hortensis .... 102 3.2.3.3. Parameters of ready-to-cook minced pork meat products produced with pea fiber and semolina ............. 109

3.3. The parameters of unripened curd cheese produced with Satureja montana and Rhaponticum carthamoides bioproducts .. 110

4. DISCUSSION ................................................................................. 119 4.1. Lactic acid bacteria – plant bioproducts – alternative preservatives for a higher value food of animal origin production .................................................................................. 119 4.2. Changes of bioactive compounds in plants during lactic acid fermentation process ........................................................... 121 4.3. Marinades based on lactic acid bacteria cultivated in an alternative substrate for improving meat quality parameters indifferent part of pork, beef and chicken .................................... 124 4.4. New by developed plant bioproducts for improving ready-to- cook minced pork, pork and beef loin quality and safety parameters .................................................................................. 129 4.5. Pea fiber incorporation in the formula of gluten-free meat products ...................................................................................... 132 4.6. Lactic acid bacteria for decreasing the polycyclic aromatic hydrocarbons content in cold smoked pork sausages ................... 134 4.7. Savory plant bioproducts for higher sustainability unrippened curd cheese production ............................................. 136

CONCLUSIONS ................................................................................ 138 REFERENCES .................................................................................. 141 PUBLICATIONS ............................................................................... 175 SUMMARY IN LITHUANIAN ........................................................ 213 APPENDIXES ................................................................................... 256 CURRICULUM VITAE .................................................................... 266 ACKNOWLEDGEMENTS ............................................................... 267

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ABBREVIATIONS

BAs – Biogenic Amines BIOR – Institute of Food Safety, Animal Health and Environment BLIS – Bacteriocin-Like Inhibitory Substances GC/MS – Gas Chromatography – Mass Spectrometry GF – Gluten free GRAS – Generally Recognized As Safe HAA – Heterocyclic Aromatic Amines HPLC – MS/MS – High-Performance Liquid Chromatography – Mass Spectrometry IF – Intramuscular Fat KTU – Kaunas University of Technology LAB – Lactic Acid Bacteria Ls – Lactobacillus sakei LUHS VA – Lithuanian University of Health Sciences Veterinary Academy MBL – Marinated Beef Loin MPL – Marinated Pork Loin MRS – de Man Rogosa Sharpe Pa – Pediococcus acidilactici Pp – Pediococcus pentosaceus Rc – Rhaponticum carthamoides RCMP – Ready-to-Cook Minced Pork meat products Sh – Satureja hortensis L. Sm – Satureja montana L. SMF – Submerged Fermentation SSF – Solid State Fermentation TPC – Total Phenolic Compounds TTA – Total Titratable Acidity UCC – Unripened Curd Cheese VC – Volatile Compounds VMU – Vytautas Magnus University VRBA – Violet Red Bile glucose Agar WHC – Water Holding Capacity RSA – Radical Scavenging Activity BaA – Benz[a]anthracene BbF – Benzo-[b]fluoranthene BaP – Benzo[a]pyrene Chr – Chrysene

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INTRODUCTION

The food industry seeks alternatives to satisfy consumers demands of safe, minimally processed food in which chemical preservatives are replaced by more natural alternatives [19, 141, 246]. Lactic acid bacteria (LAB) have been used in food fermentations all over the world for millennia. LAB are an integral part of food safety, keeping the quality and the nutritional value of many foodstuffs [122].

Traditionally LAB are natural constituents of fermented foods, however, the application of antimicrobial compounds producing protective cultures may provide an additional parameter of bioprotection in several different food matrices and ensure food quality, keeping or enhancing its organoleptic, textural characteristics and nutritional aspects in the final product (e.g., meat or cheese) [19, 41, 115, 122, 277-9, 289]. Many LAB produce a high diversity of different bacteriocins. Bacteriocinogenic LAB are generally recognised as safe (GRAS) and useful to control the frequent development of pathogens and spoilage microorganisms [233]. Bacteriocins can be produced in foods by the activity of bacteriocin-producing LAB strains or when added in foods as food preservatives [346]. The use of bacteriocins has emerged as an important strategy to increase food safety and to minimize the incidence of foodborne diseases due to their minimal impact on the nutritional and sensory properties (low toxicity and stability against proteases and temperature) of food products [83, 119, 309]. They can be used in the production of several foods (meat, chicken, dairy, eggs, seafoods, fruit and vegetables) [117]. The direct addition of bacteriocin-producing cultures into products can be a more practical and economic option for improvings of the safety and quality of the final product [233].

With the advent of biotechnological innovations, mainly in the area of the fermentation technology, many new avenues have been opened [178]. Biotechnology offers significant advantages, such as a high concentration of metabolites, to obtain product stability and the adaptability of microorganisms with a low free water content. Over the last two decades, solid state fermentation (SSF) has gained significant attention for the development of industrial bioprocesses, particularly due to a lower energy requirement associated with higher product yields and a less wastewater production with a lower risk of bacterial contamination. In addition, it is an ecofriendly alternative to produce different fermented values added products like enzymes, organic acids, bio-colours, bio-flavors, bio-pesticides, bio-surfactants, as it mostly utilizes solid plant material, which eventually results in the greener and cleaner environment [123, 234, 376].

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One of the principal challenges of the food industry is to find the best combination among various food processing methods to increase the concentration of bioactive compounds in food to ensure of their nutritional and healthy properties for humans [65, 190, 292, 331]. LAB are used as a flavour, an acidifying agent having biodegradable properties and controlled release and/or increasing the content of specific beneficial compounds [154, 329]. These compounds can be macronutrients, micronutrients (such as vitamins) or non-nutritive compounds [329]. LAB fermentation has been shown to increase the levels of nutrients including folates, soluble dietary fiber, and the total content of phenolic compounds in legumes and dietary fiber-rich pants, thus enhancing their antioxidant activity, improving the protein digestibility [86, 242, 378], increasing lignans and positively influencing the content of alkylresorcinols [308]. Consequently, fermented plants could be an excellent material for enriching the food of animal origin.

It is known that savory plants contain compounds with antimicrobial properties [249]. Such a complex (savory plants – bacteriocins producing LAB) could be promising for the preservation of food products. In this way, LAB and plants that combine different functional characteristics could be useful for developing improved or new foods.

Meat fermentation of is a well-known method to extend the shelf life, transformation and diversification of meat products [108]. Meat and meat products are a concentrated source of proteins, however, they can rapidly spoil and may allow the growth of food-borne pathogenic microorganisms. This is why treatement with fermented savory plant bioproducts become an important preservation technology. LAB and savory plant compounds can prevent the growth of undesirable food pathogens and spoilage bacteria, form texture and taste [199, 210].

Fermented dairy products are the most common fermented foods, because they are convenient, nutritious, stable, natural, and healthy [23]. LAB can show the desired technological and functional potential in milk protein coagulation, production of proteinases [385], lactic acid, acetic acid, ethanol, acetaldehyde, diacetyl, aroma compounds, bacteriocins, exopolysaccharides, and several enzymes of importance to be controlled. These compounds can enhance the shelf life and microbial safety, improve texture, and contribute to the pleasant sensory characteristics of dairy products, but they may also cause spoilage in uncontrolled conditions [134].

Food technology innovation has the potential to deliver many benefits to society from improved food safety and food risk mitigation and improved nutrition to increasing food sustainability and improving food quality [100, 312]. Technological innovation in food production may involve the application of emerging technologies associated with societal disquiet, such

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as biotechnology. However, fermented foods can contain biogenic amines (BAs) whose content should be limited in food [158] and which are often related with BAs poisoning [92]. Also, the main metabolyte of LAB is lactate, which has two otical isomers – L(+) and D(-) lactate. Increased levels of D(-) lactic acid in plasma and urine have been demonstared in cases of intesinal ischaemia, short bowel and appendicitis, and are considered as a marker of dysbiosis and/or increased intestinal permeability [391]. Therefore, the desirable lactic acid isomer in food is L(+).

The aim of the study To develop new bioproducts by using bacteriocins-producing LAB and

plants having a unique chemical composition: the high content of protein, phytoestrogens, inulin, phenolic compounds, and essential oils (EOs), and to adapt them for the production of a higher value and sustainability safer food of animal origin production.

The objectives of the study

1. To select the conditions for plant material fermentation and to evaluate the parameters of fermentation efficiency.

2. To evaluate the content changes of bioactive compounds (alkylresorcinols (ARs), lignans, β-glucans, and volatile compounds (VC)), total phenolic compounds (TPC), and radical scavenging activity (RSA) of plants during biotreatment.

3. To carry out the comparison of SSF and SMF bioprocesses and to select safe, having antimicrobial properties plant bioproducts for the production of higher value and sustainability food.

4. To create a design of technologies for higher value and safer sustainability food products, and to improve the safety of the traditional food technologies by using selected LAB – plant bioproducts.

The scientific novelty and practical usefulness According to the World Health Organization, access to sufficient

amounts of safe and nutritious food is the key to sustaining life and promoting good health. As the world’s population grows, the intensification and industrialization of agriculture and animal production to meet the increasing demand for food creates both opportunities and challenges for food safety. The development of new, higher value and sustainability food technologies and assessing their safety become very important. The exploration of naturally occurring antimicrobials for food preservation receives increasing attention due to consumer awareness of natural food

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products and the growing concern of microbial resistance to conventional preservatives. The use of the LAB-plant complex bioproducts as an alternative to the use of synthetic chemicals to preserve the quality and to increase the functional value of food of animal origin could be a choice. The dissertation is dedicated to solving this particular problem.

The scientific and practical novelty of the thesis is concentrated on developing technologies of the new higher value and safer food of animal origin, based on the treatment with the LAB-plant complex bioproducts or their incorporation in the food formula. For this purpose, in Lithuania cultivated plants (source of biologically active compounds) (I) and SSF with selected LAB (II) (for plant substrate biomodification with the purpose to produce additives for improving the technological and safety parameters of animal origin), will be used. The SSF technology and the unique properties of plants applicated in the food of animal origin production would be novel and perspective to reduce the use of synthetic additives and conservants.

Important practical results will be obtained: (I) LAB-plant complex bioproducts will be created; (II) the new, of higher value and sustainability, safer food of animal origin and its technologies prototypes will be developed, and safe food supplies will support national economies, trade and tourism, contribute to food and nutrition safety, underpin sustainable development.

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LITERATURE REVIEW

1.1. Lactic acid bacteria and their regulation for food uses

LAB are widespread microorganisms which can be found in any environment rich mainly in carbohydrates, such as plants, fermented foods and the mucosal surfaces of humans, terrestrial and marine animals. In human and animal bodies, LAB are part of the normal microbiota or microflora, the ecosystem that naturally inhabits the gastrointestinal and genitourinary tracts, which is comprised by a large number of different bacterial species with a diverse amount of strains [20, 22]. They constitute a Gram-positive heterogeneous group of microorganisms that produce lactic acid as the major metabolite during the fermentation process and initiate rapid and adequate acidification in the raw materials through the production of various organic acids from carbohydrates [12, 27]. The main members of the LAB are Lactococcus, Lactobacillus, Streptococcus, Leuconostoc, Pediococcus, Carnobacterium, Aerococcus, Enterococcus, Oenococcus, Tetragenococcus, Vagococcus and Weisella. Lactobacillus is the largest genus of this group, comprising around 80 recognized species [166, 340]. LAB have limited biosynthetic capabilities and thus require continuous supply of purines, pyrimidines, vitamins, and amino acids. These are nonsporing nonmotile organisms and usually categorized as facultative anaerobes. Lactobacillus strains are used in pickle, sauerkraut, beer, wine, juices, cheese, yogurt, and sausage production [22, 306]. Some LAB strains (including Enterococcus faecium, Lactobacillus plantarum, Lactobacillus acidophilus, Lactobacillus casei subsp. rhamnosus, and several Bifidobacterium and Propionibacterium species) from animal and human intestinal microflora have been adopted as ”probiotic“ food supplements [374]. However, especially LAB are assessed for their food protective properties. The biopreservative role of LAB is mainly due to the synthesis of a wide range of active metabolites which include: organic acids (lactic, acetic, formic, propionic, and butyric acids), or compounds, such as carbon dioxide, ethanol, hydrogen peroxide, fatty acids, acetoin, diacetyl, antifungal compounds (propioniate, phenyllactate, hydroxyphenyl-lactate, cyclic dipepetides and 3-hydroxy fatty acids), food aromas and flavors (e.g., diacetyl and acetaldehyde) [273, 289], proteinaceous, small heat-stable inhibitory peptides bacteriocins (e.g. nisin, reuterin, reutericyclin, pediocin, lacticin, enterocin), or bacteriocin-like inhibitory substances (BLIS) [12, 263, 293, 317, 352].

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It is well known that the production of acid based compounds play an important role in the preservative effect of LAB and can inhibit growth of the food spoilage microorganims [263, 314, 317]. The antimicrobial effect of acids is due to the fact that undissociated acids can pass through the microbial lipid membranes and disrupting the host cell proton motive force [263, 317]. These bacteria not only produce lactic acid but also preserve nutrients, vitamins and are used as starter cultures to convert sugars into lactic acid and other end products which give the typical flavour to fermented products [275]. The production of LAB antimicrobial compounds is dependent on the selected strain, growth conditions, and the interactions between metabolites [213].

Second group of antimicrobial compounds produced by LAB – bacteriocins, are generally defined as ribosomally synthesized peptides that have bacteriostatic or bactericidal activity against other related and unrelated microorganisms [128]. These peptides are considered natural biopreservatives and their potential application in the food industry has received great interest [68]. Their main advantage over chemical preservatives is their ability to preserve without affecting the sensory qualities of the food while adhering to the demand for natural preservatives. The ideal bacteriocin should be potent at low concentrations, active against a range of spoilage and pathogenic organisms, innocuous to the host and economical to produce [74; 309]. Bacteriocins in combination with other antimicrobial factors may be useful tools for the implementation of methods intended to significantly reduce the load of food spoilage and foodborne pathogenic bacteria [155]. In our experiment used L. sakei KTU05-6, P. pentosaceus KTU05-10 and P. acidilactici KTU05-7 produced BLIS (sakacin 05-6, pediocin Ac05-7, pediocin 05-8, pediocin 05-9 and pediocin 05-10, respectively) show wide-ranging antimicrobial activities against gram positive and gram negative strains [59]. L. sakei KTU05-6 produced BLIS inhibited both B. subtilis substrains: B. subtilis subsp. subtilis and B. subtilis subsp. spizizenii. Among gram negative bacteria 7 from 13 Pseudomonas spp. strains were inhibited by L. sakei KTU05-6 produced BLIS, whereas other tested pathogenic bacteria strains were not affected by this BLIS. P. acidilactici and P. pentosaceus strains produce BLIS were active against gram positive B. subtilis substrains only, whereas P. pentosaceus KTU05-10 produced BLIS additionally inhibited the P. fluorescens biovar [59].

Currently, the European Union (EU) focuses very intensely on food safety, and especially on both chemical and microbiological hazards. Microbiological hazards are not only pathogenic microorganisms that coincidently find their way into the food chain, but also can be microbial

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cultures deliberately added during food production. Of all 25 EU member states, only Denmark and France have legislation that explicitly regulates the addition of microbial cultures to food, and the EU itself only regulates them in infant formulae and only as to the configuration of the lactic acid molecule [96]. Vice versa, by Food and Drug Administration (FDA, USA) LAB have a GRAS status [12, 53, 98, 109, 138, 275, 294, 296, 405], and lactic acid fermentation has been associated with the reduction of certain naturally occurring or otherwise formed toxins in foods of plant origin. On the other hand, microbially produced lactic acid is usually a mixture of the L(+)- and D(-)- isomers (Figure 1.1.1). As the latter can not be metabolized by humans, excessive intake can result in acidosis, which is a disturbance in the acid-alkali balance in the blood. The potential toxicity of D(-)-lactic acid is of particular concern for malnourished and sick people. Since elevated levels of D(-)-lactic acid is harmful to humans, L(+) lactic acid is the preferred isomer in food industrie as humans have only L-lactate dehydrogenase that metabolizes L(+) lactic acid. The fermentation processes to obtain lactic acid can be classified according to the type of bacteria used. In the heterofermentative process, equimolar amounts of lactic acid, acetic acid, ethanol, and carbon dioxide are formed from hexose, whereas in the homofermentative process only lactic acid is produced from hexose metabolism. Unlike the higher animals and plants which produce exclusively the L(+) isomer, species of LAB produce either D(-)- or L(+)-lactate or even both isomers. The FAO/WHO-experts [161] suggested limiting the daily intake of D(-)-lactate to 100 mg/kg body weight and attempts to favour the L(+) isomer content in fermented food are in progress.

Figure 1.1.1. Chemical structures of L(+) and D(-)-lactic acid isomers. A regulation for reasons of safety should be proportional to perceived

risks, risk being a function not only of severity but also of probability of the adverse effect taking place [402]. The European Food Safety Authority (EFSA) has stated that several LAB strains can be considered to have “Qualified Presumption of Safety” QPS-status [196].

a) L – lactic acid

b) D – lactic acid

OHHO

H CH3

O

OHHO

O

H3C H

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The main aim of lactic acid fermentation is the conversion of carbohydrates to lactic acid. Therefore the action of LAB is desirable for the production of fermented food products. However, some of the bacteria involved in fermentation can produce BAs (Figure 1.1.2). Member States informed the EFSA that findings of certain levels of toxic BAs in fermented food could be of concern and reported a recent increase of BA content in some fermented foods. Formation of BAs in all foods of animal origin having high protein contents, as well in foods of plant origin, has been reported. It can occur as a result of activities of spoilage microflora and/or intentionally added microorganisms. The consumption of food containing higher amounts of toxic BAs may cause food intoxication with symptoms including flushing, headaches, nausea, cardiac palpitations, and increased or decreased blood pressure; in extreme cases the intoxication may have fatal outcome and indicates the need for a better hygiene process and other controls [92].

Figure 1.1.2. Chemical structures of main BAs.

The most frequent food-borne intoxication caused by BAs involves histamine (HIS) and tyramine (TYR) [92, 379]. Histamine causes dilation of peripheral blood vessels, capillaries and arteries, thus resulting in hypotension, flushing, and headache. The toxicological effect depends on histamine intake concentration, presence of other different amines, amino-oxidase activity and the intestinal physiology of the individual [150, 307].

Putrescine

Cadaverine

Spermidine

Spermine

Histamine

Tyramine

β-Phenyltethylamine

Tryptamine

H2NNH2 H2N NH2

H2N NNH2

H H2N NN NH2

H

H

H2N N

NH H2N

OH

H2N

H2N

NH

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For cured products, the FDA consider it a danger to health if the histamine level is equal to 500 mg/kg [206]. Harmful effects resulting from the consumption of foods rich in BAs can be expected only when these amines gain access to the bloodstream [150]. Dietary TYR and trace amines cause vasodilatation of the mesenteric vascular bed, increasing blood flow at the gastrointestinal level and thus facilitating their absorption [43]. Direct effects associated with specific receptors have also been reported at the cardiovascular level, causing an increase in heart rate [110]. The vasoconstriction effect of TYR, phenylethylamine (PHE) and tryptamine (TRP) cause hypertension, but other symptoms such as headache, perspiration, vomiting, pupil dilatation, migraine cases caused by the consumption of food potentially rich in TYR [92]. In this respect, the control of BAs accumulation in food products is one of the present challenges of the food industry [392].

1.2. Design of the fermentation processes – solid state and submerged

fermentation

The demand for faster, more efficient, controllable and largescale fermentation has resulted in a careful selection of starter microorganisms to guarantee the reproducibility of fermentation at industrial scale and to obtain a product with specific properties [48]. The choice of starter culture and fermentation conditions has critical impact on the final quality of fermented foods. Fermentation with well-characterized cultures, yeast or LAB, could be a potential tool to improve the palatability, processability and the nutritional value of fermented products or high-fiber ingredients [335]. The main criteria used to select microbial starters are desirable technological, sensory and nutritional aspects. The main technological factors of interest for fermentation are cells growth and acidification rate [62], synthesis of antimicrobial compounds [169] antifungal activity [63], exopolysaccharide (e.g. glucan and fructan) [55, 116], and sweeteners (e.g., mannitol) production [148].

Technologically interesting potential starter strains are usually selected from the food matrix they are going to be used for [62]. The composition of a fermentation medium influences the supply of nutrients and metabolism of cells in a bioreactor and, therefore, the productivity of a fermentation process also depends on the culture medium used. Of the major culture nutrients, carbon and nitrogen sources generally play a dominant role in fermentation productivity because these nutrients are directly linked with the formation of biomass and metabolites [49]. The understanding and modeling of microbial growth kinetics and transport phenomena play

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important roles in fermentation. The parameters like pH, temperature, agitation and aeration also need to be controlled. Moreover, such understanding is very much required in the design, scale up and process control in fermentation [238].

Fermentation processes may be divided into two systems: submerged fermentation (SMF), which is based on the microorganisms cultivation in a liquid medium containing nutrients, and solid state fermentation (SSF), which consists of the microbial growth and product formation on solid particles in the absence (or near absence) of water, which is similar to the fermentation reaction occurring in nature; however, substrate contains the sufficient moisture to allow the microorganism growth and metabolism [118, 287]. In recent years, SSF has received more interest from researchers since several studies have demonstrated that this process may lead to higher yields and productivities, better product characteristics, less investment, and low energy, water volume and sterility demand than SMF [77]. SSF has become a very attractive alternative to SMF for specific applications [287]. SMF requires the consumption of large amounts of water, energy, and space. SSF, has not gained significant use because of engineering and standardization issues, especially concerning scaling-up the SSF process to an industrial scale process [270]. These biotechnological processes, especially SSF, can be applied to reduce costs and enable the use of enzymes for human and animal consumption [270]. Many microorganisms are capable of growing on solid substrates [118]. In SSF, the water content is quite low and the microorganisms are almost in contact with gaseous oxygen and substrate, unlike in the case of SMF. The water activity levels are very low, the risk of contaminating bacterial or fungal growth is greatly reduced, thereby reducing the high energy cost of strict aseptic and sterile conditions. SSF produces products that are more heat and pH stable at a reduced risk of enzyme inhibition and protease degradation [198].

1.3. Fermented plant products for the food of animal origin nutritional,

technological, and safety parameters improving

LAB improve technological characteristics and the nutritional value of foods during fermentation by increasing the protein content and its digestibility, reducing saccharides content and antinutritional factors (phytates, tannins, and polyphenols), improving the bioavailability of minerals [238] increasing availability of functional compounds (e.g. soluble fiber, soluble arabinoxylans, free phenolic acids, bioactive peptides) [185] and increasing the energy density by hydrolyzing starch into simpler compounds such as glucose and fructose [360]. Incorporation of animal

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origin with plant additives can be a simple way to improve their nutritional value and biofunctionality [142]. Health enhancing ingredients such as plant proteins, dietary fibers, herbs and spices are rapidly increasing worldwide [399]. Plantbased products provide the majority of the carbohydrates, some proteins, oils, dietary fiber (arabinoxylans, β-glucans, cellulose, lignin and lignans), sterols, tocopherols, tocotrienols, ARs, phenolic acids, vitamins and microelements, and have positive health benefits: significantly reduce the risk of some chronic health conditions such as type 2 diabetes, cardiovascular disease, and cancer [14, 54, 209, 300, 344, 349, 382]. The technological processes such as mechanical, thermal, chemical, and biological are used to reduce antinutritional factors content and to improve the bioavailability of nutrients. Unlike thermal, chemical, and mechanical processes which can deteriorate quality of food, fermentation is one of the processes that decreases the level of antinutrients in plant food and increases the starch and protein digestibility, and nutritive value [133, 320]. During fermentation, the plant substrate constituents are modified by the action of both endogenous and bacterial enzymes, including amylases, proteases, hemicellulases and phytases [305], esterases, xylanases, phenoloxidases, thereby affecting their structure, bioactivity, and bioavailability.

Changes of the plants bioactive compounds – ARs, lignans, and VC –

during lactic acid fermentation. Fermentation with LAB positively influences nutritional and functional value of plants: increase the levels of free phenolic acids, TPC, soluble dietary fiber, lignans [168, 229, 300, 308], improve the protein digestibility [16, 93]. Also, LAB are essential for the transformation of natural compounds, e.g., lignans and ARs, to perform their bioactivities. The intake of ARs is beneficial because they reduce the absorption of cholesterol, regulate metabolism of triacylglycerols and affect levels of lipid-soluble vitamins [197; 200]. ARs (Figure 1.3.1) contain phenolic ring and belongs to antioxidants. However, in vitro investigations showed that their antioxidant activity was very low compared to some other bioactive compounds found in cereal grains such as tocols [322]. ARs display also antibacterial and antifungal activities [200; 323]. Because ARs contain hydrophobic alkyl chains, they easily react with proteins including enzymes and thereby inhibit their catalytic activity. Even at low concentrations (e.g. 900 mM), ARs considerably decrease the activities of human digestive enzymes (proteases, aldose reductases and a-glucosidases) and this way they reduce the absorption of some nutrients [323].

19

Figure 1.3.1. The five major cereal ARs homologues have saturated odd-

numbered alkyl chains with 17-25 carbon atoms (C17:0-25:0, top to bottom).

Second group of biological active compounds in plants, which are

increasing during fermentation, is lignans (Figure 1.3.2). Lignans are present in a wide range of foods, such as flaxseed, cereals, vegetable, fruit, and beverages. They afford protection against cardiovascular diseases, hyperlipidemia, breast cancer, colon cancer, prostate cancer, osteoporosis and menopausal syndrome, dependent on the bioactivation of these compounds to enterolactone and enterodiol [113, 211]. They have anticarcinogenic, antioxidant, estrogenic and activities [5, 255].

These phytochemicals are not typical to animal origin and could be an opportunity to industry develop novel food products with enhanced nutritional and health benefits, improved shelf-life, quality and usefull for animal origin functional food production [180, 203].

During the fermentation process VC (acids, alcohols, aldehydes, ketones, and esters) are produced, and they are associated with the sensory characteristics of the fermented products [203]. The SSF showed positive changes of the phenolic content with the development of special flavour compounds, and the influence of SSF on the content of biologically active compounds depended on the type of microorganisms (LAB or yeast) and the used LAB strain [176]. Volatile organic compounds comprise a chemically diverse group of organic compounds, generally with a molecular weight in the range of 50-200 Da, which exhibit appreciable vapor pressure under ambient conditions. For humans, volatiles are important as scents and contribute to the flavor of foods (flavor volatiles). As food aromas, volatiles contribute to palatability and to our appreciation of foods, and along with

OH

HO

OH

HO

OH

HO

OH

HO

OH

HO

OH

HO

20

sugars, organic acids, salts, and other components affecting taste receptors, are responsible for the flavor of food [203].

Figure 1.3.2. Chemical structures of some dietary lignan precursors. Plants and plant products for the food of animal origin quality

improving. Pea fiber. Pea dietary fiber (DF) gives the opportunitie to make innovative, healthy products. DF is the edible part of plants or analogous carbohydrates; it consists of polysaccharides that are resistant to digestion and absorption in the human small intestine with complete or partial fermentation in the large intestine [221]. The physiological effects of DF are dependent on its physicochemical properties, which are mainly influenced by particle size, cell wall structure, solubility, degree of polymerisation and substitution, distribution of side chains and degree of cross-linking of the polymers. Recent results demonstrate the efficacy of fermentation to increase the bioavailability of DF related compounds such as free ferulic acid, lignans and phenolic acids together with other phytochemicals [186, 265]. The important technological characteristics of dietary fiber that determine the possibilities for their application are water holding capacity (WHC), capacity of fat binding, viscosity, gel forming ability, chelating capacity, and the influence on food texture. The WHC is associated with the length and density of the fibers. Also, the pH of the environment affects the water retention capacity. Capacity of fat binding is more dependent on the porosity of the fibers, than the molecular affinity. The ability to form a gel is the most important feature in using fibers as a fat replacer. This ability is

Secoisolariciresinol

Matairesinol

Lariciresinol

Pinoresinol

OH

OHH3CO

HO

OCH3

OH

H3CO

HO

OCH3

OH

O

O

OOH

OCH3

OH

H3CO

HO

OCH3

OH

O

OH3CO

HO

21

provided by cross-linking of polymeric units and by retention of water or other solvents in the gel structure. This characteristic depends on a number of factors, such as concentration, temperature, the presence of certain ions, and the pH of the environment [85]. This is why DF and high-fiber food products have attracted great attention because of their significant health benefits to consumers [14, 57].

Soya. Soybean is one of the main agricultural commodities cultivated worldwide. It is an excellent source for plant protein, oligosaccharides, VB, VE and mineral substance [72]. The health benefits of soybean intake are attributed to various phytochemicals: phenolic acids (mainly vanillic acid, caffeic acid, ferulic acid, protocatechuic acid, and coumaric acid), flavonoids (mainly quercetin and the glycosylated isoforms of isoflavones genistin, daidzin, and glycitin), carotenoids, and tocopherols [90, 205]. They are well known for their antioxidant, antiinflammator, anticarcinogenic activities and they can reduce the risk of cardiovascular diseases [13, 90]. Different from phenolic acids and flavonoids, which are hydrophilic constituents, carotenoids and tocopherols are lipophilic and are found in the oil fraction. Carotenoids possess antioxidant activity due to their provitamin A and RSA, and they present the capability to prevent carcinogenesis, coronary, and neurodegenerative diseases [204, 261]. SSF and SMF are current processing techniques traditionally used to preserve and to enhance the nutritional quality and health promotion properties of legumes [174, 378]. SSF of the soybean is a more economical and simple fermentation technology in order to produce probiotics carrier food [410]. The type of microorganism plays a key role in the fermentation process [174, 378]. Fermentation of soya brings several advantages: decreases the non-nutritional factors (phytates, tannins, trypsin inhibitors and oligosaccharides), improves nutrient digestibility, reduces their allergenicity [111, 318, 364], microbial enzymes bring about the bioconversion of polyphenols into more biologically active compounds [220], reduces the toxins and release many small peptides by the hydrolysis of soybean proteins by microbial proteases [241, 338, 378].

[395] have found that lactic fermentation reduced stachyose and raffinose content and transformed b-glucoside-, acetyl- and malonyl-glucosides isoflavones in soymilk into aglycone, the bioactive form of isoflavones. Furthermore, it has also been found that fermentation enhanced the antioxidant and antimutagenic activity. This is why fermentation with LAB is very usefull on purpose to improve the protein content, supply essential amino acids, reduced the tannin, phytate, trypsin inhibitor and protease inhibitor [398].

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Jerusalem artichockes (Helianthus tuberosus L.) Jerusalem artichoke (JA) is a perennial sunflower species with origins in central North America and has been grown in Europe since the 17th century [370]. Fresh tuber of JA contains 75-80% water, 15-20% carbohydrates (main sources of inulin (70-90%)) and 2% protein [164]. Inulin from JA is a non-starch carbohydrate known as a fructan which is considered as a functional food ingredient with similar characteristics to DF and important technological benefits [78, 326, 395]. Because of its desirable textural and nutritional properties inulin from JA has been used as a prebiotic [326], a source of low glycemic index food [310], foam/emulsion stabilizer and a fat/sugar replacer and texturizer [81] while it promotes further health benefits [240].

JA is a good source of minerals (calcium, iron, selenium, potassium, phosphorus) and vitamins (vitamin B complex, vitamin C and β – carotene) [177], and rich in biologically active substances, including the naturally occurring isomers of caffeoylquinic acid [179], coumarins, unsaturated fatty acids, polyacetylenic derivatives and sesquiterpenes [285], and polyphenols (antioxidants) [334, 343]. The higher antioxidant activity of JA tubers prevent from oxidative stress [299]. Phenolics are secondary plant metabolites found in the majority of herbs, vegetables with well pronounced RSA [121, 247]. The phenolic content exhibits various medicinal properties, such as antioxidant, anticancer, antiallergenic, anti-inflammatory and antiviral [232].

Flaxseed. Flaxseed (Linum usitatissimum L.) accumulates many biologically active compounds and elements including linolenic acid, linoleic acid, phenolic compounds such as lignans, phenolic acids (p-coumaric, ferulic, p-hydroxybenzoic, caffeic, and sinapic acids), and their glucosides, as well as flavonoids (herbacetin and campherol diglucoside), cyclic peptides, polysaccharides, alkaloids, cyanogenic glycosides, and cadmium. Defatted flaxseed contains high levels of dietary fibers and phytochemicals such as lignans [182, 183, 351, 394], are resistant to oxidation [7], and could be incorporated in many types of food as defattening agent [152]. Among the phenolic compounds, flaxseed lignans are in focus because of their estrogenic/antiestrogenic and antioxidant activity [358]. Therefore, flaxseed and their products (whole seed, ground seed and partially defatted flaxseed, which contains the highest content of dietary fiber) are used as a component of functional food [282, 351]. Besides, flaxseed is a good source of soluble and insoluble fibers and has been used as a traditional medicine for centuries to treat constipation.

Savory plants. A long time ago, herbs and spices have been added to different types of food to improve the flavour and organoleptic properties [51]. Consumers increasingly demand natural antimicrobials as alternative

23

preservatives in foods because the safety of additives has been questioned in the last few years. Alternative preservation techniques with such naturally derived ingredients are under investigation for their application in food products. Due to negative consumer perceptions of chemical preservatives, attention is shifting toward alternatives that consumers perceive as natural, especially plant extracts, including the EOs and essences of plant extracts [129]. In this context, they have attracted increasing interest because of their relatively safe status (many of them are considered GRAS by the FDA); they are easily decomposed, environmentally friendly and non phytotoxic [71, 129]. EOs are volatile, natural, complex compounds characterized by a strong odor and are formed by aromatic plants as secondary metabolites [8]. Many spices and herbs exert antimicrobial activity due to their EOs fractions can be used in food system like antifungal, antibacterial and antioxidant agents by inhibiting the growth of pathogenic microorganisms, ensuring the microbiological safety of food products [ 246, 345]. Their inherent antimicrobial activity is commonly related to the chemical structure of their components, the concentration in which they are present, and their interactions, which can affect their bioactive properties. They also may contain various antioxidant compounds such as polyphenols, phenols, flavonoids, etc. which have been thought to be the basis of their antimicrobial properties [71]. Antimicrobials are used in food for two main reasons: (1) to control natural spoilage processes (food preservation), and (2) to prevent/control growth of microorganisms, including pathogenic microorganisms (food safety). There is considerable potential for utilization of natural antimicrobials in food, especially in fresh fruits and vegetables. However, mechanisms of action, as well as the toxicological and sensory effects of natural antimicrobials, are not completely understood [371].

Satureja montana L. and Satureja hortensis L. S. montana L., commonly known as winter savory or mountain savory, belongs to the Lamiaceae family, Nepetoideae subfamily and Mentheae tribe and is a perennial semi-shrub (20–30 cm) that inhabits arid, sunny and rocky regions. S. montana L. is native to the Mediterranean and is found throughout Europe, Russia and Turkey. This is a intensive aromatic herb and has been used for centuries as a spice for food and teas; is used in Mediterranean cooking, mainly as a seasoning for meats and fish and in flavoring agents for soups, sausages, canned meats and spicy sauces [280]. S. montana L. has biological properties that are related to the presence of its major EOs chemical compounds, in general, carvacrol, p-cymen and thymol are main phenolic compounds of savory species oil [167]. S. montana L. have been demonstrated antibacterial, antifungal, antioxidant, anti-diabetes, anti-HIV, anti-hyperlipidemic, reproduction stimulatory, expectorant and

24

vasodilatory activities. Carvacrol and thymol play the fundamental roles in antimicrobial activity of this genus [257, 280, 345]. S. hortensis L. (Lamiaceae), summer savory, is an annual, herbaceous aromatic and medicinal plant native to southern Europe and naturalized in parts of North America. This herb is popular in most regions of the world and are widely used in foods as a flavor component, in herbal teas and possess at a broad spectrum of potent antibacterial and antifungal activities. In folk and traditional medicines this herb is used to treat ailments such as nausea, cramp, indigestion, muscle pain, diarrhea and infectious diseases. Evidence shows that this plant contains phenolic compounds such as thymol and carvacrol with a relatively wide spectrum of antimicrobial activity [311].

Rhaponticum carthamoides CD. R. carthamoides CD. (Willd.) Iljin, a member of the Asteraceae family, is a perennial, herbaceous species naturally growing in the mountains of South Siberia, Middle Asia, and Mongolia [194]. This species often occurs in scientific literature equally under synonyms Leuzea carthamoides (Willd.) DC., preferred primarily in Eastern Europe and post-Soviet countries, or less common Stemmacantha carthamoides (Willd.) [409]. R. carthamoides CD. possess a wide range of biological activities: adaptogenic or anabolic dietary supplement, cardioprotective, immunomodulatory, antihyperglycemic, antioxidant (radical scavenging) and antimicrobial effects. It is valued as a rich natural source of ecdysteroids that are present in all parts of the plant [194]. R. carthamoides CD. are considered to be highly promising in developing new classes of biologically active food additives and ecologically safe products against pests [239] and functional food preparation [44]. Therefore the use of certain aromatic plants as innovative food additives may help to prevent the external growth of fungal spoilers and thus avoid consumer exposure to mycotoxins. In addition, recent research on spices and aromatic herbs suggests that they may be more effective in improving flavour and preserving food than artificial flavourings [225].

1.4. Changes of the meat during lactic acid fermentation process and

the influence of the lactic acid bacteria on smoked meat products safety parameters

Fermentation of meat causes number of physical, biochemical and

microbial changes, which eventually result in functional characteristics of the products. Those changes include acidification (carbohydrate catabolism), solubilization and gelation of myofibrilla and sarcoplasmic proteins, degradation of proteins and lipids, reduction of nitrate into nitrite, formation of nitrosomyoglobin and dehydration [137]. Decrease in pH

25

caused by lactic acid affects the firmness, colour, aroma, texture and flavour development and preservation effect of meat. The hardness of meat is a measure of degree of maturation, resulting from the denaturation and gelation of meat proteins and the loss of water [132]. Fast pH decrease can influence myofibrillar protein functionality, thereby altering meat tenderness, colour, WHC, and meat protein binding ability [175]. Weight loss in meat products is mainly associated with loss in water and WHC of meat. Increasing amounts of released and expressible water are possibly responsible for an increase in weight loss caused by proteolysis and denaturation of proteins during fermentation. Denaturation of sarcoplasmic proteins contributes to the decreased WHC of pork myofibrils [403] and that may cause the increased drip loss [258]. The taste of fermented meat products is mainly due to lactic acids and production of low molecular weights flavor compounds such as peptides and free amino acids, aldehydes, organic acids and amines resulted from proteolysis of meat [264]. It is well known that Lactobacillus species are weakly lipolytic [259]. Lipid oxidation products, free fatty acids, and VC produced from the process of fermentation are responsible for the aroma of a meat product [60, 284]. In most food fermentations, lactic and acetic acids produced by LAB and the resulting decrease in pH are responsible for the preservation effect. In meats, the main organic acid formed is lactic acid, and only low concentrations of acetic acid are acceptable from a sensory point of view [231]. Influence of microorganisms on the change in colour can be associated with acid production, protein denaturation. The production of organic acids is undoubtedly the determining factor on which the shelf life and the safety of the final product depends. The inhibition of pathogenic and spoilage flora is also dependent on a rapid and adequate formation of these organic acids [286]. Meat is an excellent source of protein in human diet and it is highly susceptible to microbial contaminations, which can cause spoilage and food borne infections in human, resulting in economic and health loses [195]. Many factors influencing meat shelf life can promote spoilage, bacterial growth and oxidative processes during storage. These in turn provoke deterioration in the flavour, texture and colour of meat [79]. Biopreservation has gained increasing attention as means of naturally controlling the shelf-life and safety of meat products, where antagonistic microorganisms or their antimicrobial metabolites can prevent the growth of pathogenic bacteria and fungus in food [389]. Consumer demand for greater stringency in relation to hygiene and safety of fresh and processed meat products with natural flavor and taste, free from chemical additives and preservatives [1]. Some microorganisms commonly associated with meat have proved to be antagonistic towards pathogenic and spoilage bacteria

26

[98]. LAB mainly the species Lactobacillus sakei, Lactobacillus curvatus, Staphylococcus carnosus, Staphylococcus xylosus and Staphylococcus saprophyticus are therefore often used as starter and bioprotective cultures in industrial meat fermentation [131]. LAB are essential agents during meat fermentation improving hygienic and sensory quality of the final product. Its fermentative metabolism prevents the development of spoilage and pathogenic microflora by acidification of the product, also contributing to its colour stabilization and texture improvement [98]. pH is one of the most important environmental parameters affecting food fermentation. pH is closely related to microbial growth and the structural changes in phytochemicals during fermentation [367]. Reducing the pH of meat and meat products to 5.0 causes a reduction in lipoxygenase activity. An increase in the enzyme’s oxidative activity was noted for the pH ranging between 6.0–9.0 by [171]. The main cause of the quality defect is denaturation of sarcoplasmic proteins and myosin, leading to a decrease in water-binding capacity of the protein, as a result, a decrease in the WHC of the meat [26]. Low pH and high temperature conditions caused protein denaturation, including denaturation of µ-calpain, and as a consequence, limited post-mortem proteolysis and pale colour of the meat [191]. It is important to be able to predict a high WHC of meat because it is responsible for weight loss in raw, cooked and processed meats. WHC is also responsible for poor colour development in cured meat products, such as ham, and can influence meat palatability traits [328]. During meat processing, one common problem is water loss, which is frequently expressed as drip loss, expressible water, cook loss, and cooling loss depending upon the stage during processing in which it was measured [56]. As a consequence of these processes, the muscle protein coagulates, resulting in the slice ability, firmness and cohesiveness found in the final product. The development of curing colour occurs also in acidic conditions when nitric oxide is produced from nitrite and can then react with myoglobin [210]. The safety of meat and meat products is the most attention problem for consumers. BAs are related to quality and freshness of meat and meat products [281, 354] and has been used as a quality index of unwanted microbial activity [379]. Due to increases in the global demand for foods of animal origin, suppliers are obliged to implement specific controls to guarantee food safety and high quality [379]. BAs are basic nitrogenous compounds present in food and produced by different mechanisms, such as decarboxylation of amino acids or by the normal cellular metabolism of tissues [76, 345].

Biodegradation of toxic compounds by LAB is one of the most important mechanisms for the breakdown of organic compounds and the

27

microorganisms are the most important agents for such degradation. It is known that LAB are able to adsorb some toxic substances (p-cresol, heterocyclic aromatic amines (HAA), ochratoxin A) to their cell wall [2, 271, 272, 302, 377] and remove several toxic compounds [136, 145].

Smoking of meat and meat products is one of the oldest processing methods applied in food preservation. Smoke not only gives special taste, colour and aroma to food but also enhances preservation due to the dehydrating, bactericidal and antioxidant properties of smoke [369]. However, it is known that polycyclic aromatic hydrocarbons (PAHs) can be found in smoked meats and they are a large group of organic compounds, belonging to the food and environmental contaminants [390] (Figure 1.4.1).

Figure 1.4.1. Chemical structures of some policyclic aromatic hydrocarbons.

These contaminants generate considerable interest, because some of them

are highly carcinogenic in laboratory animals and have been implicated in breast, lung, and colon cancers in humans. Dietary intake of PAHs constitutes a major source of exposure in humans [153]. Food can be contaminated by PAHs that are present in air, soil or water, by industrial food processing methods (e.g. heating, drying and smoking processes) and during domestic food preparation (e.g. grilling and roasting processes) [159, 304].

1.5. Lactic acid fermentation in unripened cheese production

LAB are an important group of industrial starter cultures applied in the

production of fermented dairy products [260, 380, 381]. The application of antimicrobial compounds producing LAB in the manufacture of dairy

Benz[a]anthracene

Benzo-[b]fluoranthene

Chrysene

Benzo[a]pyrene

28

products, which can be incorporated into fermented or nonfermented dairy products, implies a processing additional advantage to improve the safety and increase the quality of dairy products, reduce the likelihood of food-borne diseases [19].

Fresh curd cheeses (acid curd cheeses, tvarogs, cottage cheeses) count among traditional dairy products. The technology of their production varies in different regions and different dairy traditions all over the world. For this reason, white fresh cheeses have strongly varied quality characteristics, especially those concerning their chemical composition and sensory characteristics [36]. They are regarded high quality products in our diet, that are rich in protein, macro elements, organic acids and vitamins [368]. Curd cheese are a large and diverse group of fermented dairy products. A common feature of all curd cheese is their processing, which is the coagulation of milk protein (mainly casein) by lactic acid fermentation or an acid-rennet combination (a coagulant enzyme simultaneously in conjunction with the LAB). Curd cheeses have mild, clean, slightly acidic taste and smell. Their structure and texture are uniform, compact, without lumps, and slightly loose, and it may be slightly granular. The colour of curd cheeses should be white to light cream and be uniform throughout the whole cheese [412].The manufacture of dairy foods is not a sterile process, and BA producers are likely to enter the food chain as non-starter LAB that are indigenous to the raw material. The presence of BAs in nonfermented foods generally indicates inadequate or prolonged storage; on the other hand, their presence in fermented foods could be unavoidable due to the diffusion of decarboxylases LAB [362]. BA formation is only possible if there is availability of the free substrate amino acids and the environment conditions are favorable to the decarboxylation activity [330]. Formation of BA in cheese depends on various factors; such as ripening time, ripening temperature, pH, the presence of microorganisms having BA-producing capability through their proteolytic and decarboxylase “activities” [226] and the bacterial density and synergistic effect between microorganisms are the most important [363].

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2. MATERIALS AND METHODS

2.1. Investigation venue

The experiments were conducted between 2012 and 2016 at the Lithuanian University of Health Sciences Veterinary Academy (LUHS VA) Department of Food Safety and Quality; Institute of Animal Rearing Technologies Laboratory of Meat Characteristics and Quality Assessment (LUHS VA), Kaunas University of Technology (KTU) Department of Food Science and Technology; Vytautas Magnus University (VMU) Department of Biology / Environmental Research Centre; Kaunas Botanical Garden (VMU); University of Latvia (LU) Centre of Food Chemistry (Riga, Latvia); Institute of Food Safety, Animal Health and Environment – “BIOR” (Riga, Latvia); at the enterprises “Nematekas” (Dovainonys, Lithuania), and “Judex” (Kaunas, Lithuania).

2.2. Materials

Microorganisms used in experiments. Lactobacillus sakei KTU05-6,

Pediococcus acidilactici KTU05-7, Pediococcus pentosaceus KTU05-8 and Pediococcus pentosaceus KTU05-9 previously isolated from spontaneous rye sourdough and selected due to their preliminary inhibiting properties [82] were obtained from the culture collection of the Kaunas University of Technology, Department of Food Science and Technology, cereal and cereal products research group collection (Kaunas, Lithuania).

Bacillus cereus ATCC 10876, Bacillus subtilis, Escherichia coli 1.10, Escherichia coli ATCC25922, Escherichia coli, Listeria monocytogenes 1.1, Pseudomonas fluorescens biovar. V and Pseudomonas fluorescens biovar. III were obtained from the Institute of Botany of the Nature Research Centre (Vilnius, Lithuania).

Pseudomonas aeruginosa NCTC 6570, Pseudomonas aeruginosa VUL-13, Staphylococcus aureus ATCC 9144, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 8739, Listeria monocytogenes ATCC 35152, Salmonella enterica serovar typhimurium ATCC 13311, Bacillus cereus ATCC 11778, and Yersinia enterolitica DSM 13030 and Yersinia pseudotuberculosis III HH 146-36/84, previously isolated from the pork production chain, were obtained from the LUHS VA Department of Food Safety and Quality collection (Kaunas, Lithuania).

30

Plant material. In experiment used plant material is presented in Table 2.2.1.

Defatted soy flour (Maxima, Kaunas, Lithuania) and pea fiber (M plant Ltd., Hamburg, Germany) were purchased at a local pharmacy (Kaunas, Lithuania).

Winter savory (Sm), Summer savory (Sh) and Maral root or Rhaponticum carthamoides (Rc) were grown and collected in an experimental field located in the Kaunas Botanical Garden of VMU in 2012.

Savory plants (Sm ans Sh) were used for RCMP production and for the beef and pork meat treatment.

Table 2.2.1. In experiment used plant material.

The main characteristic of selected plant

Plant

With the high content of proteins Defatted soy flour With the high content of phenolic compounds

Winter savory (Satureja montana L.) (Sm), Summer savory (Satureja hortensis L.) (Sh), Maral root or Rhaponticum (Rhaponticum carthamoides CD.) (Rc)

With the high content of inulin Jerusalem artichokes (Helianthus tuberosus L.) With the high content of lignans Defatted flaxseed (Linum usitatissimum L.) With the high content of dietary fiber

Pea fiber

Rc products were used for unripened curd cheese (UCC) production. Defatted flaxseed (Institut Wlokien Naturalnych, Poznan, Poland) were

purchased at a local super-market (Kaunas, Lithuania). Defatted flaxseeds were used for the production of functional additives with a high content of lignans production.

Jerusalem artichoke (Helianthus tuberosus L.) (harvest of 2011) was received from the Lithuanian Institute of Horticulture Experimental Farm (Babtai, Lithuania). Jerusalem artichoke tubers were used for ready-to-cook minced pork meat products (RCMP) enrichement [365].

Pea fiber was used for gluten-free RCMP production. Material of animal origin. Fresh pork and beef meat from five different

muscles including neck, shoulder, ham, M. longissimus dorsi and loin were obtained from the local market (Kaunas, Lithuania) (less than 4 days after slaughter). Pork and beef muscles were specifically chosen to span as large range of the concentrations of protein, moisture and fat as possible. All meat samples were cut into chops with a thickness of 2.5 cm, placed into plastic containers and stored under refrigeration (+4 ± 1 °C) for 12 hours and then

31

were used for marination with selected LAB-based marinades, and Sm and Sh bioproducts were used for fresh pork and beef loin treatment.

Fresh, raw, boneless chicken breast, drumsticks and thigh muscles were obtained from the local market (Kaunas, Lithuania) and from an ecological farm (Kaunas, Lithuania).

Fresh pork loin (used for minced meat production) was obtained from the local market (Kaunas, Lithuania) (less than 4 days after slaughter). Loin was cut into chops with a thickness of 2.5 cm, placed into plastic containers and stored under refrigeration (+4 ± 1 °C) for 12 hours. Fresh pork loin minced meat was used for RCMP production.

Fresh pork and frozen back fat (used for sausage production) was obtained from the local market (Kaunas, Lithuania), minced and used for sausage production.

Raw cow milk was collected from a local farm (Mazeikiai, Lithuania). Before cheese production, the raw cow milk was stored in a refrigerator at +4 ± 1 °C no longer than 12 hours. Raw cow milk was used for UCC production.

2.3. Lactic acid bacteria cultivation and plant material fermentation

LAB cultivation. LAB (L. sakei; P. acidilactici; P. pentosaceus 8,

P. pentosaceus 9) before the experiment had been stored at -80 °C (PRO-LAB Diagnostics, United Kingdom) supplemented with 20% of glycerol. Before the experiment, LAB had been defrosted and propagated in de Man Rogosa Sharpe (MRS) broth (CM 0359, Oxoid Ltd, Hampshire, United Kingdom): L. sakei at 30 °C, P. acidilactici at 32 °C, P. pentosaceus 8, P. pentosaceus 9 at 35 °C temperature by keeping for 48 hours in a thermostat (Binder, Germany). Before the use, 40 mM of fructose and 20 mM of maltose had been added.

Plant bioproducts production. The fermentation of plant products was

performed with pure P. acidilactici, L. sakei, and P. pentosaceus strains (2%, m/m). The substrate water content was calculated with reference to the moisture content of the raw materials, water absorption capacity and the required humidity of the end product for SSF was ≤ 50%, and for SMF was ≥ 70%. The fermentation was carried out for 48 hours at optimal temperatures for P. acidilactici, L. sakei and P. pentosaceus (30 °C; 32 °C and 35 °C, respectively).

Fermentation of defatted soy flour and pea fiber was carried out for 48 hours at optimal temperatures for LAB cultivation, under SSF and SMF conditions.

32

The upper parts of Sm, Sh and Rc were collected during the flowering vegetative phase. The plants were dried in a vacuum oven (Model SZG, China) until the moisture content was approximately 7%, and stored in the dark at the ambient temperature. Before starting the experiment, the plants had been ground in a laboratory-scale impact mill (Bühler-Miag, Brunswick, Germany) into particles with a size of 0.5–2.0 mm. The ground savory plant samples were weighed in glass beakers covered with the aluminium foil. The samples were autoclaved at the temperature of 121 °C for 10 min and soaked in sterile water. Two types of plant bioproducts were prepared: I – bioproducts containing 50% of water (SSF); and II – bioproducts containing 70% of water (SMF). 2% of a fresh LAB culture suspension was added and mixed under aseptic conditions. The prepared plant samples were fermented for 48 hours at the optimal temperature (described above).

Partially defatted flaxseed (Linum usitatissimum L.) flour was fermented for 48 hours at optimal temperatures for LAB cultivation, using SSF and SMF conditions.

Tubers of Helianthus tuberosus L. were cut into 1–2 mm slices, dried in a vacuum oven (Model SZG, China) at +45 °C and ground into powder. Fermentation was carried out for 48 hours at optimal temperatures for LAB cultivation, using SSF and SMF conditions.

2.4. Methods of evaluating plant bioproducts microbiological,

physical chemical, enzymatical and antimicrobial activity parameters

Microbiological parameters evaluation methods of plant bioproduct.

LAB counts were determined on MRS agar (Liofilchem, Roseto degli Abruzzi, Teramo, Italy) using standard plate count techniques. The plates were incubated at a temperature of 30 °C, 32 °C and 35 °C for 72 hours under anaerobic conditions (using the atmosphere generation system AnaeroGen, Oxoid, Basingstoke, UK).

The number of spore-forming aerobic mesophilic bacteria was determined on Plate Count Agar (CM0325, Oxoid, UK) after incubation at a temperature of 30 °C for 72 hours. Before their inoculation on agar plates, the diluted samples had been heated at a temperature of 80 °C for 10 min to remove the vegetative bacteria cells and to evaluate the amount of spores.

Enterobacteria were determined on the Violet Red Bile Glucose Agar (Liofilchem, Italy) after the incubation at a temperature of 37 °C for 24 hours.

33

The yeasts and fungi were determined on the Rose-Bengal Chloramphenicol Agar (CM0549, Oxoid, UK) after a five-day incubation at a temperature of 25 °C.

All experiments were carried out in triplicate, and the number of microorganisms was expressed as log10 of colony-forming units per gram (log10 CFU/g).

The evaluation of the antimicrobial activity of tested LAB/plant extracts against foodborn pathogens is described below.

Methods of evaluating antimicrobial activity of plant bioproduct and

LAB supernatants. EOs from Sm, Sh and Rc and their bioproducts were extracted using the supercritical carbon dioxide (99.9% purity, AGA) extraction. For supercritical fluid extraction, an HP 7680 T (Hewlett Packard, Palo Alto, CA, USA) apparatus was used. The extraction according to [176] was carried out. The antimicrobial activity of EOs extracted from plantbased bioproducts and nonfermented savory plants by applying an agar diffusion assay method were tested. The LAB propagated in MRS media for 24 hours were centrifuged (4 °C, 10 min, 6000g) to remove LAB cells and obtained LAB supernatants for antimicrobial activity determination were used. The indicator strains (Bacillus cereus ATCC 10876, Bacillus subtilis, Escherichia coli 1.10, Escherichia coli ATCC25922, Escherichia coli, Listeria monocytogenes 1.1, Pseudomonas fluorescens biovar. V and Pseudomonas fluorescens biovar. III) for antimicrobial activity determination in a nutritional medium (per 1 L used: 5 g peptone, 1.5 g meat and yeast extract and 5 g NaCl) at 37 °C were propagated. The antimicrobial activities against the indicator microorganisms by measuring the inhibition zones (mm) were determined.

Methods of evaluating plant bioproduct moisture and acidity parameters. The determination of the moisture required samples to be dried at a (103 ± 2)°C temperature in a thermostat (UNB400, Memmert, Germany) till the constant mass, and the percentage of the mass change was calculated.

The pH value was measured and recorded by a pH electrode (PP – 15, Sartorius, Goettingen, Germany).

The total titratable acidity (TTA) was expressed in mL of 0.1 M NaOH solution per 10 g of a sample to obtain pH = 8.5. The TTA was assessed in Neiman degrees (° N).

The concentrations of L(+)- and D(-)-lactic acid were determined with an enzyme test kit as reported elsewhere [75, 165]. L(+)-lactic acid was determined using an enzymatic test kit K-DLATE 08/11 (Megazyme

34

International Ireland Limited), and D(−)-lactic acid was also determined using the same test kit modified by replacing L(+)-lactate dehydrogenase of D(−)-lactate dehydrogenase (from Leuconostoc mesenteroides, Sigma). The enzyme L(+)- or D(−)-lactate dehydrogenase catalyzed the oxidation of L(+)- or D(−)-lactate in the presence of nicotinamide adenine dinucleotide (NAD+), and the product, pyruvate, was trapped by hydrazine while the NADH formed was quantified by measuring the absorbance at 340 nm.

Methods of TPC, β-glucans, ARs and lignans content evaluation in

plant bioproducts. The analysis of the ARs, lignans and RSA were carried out according to the procedures described by [30].

TPC content was carried out according to the procedures developed by [387].

Total β-glucans content was determined by the method of [243], in which a specific (1→3)(1→4)-β-D-glucan-4- glucanohydrolase (EC 3.2.1.73; lichenase) and β-glucosidase (EC 3.2.1.21) were used to hydrolyze β-glucan to glucose using an assay kit (Megazyme International, Bray, Ireland).

BAs evaluation procedure. Extraction and determination of BAs was

carried out according to the procedures developed by [33]. Perchloric acid (0.4 mol/L, 10 mL) containing a known amount of 1.7-diamino-heptane used as an internal standard was added to 3 g of sample, and the mixture was homogenized with Ultra-Turrax (IKA Labortechnik, Staufen, Germany) and centrifuged at 3000 × g for 10 min. The residue was extracted again with an equal volume of 0.4 mol/L perchloric acid.

Both supernatants were combined, and the final volume was adjusted to 30 mL with 0.4 mol/L of perchloric acid. The extract was 6 filtered through Whatman No. 1 paper. One millilitre of extract or standard solution was mixed with 200 mL of 2 mol/L sodium hydroxide and 300 mL of saturated sodium bicarbonate. A 5-(dimethylamino)naphtha-lene-1-sulphonyl chloride (dansyl chloride reagent) (10 mg/mL, 2 mL) prepared in acetone was added to the mixture and incubated at 40 °C for 45 min. Residual dansyl chloride was removed by the addition of 100 mL of 25 mg.L ammonium hydroxide. After incubation at room temperature for 30 min, the mixture was adjusted to 5 mL with acetonitrile. Finally, the mixture was centrifuged at 3000 × g for 5 min, after the supernatant was filtered through 0.2 µm filters (Millipore Co. Bedford, MA, USA) and stored at 225°C until HPLC analysis. An Agilent 1200 HPLC (Carlsbad, CA, USA) equipped with diode-array detector and Chemstation LC software was employed. A Chromolith C18 HPLC column (100 mm × 4.6 mm × 4 mm, Merck KGaA/EMD Chemicals, Darmstadt, Germany) was used. Ammonium

35

acetate (0.1 mol/L) and acetonitrile were used as the mobile phases by a flow rate of 0.45 mL/min. The sample volume injected was 10 mL and the amines were monitored by 254 nm. The detection limits for standard amine solutions were approximately 0.1 mg/kg.

Methods of evaluating LAB excreated enzymes activity. Amylolytic

enzymes activity in fermented plant bioproducts was determined according [268].

Proteolytic enzymes activity in fermented plant products was determined according procedures developed by [69].

2.5. Technology of food products

In the experiment, marinated pork, beef, organically and conventionally

produced chicken meat, minced meat products, cold smoked pork meat sausages and UCC were produced.

2.5.1. The use of fermented Sm and Sh bioproducts for beef and pork

loin marination. Fresh pork and beef loin was divided into three subsamples (each 100 g). Then the meat surface was treated with 2% (m/m) of Sm and Sh bioproducts, placed into plastic containers and stored under refrigeration (+4 ± 1°C) for 24 hours. Meat samples without Sm and Sh bioproducts were stored under refrigeration (+4 ± 1°C) for 24 hours and analysed as a control.

2.5.2. The use of LAB – potato juice marinade for meat treatment.

The parameters of natural marinade (based on potato juice fermented with different LAB) are given in Table 2.5.2.1. For all analyses, samples were collected 12 hours after the manufacturing process. Untreated chicken meat samples were analysed as a control.

Pork and beef meat marination. Fresh pork and beef meat from five different muscles (neck, shoulder, ham, M. longissimus dorsi and loin) were divided into three subsamples (each 100 g). Meat was placed individually in a container with 100 mL of natural marinade based on LAB, and coveredg with a plastic film. The meat subsamples were stored for 24 hours at a temperature of +4 ± 1 °C in a refrigerator. Untreated pork and beef meat was analysed as a control.

Organically and conventionally produced chicken meat marination. Organically and conventionally produced chicken meat (120 ± 10 g) was cut into three subsamples 40 g each. Chicken meat surface was sprayed with

36

natural marinade based on LAB, placed individually in a container, covered with a plastic film and stored in a refrigerator at +4 ± 1 °C for 12 hours.

Table 2.5.2.1. The pH, TTA and LAB cell count in fermented (72 hours) with different LAB potato juice marinade.

Time Potato juice

pH TTA, °N LAB count (log10

CFU/mL) Pa Pp Ls Pa Pp Ls Pa Pp Ls

72 4.1± 0.04

4.15±0.07

4.24±0.08

8.92± 0.16

8.87± 0.13

8.53± 0.12

9.74± 0.15

9.60± 0.17

9.41± 0.11

Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; TTA – total titratable acidity; h – hours. Values are mean±standard deviation of three replicate analyses. Different letters indicate significant differences between mean values of treatments (P < 0.05).

2.5.3. Production of cold smoked pork sausages.

The cold smoked pork sausages production was performed in the

agricultural enterprises “Nematekas” (Dovainonys, Lithuania). Sausages were made of 76.9% fresh pork, 19.4% frozen back fat, 2.4% salt (containing 0.4% sodium nitrite (NaNO2)), 0.4% glucose, and 0.4% spice mix. The diagram of sausages preparation, additional biotreatment with LAB and sampling schema is presented in Figure 2.5.3.1. Control sample was prepared without treatment by LAB suspension.

Meat and fat was ground. After grinding, the batch was mixed 2 min in a mixer to distribute the added fat and spice mix. The mixture was vacuum-stuffed into natural casing (40 mm diameter, 240 mm length). Sausages treatment with LAB have been performed before (I) and after (II) smoking. (I) Formed fresh pork meat sausages, 400 g of each, were placed individually in a container with 1000 mL of fermented potatoes juice (containing on average 9.60 log10 cfu m/L of LAB) and were immersed for 60 min at 18 oC.

After immersion the sausages were drained and covered with plastic film. Samples were stored at room temperature (18-20 °C) for 24 hours. After 24 hours sausages were ripened for 78 hours in 24 °C temperature under 93-86% humidity. After ripening fresh pork meat sausages were smoked at 16 °C temperature for 130 min under 80–82% humidity. After smoking sausages were dried (8 days at 15 °C under 75% humidity). The smoking and drying was carried out in the universal thermal camera (Bastramat 850 C-UP, Armsberg, Germany) with separate sawdust smoke generator. (II) The sausages after smoking and drying (400 g of each) were placed individually in a container with 1000 mL fermented potatoes juice

37

(containing on average 9.60 log10 CFU µg of LAB) and covered with plastic film. Samples were stored at room temperature (18-20 °C) for 24 hours.

Figure 2.5.3.1. The diagram of sausages preparation, additional bio-treatment with LAB and sampling.

2.5.4. Ready-to-cook minced pork meat products technology

For the production of RCMP, meat slices were minced twice (4 mm

diameter of holes in a plate) and after addition of 3, 5 and 7% of Sm and Sh bioproducts carefully mixed to obtain a homogeneous sample.

Also, minced meat samples with the addition of 5% of pea fiber (instead of semolina) were produced. RCMP produced with 5% of semolina as a

38

control were analysed. Prepared samples were divided into subsamples (50 g each) and after covering with a plastic film were stored under refrigeration (+4 ± 1 °C) for 5 days. For all analyses the samples were collected 12 hours after the manufacturing process.

2.5.5. Unripened curd cheese technology

The curd cheese was made from 10 L of raw cow milk. The raw milk,

intended for the production of UCC (fresh cheese), was pasteurised at 72–73 °C for 15–20 s and then cooled to 30 ± 2 °C. Next, 3% w/vol of Sm and Rc bioproducts were added. After spontaneous coagulation of the milk, the curd was mixed and gently cut into cubes of 100 g. Then the cubes of curd were drained, placed in nylon containers, pressed for 12 hours and held in 4 ± 1°C. In addition, curd cheese samples with nonfermented plants were prepared. Samples of curd cheese without fermented bioproducts were analysed as control samples.

2.6. Methods of evaluating food products microbiological, physical chemical, sensory and technological parameters

For marinated meat pH, moisture content, cooking loss, tenderness, drip

loss, intramuscular fat (IF), WHC, colour characteristics, BAs content and overall acceptability were evaluated.

For cold smoked pork sausages BAs content, PAHs and overall acceptability were evaluated.

For RCMP the number of spore-forming aerobic mesophilic bacteria, Enterobacteria, fungi and yeasts, pH, technological parameters (moisture content, cooking loss, dry matter content, tenderness, drip loss, IF, WHC, colour characteristics, BAs content, VC, and overall acceptability were evaluated.

For UCC anti-mold activity of plant bioproducts on the curd cheese surface, TTA, pH, L(+) and D(-)-lactic acid, BAs, VC were evaluated and sensory analysis was performed.

Microbiological analysis methods of food products. The number of

spore-forming aerobic mesophilic bacteria, Enterobacteria and yeasts and fungi was determined according procedures described at 2.4. paragraph.

Anti-mold activity of plant bioproducts on the unripened curd cheese surface: L. sakei bioproducts, prepared under SSF conditions, were tested for anti-mold activities on the unripened curd cheese surface during cheese storage. The prepared fresh curd cheese was covered with paper and stored

39

in the fridge at the temperature of +4 °C. The fresh curd cheeses, made using dried plants without adding any bioproduct, were used as control samples. The cheese samples were examined externally for 13 days to check the levels of mold appearing on the surface of the cheese according to the following scale: (-) no mold; (+) the first traces of mold; (++) large mold colonies.

Methods of evaluating food products acidity parameters. The TTA, pH was evaluated according procedures described in 2.4. chapter.

Methods of evaluating meat products technological parameters. The moisture content was determined according to the AOAC 950.46

method [18]. Cooking loss was determined after cooking of 100 g sample in a 100 oC

temperature water for 15 min and expressed as a percentage of the initial weight.

Dry matter content was determined by water loss during heating in an oven (UNB400, Memmert, Germany) at 102-105 °C temperature until weight constancy

Tenderness of meat samples was determined using Warner – Bratzler device, which measured as shear force, needed to cut meat sample with specifical knife.

Drip loss was measured as fluid lost from fresh meat via passive exudation (+4 oC; 24 h) and expressed as a percentage of the initial weight of the product according [105].

The amount of IF was measured by the Soxhlet method with petroleum ether as solvent according [107].

WHC was determined with the filter paper method according to [127] and expressed as percent of loose water in meat. A meat sample was compressed between 2 layers of filter paper and 2 plaques of acyclic Plexiglas at a force of 5 N for 60 s using the compression technique (Lloyd Instruments Ltd., Hampshire, UK).

Colour characteristics evaluation method. The colour characteristics were evaluated at three different positions of the surface using CIELab system (CromaMeter CR-400, Conica Minolta, Japan). L* is a measure of lightness, from completely opaque (0) to completely white (100), a* is a measure of redness (or -a* of greenness), and b* of yellowness (or -b* of blueness) [244].

Sensory analysis. The overall acceptability of meat products was evaluated according to the [162] by fifteen judges for overall acceptability using a 6 scores hedonic line scale ranging from 6 (extremely like) to 1 (extremely dislike).

40

Sensory analysis of curd cheese was carried out by ten assessors according to the [162].

Also, the acceptability of cheese samples was evaluated by applying FaceReader software (Noldus Information Technology, Wageningen, the Netherlands) and using a scale of the 3 basic emotions (happy, sad and angry).

Methods of evaluating food products safety parameters. The influence of fermented plant bioproducts on BAs and D(-) lactic acid (methods described in chapters 2.4.5 and 2.4.3, respectively) and PAH content in food products was evaluated.

Method of PAH determination in smoked pork meat sausages. Solvents employed were cyclohexane, hexane, dichloromethane, and ethyl acetate, all of them were of pesticide purity grade. Other reagents and materials used were anhydrous sodium sulphate, and 6 mL (500 mg) Phenomenex Strata SI-1 Silica solidphase extraction (SPE) tubes. All the aforementioned solvents, reagents, and materials were commercially purchased from SigmaeAldrich (Steinheim, Germany), Supelco (Bellefonte, PA), and Merck (Darmstadt, Germany). Mixture of four PAHs standards: benz[a]anthracene (BaA), benzo-[b]fluoranthene (BbF), benzo[a]pyrene (BaP), chrysene (Chr), and deuterated standards benzo[a]pyrene-d12, benzo[b]fluoranthened12, chrysene-d12, and benz[a]anthracene-d12 were purchased from Dr. Ehrenstrofer (Augsburg, Germany). The standard mix of PAHs consisted of a solution in acetonitrile with concentration 50 mg/L and the concentration of deuterated benzo[a]-pyrene-d12, benzo-[b]fluoranthene-d12, chrysene-d12, benz[a]anthracene-d12 dissolved in cyclohexane was 1000 ng/μl. The mixtures were stored at 4 °C.

For analysis whole samples were homogenized, minced into smaller pieces, and blended. Each homogenised smoked pork meat sausage sample (2.75 g) was thoroughly mixed with 15 g of anhydrous sodium sulphate to absorb moisture. An aliquot of 27.5 μL toluene solution of internal standards including BaP, Chr, BbF, and BaA with concentration 0.500 μg/mL was added. The PAHs were extracted from smoked sausages samples by adding 25 mL of dichlormethane/hexane (1:1, v/v) mixture and performing sonication of about 20 min. After sonication the supernatant of the extracts were decanted and 15 mL of fresh solvent was added for another 20 min of sonication. To avoid the presence of solid particles, all the extracts were filtered. The combined extracts (~40 mL) were rotary evaporated (30 °C, 500-100 mbar) to eliminate the solvents, and the fat residue was reconstituted in 5.5 mL of cyclohexane/ethyl acetate (1:1, v/v) solution for further elimination of high molecular compounds by means of gel permeation chromatography (GPC). The extracts were centrifuged at

41

3000 rpm for 10 min and the solution was transferred into a glass GPC vial. The sample extracts were injected into LC Tech Freestyle™ GPC system (Dorfen, Germany) consisting of an HPLC pump, autosampler, and fraction collector. High molecular substances were removed on a glass column (500×40 mm, 25 mm ID) filled with 50 g of Bio-Beads SX3 (Bio-Rad, Philadelphia, USA) stationary phase with cyclohexane/ ethyl acetate (1:1, v/v) mobile phase at a flow rate of 5 mL/min. The automated GPC programwas as follows: dump time 0-21 min, collection time 21-45 min. The collected fractions were transferred to round-bottom flasks and then rotary evaporated (30 °C, 130 mbar) to dryness. The dry residue was dissolved in 3 mL of cyclohexane. Further clean-up was done using SPE cartridges filled with 500 mg of silica. The sorbent of the SPE cartridges was first conditioned with 5 mL of cyclohexane, after which extracts were loaded onto the cartridges. The analytes of interest were eluted from the column with cyclohexane (3×3 mL), the obtained fraction was evaporated under a stream of nitrogen at 40 °C, dissolved in 50 μL of cyclohexane, and transferred into a GC-MS/MS vial for the further PAHs analysis.

The PAHs analysis was carried out by ThermoScientific TSQ Quantum XLS Ultra GCeMS/MS system equipped with a DB-17 capillary column (30 m long ×0.25 mm i.d. ×0.25 μm film thickness) and operating in a splitless mode. The operating conditions were as follows: helium gas was used as the carrier gas at a constant flowof 1.2 mL/min; inlet temperature 260 °C; MS transfer line temperature 280 °C; source temperature 250 °C. The oven temperature was set initially at 80 °C (2 min hold), increased to 265 °C at 15 °C /min. At 265 °C, temperature increased at a rate of 5 °C /min to 290 °C and then to 320 °C at a rate of 20 °C /min (20 min hold). The total run time was 45.8 min. The injection volume was 1 μL. The data were acquired by operating the MS in selective reaction monitoring mode (SRM).

Method of evaluating volatile compounds. Samples for gas chromatographic analysis were prepared using solid-phase microextraction (SPME). SPME device with Supelco 57750-U StableflexTM fibre coated with 65-mm PDMS-DVB layer (Sigma-Aldrich, St. Louis, MO, USA) was used for the preparation of samples. For extraction, 0.01 g of sample was placed in a 10-mL glass vial with the PTFE-lined silicone septa. The sample was placed in the headspace of the sample at 25 °C for 1 hours. The injection was performed by thermal desorption of the volatiles in the injection port of the gas chromatograph at 230 °C. For GC-MS measurements, a model GCMS-QP2010 gas chromatograph with a mass spectrometric detector (Shimadzu, Tokyo, Japan) was used. The ionization of the analytes was performed using an electron ionization mode at 70 eV. For the separation of volatiles, a low polarity Rtx®-5MS column (Restek

42

Corporation, Bellefonte, PA, USA; length 30 m, coating thickness 0.25 mm, 0.25 mm i.d.) was used. Ion source temperature was set at 220 °C, and interface temperature was 260 °C. Sample injection was carried out for 1 min in order to ensure full desorption of volatiles from the SPME fibre. Split mode injection (1:10) was used. Temperature gradient program was set as follows: from 30 to 200 °C at 5 °C/min and up to 280 °C at 20 °C/min, then maintained for 2 min. The carrier gas was 99.999% helium (AGA, Vilnius, Lithuania) with a pressure of 90 kPa at the column head, and the column flow of 1.61 mL/min. The compounds were identified according to the mass spectral library NIST v. 8.0 (The National Institute of Standards and Technology, Gaithersburg, MD, USA). Five identical samples were repeatedly measured for solid-phase microextraction coupled with GC-MC. The relative standard deviation (RSD) for peak area did not exceed 5.15% [176].

2.7. Statistical analysis

All analytical experiments were carried out in triplicate. In order to evaluate the influence of three different factors (fermentation method, plant bioproduct, and the application of several microorganisms) and their interaction on the quality parameters of the final product, the data were subjected to the analysis of variance (ANOVA) and the Tukey HDS test as a post-hoc test (statistical program R3.2.1, Core Team 2015). The results were referred to be statistically significant at p ≤ 0.05.

43

3. RESULTS

3.1. Microbiological and physical chemical parameters of plants and their bioproducts

3.1.1. Parameters of fermented and nonfermented savory plants

The moisture content, total TTA, L(+) and D(-)-lactic acid isomers

content and the pH of SMF and SSF fermented savory plants are presented in Figure 3.1.1.1. Moisture content in SMF samples ranged from 69.53 ± 0.95% (in SMF with P. acidilactici Sm) to 80.14 ± 1.32% (in SMF with P. pentosaceus Sh) and in SSF samples from 38.24 ± 0.67% (in SSF with P. acidilactici Sm) to 44.63 ± 1.13% (in SSF with P. acidilactici Sh). In all the cases, a higher TTA was found after SMF of savory plants as compared with SSF (except SMF with P. pentosaceus Sm) and ranged from 10.3 ± 0.27 ºN to 14.3 ± 0.69 ºN (in SSF with L. sakei Sm and in SMF with P. pentosaceus Sh, respectively). Results of the ANOVA test indicated that the TTA was significantly affected by the fermentation method (p ≤ 0.0001) and the type of the savory plant (p ≤ 0.0001). The interaction of factors (fermentation method, LAB used for fermentation, and the type of the savory plant) has a significant influence on the TTA of savory plants (p ≤ 0.003). A moderate significant correlation between the substrate moisture content and TTA (r = 0.577; p = 0.0001) was found.

In all the cases, a higher sample pH after SSF of savory plants was found, and it ranged from 4.41 ± 0.05 to 5.93 ± 0.07 (in SMF with P. acidilactici Sh and in SSF with P. acidilactici Sm, respectively). After 48 hours of fermentation, the pH of samples was significantly influenced by the fermentation method (p ≤ 0.0001) and by the type of the savory plant (p ≤ 0.0001). The interaction of factors (fermentation method, LAB used for fermentation, and the type of savory plant) was significant on the pH of savory plants (p ≤ 0.0001). A moderate negative significant correlation between the fermentable substrate moisture content and pH (r = -0.790; p = 0.0001) and a moderate positive significant correlation between the pH and TTA of the fermentable substrate (r = 0.565; p = 0.0001) were found.

The ratio of L(+)/D(-)-lactic acid isomers in Sm, Sh and Rc ranged from 1.2 to 2.1, from 1.3 to 2.0, and from 0.08 to 2.2, respectively, with the L(+)-lactic acid as the predominant isomer. The content of the L(+) and D(-)-lactic acid isomers was significantly influenced by the fermentation method (p ≤ 0.0001) and by the type of the savory plant (p ≤ 0.0001). The interaction of the factors (fermentation method and the type of a savory

44

plant) have a significant influence on the content of the L(+) and D(-) isomers in savory plants (p ≤ 0.0001).

a)

b)

c)

d)

Figure 3.1.1.1. Moisture content (%) (a), TTA (°N) (b), pH (c), and L-(+) and D-(-) lactic acid isomers content (g/100 g) (d) of SMF and SSF

fermented savory plants (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus; SMF – submerged fermentation; SSF – solid state fermentation, TTA – total

titratable acidity).

The results of the microbiological parameters of SMF and SSF savory plants are presented in Figure 3.1.1.2. Savory plants were a suitable substrate for LAB cultivation, and the highest content of valuable LAB in SSF with P. acidilactici Sh (9.74 ± 0.10 log10 CFU/g) was found. The LAB count in fermented plants was significantly influenced by the fermentation method (p ≤ 0.0001). A moderate negative significant correlation between LAB count and moisture content (r = -0.464; p = 0.0001) and between LAB count and TTA (r = -0.496; p = 0.0001) of the substrate was found.

The count of spores of aerobic mesophilic bacteria in SMF and SSF savory plants ranged from 0 to 2.12 ± 0.02 log10 CFU/g and in comparison with nonfermented samples, was found to be from 3.98% to 2.86% lower (in

SMF

SSF

SMF

SSF

SMF

SSF

Ls

Pa

Pp

R. carthamoides S. hortensisS. montana

0 5 10 15

SMF

SSF

SMF

SSF

SMF

SSF

Ls

Pa

Pp

°N

R. carthamoides S. hortensisS. montana

0

2

4

6

8

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

24 h 48 h 24 h 48 h 24 h 48 h

Ls Pa PpS. montana S. hortensisR. carthamoides

0

2

4

6

8

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

SMF SSF SMF SSF SMF SSF

Ls Pa Pp

g/10

0g

S. montana S. hortensisR. carthamoides

45

SMF with L. sakei Sm and in SMF with L. sakei Sh, respectively). Results of the ANOVA test indicated that the count of spores of aerobic mesophilic bacteria was significantly influenced by the fermentation method (p ≤ 0.0001) and by the LAB used for fermentation (p ≤ 0.0001). A significant negative correlation between the LAB count and the count of aerobic mesophilic bacteria spores (r = -0.773; p = 0.0001), between the Enterobacteria and the count of spores of aerobic mesophilic bacteria (r = -0.797; p = 0.0001) was found. Between the yeasts and molds count and the count of spores of aerobic mesophilic bacteria, a moderate positive significant correlation (r = 0.718; p = 0.0001) was found.

In all fermented samples, the count of Enterobacteria, yeasts and molds was reduced, as compared with nonfermented samples. The count of Enterobacteria, yeasts and molds was significantly influenced by the fermentation method (p ≤ 0.0001 and p ≤ 0.0002, respectively) and by the LAB used for fermentation (p ≤ 0.0001). A moderate negative significant correlation between the LAB and Enterobacteria count and between the count of yeasts and molds (r = -0.797; p = 0.0001 and r = -0.718; p = 0.0001, respectively) was found.

Figure 3.1.1.2. Microbiological parameters of SMF and SSF fermented savory plants (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus; SMF –

submerged fermentation; SSF – solid state fermentation; Sm – S. montana; Sh – S. hortensis; Rc - R. carthamoides; LAB –lactic acid bacteria; SAMB – spore of aerobic

mesophilic bacteria; Ent – Enterobacteria; Y/M - yeasts and mold).

The antimicrobial activity of savory plant bioproduct extracts and P.

acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 supernatants. The antimicrobial activities of fermented with selected LAB

0

2

4

6

8

10

SmLAB

ShLAB

RcLAB

SmSAMB

ShSAMB

RcSAMB

SmEntB

ShEntB

RcEntB

SmY/M

ShY/M

RcY/M

CFU/g log 10

Ls SMF Ls SSF Pa SMF Pa SSFPp SMF Pp SSF Non fermented

46

and nonfermented savory plants extracts are presented in Table 3.1.1.1. As could be seen from the obtained results, the extracts inhibited the growth of all tested bacteria. The diameters of the inhibition zones toward pathogenic strains varied between 5.8 ± 0.06 mm and 18.8 ± 0.19 mm.

The highest antimicrobial activity was demonstrated by the extract of Sm fermented with P. pentosaceus against Pseudomonas fluorescens biovar. V. The lowest inhibition zones were those fermented with P. pentosaceus Sh extracts against Bacillus subtilis. The antimicrobial activity of savory plant extracts and their bioproducts was significantly influenced by the type of LAB (against E.coli p ≤ 0.010, B. subtilis p ≤ 0.0001, P. fluorescens biovar. III p ≤ 0.023, and P. fluorescens biovar. V p ≤ 0.001) and by the type of a savory plant (against E. coli p ≤ 0.002 and against B. subtilis, P. fluorescens biovar. III and P. fluorescens biovar. V p ≤ 0.0001, respectively). The interaction of factors (LAB used for fermentation and the type of a savory plant) have a significant influence on the B. subtilis inhibition (p ≤ 0.0001).

Table 3.1.1.1. The antimicrobial activity of savory plant extracts and their bioproducts. *Antimicrobial activity including wall diameter (5 mm).

Microorganisms Zone of inhibition/mm* Pa Pp 8 Ls Nf

S. montana E. coli 9.0±0.09b 11.8±0.11d 9.4±0.10b 8.6±0.09a B. subtilis 6.5±0.07a 6.8±0.07b 8.8±0.09d 6.8±0.08b P. fluorescens biovar. III 13.2±0.14c 14.9±0.15d 14.9±0.16d 11.0±0.12a P. fluorescens biovar. V 16.0±0.18d 18.8±0.19e 18.0±0.20e 14.5±0.15a

S. hortensis E. coli 7.5±0.09a 9.8±0.09d 8.6±0.09c 7.3±0.07a B. subtilis 6.2±0.05c 5.8±0.06a 7.5±0.08d 6.6±0.07c P. fluorescens biovar. III 14.6±0.11d 12.4±0.10a 14.5±0.017d 13.8±0.14c P. fluorescens biovar. V 17.2±0.21e 16.4±0.15c 17.8±0.18e 15.6±0.16a

R. carthamoides E. coli 8.6±0.09b 8.6±0.09b 9.0±0.10c 8.0±0.09a B. subtilis - - - - P. fluorescens biovar. III 12.8±0.13e 11.2±0.12c 12.0±0.12e 9.2±0.09a P. fluorescens biovar. V 12.0±0.10c 12.0±0.12c 12.0±0.13c 10.0±0.11a Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, Nf - nonfermented

3.1.2. Parameters of fermented defatted soy flour, pea fiber, flaxseed

and Jerusalem artichoke

The moisture content, TTA, L(+) and D(-)-lactic acid isomer content and the pH of SMF and SSF fermented defatted soy flour, pea fiber, flaxseed

47

and Jerusalem artichoke are presented in Figure 3.1.2.1. In all cases, a higher TTA after SMF of defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke as compared with SSF (except in SMF with P. acidilactici and P. pentosaceus flaxseed) was found. The TTA of the tested plants ranged from 2.2 ± 0.14 ºN to 22.8 ± 0.43 ºN (in SSF with P. pentosaceus defatted soy flour and in SSF with P. pentosaceus flaxseed, respectively). It was found that the TTA was significantly influenced by the fermentation method (p ≤ 0.0001), by the type of LAB applied for the fermentation (p ≤ 0.0001), by the type of the plant product (p ≤ 0.0001), and the interaction of these factors was significant (p ≤ 0.0001). A moderate significant correlation between the substrate moisture and TTA (r = 0.398; p = 0.001) was found.

In most of the analysed plant samples, pH after SSF (except pea fiber) was found higher and ranged from 4.16 ± 0.07 to 6.19 ± 0.04 (in SMF with L. sakei and in SSF with P. acidilactici Jerusalem artichoke, respectively). The pH after 48 hours of fermentation was significantly influenced by the fermentation method (p ≤ 0.0001), by the type of LAB applied for fermentation (p ≤ 0.0001), by the type of plant product (p ≤ 0.0001), and the interaction of these factors was significant (p ≤ 0.0001). A moderate significant correlation between the substrate moisture content and pH (r = -0.655; p = 0.0001) and between the pH and TTA of the substrate (r = -0.305; p = 0.009) was found.

The results showed that all analysed LAB produced a mixture of L(+) and D(-)-lactic acid (Figure 3.1.2.1). The ratio of L(+) / D(-)-lactic acid isomers in defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke ranged from 1.2 to 2.4, from 1.6 to 2.5, from 0.6 to 9.9, and from 1.5 to 15.6, respectively, with the L(+)-lactic acid as the predominant isomer. The content of L(+) and D(-) lactic acid isomers was significantly influenced by the fermentation method (p ≤ 0.0001 and p ≤ 0.013, respectively), the type of LAB used for fermentation (p ≤ 0.0001), the type of plant product (p ≤ 0.0001). The interaction of these factors was significant (p ≤ 0.0001) for the content of L(+) and D(-) isomers. A moderate positive significant correlation between the pH and the L(+)-lactic acid content (r = 0.334; p = 0.004) was found. A moderate negative significant correlation between the content of TTA and L(+) and D(-)-lactic acid isomers (r = -0.554; p = 0.0001 and r = -0.392; p = 0.001, respectively), between the fermentable substrate moisture content and the L(+)-lactic acid content (r = -0.425; p = 0.0001), and between the L(+)-lactic acid and D(-)-lactic acid contents (r = 0.629; p = 0.0001) was found.

48

a)

b)

c)

d) Figure 3.1.2.1. Moisture content (%) (a), TTA (°N) (b), pH (c), and L-(+)

and D-(-) lactic acid isomers content (g/100 g) (d) of SMF and SSF fermented savory plants (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P.

pentosaceus; SMF – submerged fermentation; SSF – solid state fermentation). The count of LAB in SMF and SSF plant products is presented in Figure

3.1.2.2. Plants used in the experiment were a suitable substrate for LAB cultivation, and the LAB count in fermented plants ranged from 6.21 ± 0.05 log10 CFU/g to 9.98 ± 0.11 log10 CFU/g (in SMF with P. acidilactici Jerusalem artichoke and in SSF with P. pentosaceus defatted soy flour, respectively). The LAB count was significantly influenced by the fermentation method (p ≤ 0.0001), the type of LAB applied for the fermentation (p ≤ 0.0001), and by the type of plant product (p ≤ 0.0001). The interaction of the analysed factors (fermentation method, LAB used for the fermentation, and the type of plant product) was significant (p ≤ 0.0001) for the content of valuable LAB in the fermentable substrate. A moderate negative significant correlation between the LAB count and the moisture content (r = -0.357; p = 0.002), between the LAB count and TTA (r = -

0 50

SMF

SSF

SMF

SSF

SMF

SSFL

sP

aP

p %

H. tuberosus L. L. usitatissimum L.Pea fiber Deffated soy flour

0 5 10 15 20 25

SMF

SSF

SMF

SSF

SMF

SSF

Pa

Pp

Ls

Pea fiber L. usitatissimum L.H. tuberosus L. Deffated soy flour

0

5

10

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

24 48 24 48 24 48

Ls Pa PpDefatted soy flour H. tuberosus L.L. usitatissimum L. Pea fiber

0

5

10

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

L(+

)D

(-)

SMF SSF SMF SSF SMF SSF

Ls Pa Pp

g/100g

Defatted soy flour H. tuberosus L.L. usitatissimum L. Pea fiber

49

0.498; p = 0.0001), and between the LAB count and pH (r = 0.323; p = 0.006) of the substrate was found.

Figure 3.1.2.2. LAB count (log10 CFU/g) in SMF and SSF fermented

defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke. (Remarks: LAB – lactic acid bacteria, Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus; SMF –

submerged fermentation; SSF – solid state fermentation).

The activity of amylolytic and protelytic enzymes excreated by LAB in defatted soy flour, pea fiber, flaxseed, and Jerusalem artichoke is presented in Figure 3.1.2.3. Different tendencies of amylolytic enzymes activity in different plant products were estimated. The activity of amylolytic enzymes in fermented plants ranged from 1280.7 ± 5.65 AU/g to 22.1 ± 0.62 AU/g (in SMF with L. sakei Jerusalem artichoke and in SMF with P. acidilactici defatted soy flour, respectively). The results of the ANOVA test indicated that the activity of amylolytic enzymes was significantly influenced by the fermentation method (p ≤ 0.0001), by the type of LAB used for fermentation (p ≤ 0.0001) and by the type of plant product (p ≤ 0.0001). The interaction of the analysed factors (fermentation method, LAB used for fermentation, and the type of the plant product) was significant (p ≤ 0.0001) for the amylolytic enzymes activity in fermented plants. A moderate significant correlation between amylolytic enzymes activity and the TTA of fermented plants (r = 0.307; p = 0.009) was found.

In all the cases, a higher proteolytic enzymes activity in SMF defatted soy flour, pea fiber, flaxseed, and Jerusalem artichoke, as compared with SSF samples was found. Proteolytic enzymes activity in fermented samples ranged from 309.1 ± 1.65 AU/g to 1178.2 ± 3.26 AU/g (in SSF with L. sakei flaxseed and in SMF with P. acidilactici Jerusalem artichoke, respectively). It was found that the proteolytic enzymes activity was significantly influenced by the fermentation method (p ≤ 0.0001), by the type of LAB applied for fermentation (p ≤ 0.0001), and by the type of plant product (p ≤ 0.0001). The interaction of factors (fermentation method, the type of LAB

0

5

10

15

SMF SSF SMF SSF SMF SSF

Ls Pa Pp

CF

U/g

log 1

0

Defatted soy flour Pea fiber L. usitatissimum L. H. tuberosus L.

50

used for fermentation, and the type of plant) was significant for the activity of proteolytic enzymes (p ≤ 0.0001) of the fermentable substrate. A moderate negative significant correlation between the activity of proteolytic enzymes and the TTA (r = -0.461; p = 0.0001) of the substrate was found.

a)

b)

Figure 3.1.2.3. Amylolytic enzymes activity (AU/g) (a), Proteolytic enzymes activity (AU/g) (b) of fermented soy flour, pea fiber, flaxseed, and Jerusalem

artichoke (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus; SMF –

submerged fermentation; SSF – solid state fermentation; AU – activity units).

ARs, lignans, β-glucans, and the content of TPC in pea fiber. The content of ARs in fermented and nonfermented pea fiber is presented in Figure 3.1.2.4. It was found that the content of all ARs homologue decreased in all samples after SMF and SSF. The ARs content in SMF samples ranged from 3.0 ± 0.4 μg/g (C:21 homologue in SMF with L. sakei pea fiber) to 55 ± 0.3 μg/g (C:19 homologue in SMF with L. sakei pea fiber), in SSF samples – from 11.0±0.1 μg/g (C:21 homologue in SSF with P. acidilactici pea fiber) to 60.0 ± 0.7 μg/g (C:23 homologue in SSF with P.pentosaceus pea fiber).

The ARs homologue content was significantly influenced by the type of LAB applied for fermentation: C15:0 (p ≤ 0.004), C19:0 (p ≤ 0.001), C21:0 (p ≤ 0.0001), and C23 (p ≤ 0.001). A moderate positive significant correlation between ARs homologues C15:0 and C19:0 (r = 0.527; p = 0.014), a moderate negative significant correlation between the homologues C15:0 and C21:0 (r = -0.470; p = 0.032) and C15:0 and C23:0 (r = -0.793; p = 0.0001) was found.

The content of lignans in fermented and nonfermented pea fiber is presented in Figure 3.1.2.4. The content of matairesinol (MAT) increased in all samples after SMF and SSF. The MAT content in samples ranged from 78.3 ± 1.4 μg/g to 98.3 ± 2.0 μg/g, and in comparison with nonfermented

0

500

1000

1500

SMF SSF SMF SSF SMF SSF

Ls Pa Pp

AU

/g

Defatted soy flourH. tuberosus L.L. usitatissimum L.Pea fiber

0

500

1000

1500

SMF SSF SMF SSF SMF SSF

Ls Pa Pp

AU

/g

Defatted soy flourH. tuberosus L.L. usitatissimum L.Pea fiber

51

samples was found by 15.43% to 35.40%, 21.63% to 28.37% and 33.88% to 7.85% higher (in SMF and in SSF with L. sakei; in SMF and in SSF with P. acidilactici; in SMF and in SSF with P. pentosaceus samples, respectively).

a)

b) Figure 3.1.2.4. ARs homologues (μg/g) (a) and lignans (MAT and SECO) content (μg/100g) (b) in SMF and SSF fermented pea fiber (Remarks: ARs –

alkylresorcinols, Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, SMF – submerged fermentation,; SSF – solid state fermentation, Nf – nonfermented; secoisolariciresinol –

SECO, matairesinol - MAT).

In all the cases, a higher content of MAT in SSF samples was found. The MAT content was significantly influenced by the type of LAB applied for fermentation (p ≤ 0.001). The Secoisolariciresinol (SECO) content in SMF and SSF pea fiber samples was found higher as compared with nonfermented samples and ranged from 74.5 ± 1.5 μg/g to 121.2 ± 2.4 μg/g (in SMF with L. sakei and in SMF with P. acidilactici samples, respectively). A moderate significant correlation between the MAT and the SECO content in samples (r = 0.770; p = 0.0001) was found.

The content of TPC, ARs, lignans and β-glucans in nonfermented, SMF, and SSF fermented pea fiber is presented in Table 3.1.1.2. In all cases, a higher TPC content in SSF pea fiber samples as compared with SMF was found. The TPC content in the samples ranged from 22.05 ± 0.25 mg GAE/100g to 119.18 ± 3.51 mg GAE/100g (in nonfermented pea fiber and in SSF with P. pentosaceus pea fiber, respectively). It was found that the TPC content increased in all samples after SMF and SSF with L. sakei, P. acidilactici and P. pentosaceus (from 78.4% to 79.2%, from 78.4% to 78.6, from 79.5% to 81.5%, respectively) as compared with nonfermented samples. The results of the ANOVA test indicated that the TPC content was significantly influenced by the fermentation method (p ≤ 0.037) and the type of LAB used for fermentation (p ≤ 0.001). A strong negative significant correlation between the TPC content and the TTA (r = -0.897; p = 0.0001)

0

50

100

150

200

SMFSSF SMFSSF SMFSSF

Ls Pa Pp Nf

μg/g

C:15 C:19C:21 C:23

0

50

100

150

200

0

50

100

150

SMF

SSF

SMF

SSF

SMF

SSF

Ls Pa Pp Nf

μg/100gμg/100g

Matairesinol Secoisolariciresinol

52

of samples and a strong negative significant correlation between the TPC content and the pH (r = -0.966; p = 0.0001) of samples was found.

In all cases, a higher RSA after SSF of pea fiber as compared with SMF and nonfermented samples was found, and it ranged from 5.83 ± 0.07% (in nonfermented pea fiber) to 18.95 ± 0.19% (in SSF with P. pentosaceus pea fiber). The RSA increased in all samples after SMF and SSF with L. sakei, P. acidilactici and P. pentosaceus (from 3.02 to 3.24 times, from 2.92 to 3.06 times, and from 3.14 to 3.25 times, respectively) as compared with the nonfermented pea fiber. The RSA of the samples was significantly influenced by the type of LAB applied for the fermentation (p ≤ 0.002). A very strong positive significant correlation between the RSA and TPC content (r = 0.992; p = 0.0001), a very strong negative significant correlation between the RSA and the TTA (r = -0.929; p = 0.0001) and a very strong negative significant correlation between the RSA and the pH (r = -0.966; p = 0.0001) of the substrate was found.

In most of SSF pea fiber sampes a higher total content of β-glucans was found (except in SSF with P. pentosaceus pea fiber) as compared with SMF samples and ranged from 0.90 ± 0.01% to 1.40 ± 0.01%. In SMF samples, the total content of β-glucans was by 0.71% to 36.17% lower (in SMF with L. sakei and in SMF with P. pentosaceus pea fiber, respectively) compared with nonfermented. The total content of β-glucans in the samples was significantly influenced by the fermentation method (p ≤ 0.0001) and by the type of LAB applied for fermentation (p ≤ 0.0001), and the interaction of these factors was significant (p ≤ 0.0001). A moderate negative significant correlation between the total content of β-glucans and of TPC (r = -0.522; p = 0.015), a moderate negative significant correlation between the total content of β-glucans and RSA (r = -0.471; p = 0.031) and a very strong negative significant correlation between the total content of β-glucans and the pH (r = -0.976; p = 0.0001) of the substrate was found.

After SMF and SSF, the total content of ARs in pea fiber samples decreased and ranged from 74.0 ± 0.69 μg/g (in SMF with P. acidilactici samples) to 161 ± 0.80 μg/g (in SMF with P. acidilactici samples). Compared with nonfermented pea fiber, the total content of ARs in fermented samples was by 0.28% to 0.60% lower (in SMF with L. sakei and in SMF with P. pentosaceus pea fiber, respectively). The total ARs content in the pea fiber was significantly influenced by the fermentation method (p ≤ 0.021) and by the type of LAB applied for fermentation (p ≤ 0.014). A very strong negative significant correlation between the total ARs and TPC content (r = -0.887; p = 0.0001)and between the total ARs content and RSA (r = -0.857; p = 0.0001), a moderate negative significant correlation between the total content of ARs and the total lignans content (r = -0.527; p = 0.014),

53

a very strong positive significant correlation between the total ARs content and TTA (r = 0.864; p = 0.0001) and a strong positive significant correlation between the total ARs content and pH (r = 0.789; p = 0.0001) of the samples was found.

In most of the SSF pea fiber sampes, a higher total content of lignans was found (except in SSF with P. pentosaceus pea fiber), and it ranged from 158.3 ± 2.61 μg/100 g to 238.6 ± 4.85 μg/100 g. However, in all fermented samples from 14.97% to 43.59% higher total content of lignans (in SMF with L. sakei and in SSF with P. acidilactici pea fiber, respectively) was found as compared with nonfermented samples. The total content of lignans was significantly influenced by the fermentation method (p ≤ 0.047), the type of LAB applied for the fermentation (p ≤ 0.0001), and the interaction of these factors was significant (p ≤ 0.0001). A moderate positive significant correlation between the contents of total lignans and TPC (r = 0.479; p = 0.028), a moderate positive significant correlation between the total lignans content and RSA (r = 0.487; p = 0.025), a moderate negative significant correlation between the total lignans and ARs contents (r = -0.527; p = 0.014), a strong negative significant correlation between the total lignans and TTA contents (r = -0.601; p = 0.004) and a moderate negative significant correlation between the total lignans content and the pH (r = -0.485; p = 0.026) of the substrate was found.

Table 3.1.1.2. TPC, β-glucans, ARs and lignans content in pea fiber.

Pea fiber TPC content, mg GAE/100g

RSA, % Total β-glucans, %

Total ARs

content, μg/g

Total lignans content, μg/100 g

Ls SMF 102.13±1.81c 17.61±0.18d 1.40±0.01e 97±0.87c 158.3±2.61b SSF 105.98±1.53c 18.88±0.20e 1.32±0.02d 125±0.5d 176.4±2.93d

Pa SMF 102.31±2.10c 17.04±0.17c 1.29±0.01c 72±0.36a 218.4±4.01e SSF 102.94±2.15c 17.86±0.21d 1.24±0.02c 122±0.6d 238.6±4.85e

Pp SMF 107.63±4.12c 18.33±0.20e 0.90±0.01a 161±0.8e 167.8±2.84c SSF 119.18±3.51d 18.95±0.19e 0.95±0.01a 74±0.69a 165.5±3.12c

Nf 22.05±0.25a 5.83±0.07a 1.41±0.03e 267±0.8f 134.6±4.50a Data are the mean ± SD (n = 3); SD – standard deviation Mean values within column with different letters are significantly different (p < 0.05) SMF – submerged fermentation; SSF – solid state fermentation; TPC – total phenolic compounds; RSA – radical scavenging activity; ARs – alkylresorcinols; Ls – L. sakei; Pa – P. acidilactici; Pp – P. pentosaceus, Nf – nonfermented

The content of biogenic amines in defatted soy flour, pea fiber,

flaxseed and Jerusalem artichoke. The BAs content in defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke is presented in Table 3.1.1.3.

54

The main BA determined in deffated soy samples was putrescine (PUT), and its content ranged from 34.4 mg/kg (in SSF with P. pentosaceus samples) to 69.4 mg/kg (in SMF with P. acidilactici samples). Results of the ANOVA test indicated that the content of PUT was significantly influenced by the fermentation method (p ≤ 0.0001), by the type of LAB applied for fermentation (p ≤ 0.030) by the type of the plant product (p ≤ 0.0001), and the interaction of the factors was significant for the PUT content in defatted soy samples (p ≤ 0.0001). A moderate negative significant correlation between the PUT and PHE (r = -0.464; p = 0.0001), a moderate positive significant correlation between the PUT and TTA (r = 0.471; p = 0.0001), and a moderate positive significant correlation between the PUT and pH (r = 0.340; p = 0.003) of the substrate were found.

The main BA determined in the pea fiber was PHE, and its content ranged from 33.1 ± 3.31 mg/kg (in SMF with P. pentosaceus samples) to 48.74 ± 4.96 mg/kg (in SSF with L. sakei samples). In most of the cases, a higher (from 20.16% to 38.89%, respectively) PHE content was found after the SSF of the pea fiber as compared with SMF (except SSF with P. pentosaceus samples). The content of PHE was significantly influenced by the fermentation method (p ≤ 0.038), by the type of LAB used for fermentation (p ≤ 0.0001), and by the type of the plant product (p ≤ 0.0001). A moderate negative significant correlation between the PHE and PUT (r = -0.464; p = 0.0001), a strong positive significant correlation between the PHE and CAD (r = 0.749; p = 0.0001), a moderate positive significant correlation between the PHE and TTA of samples (r = 0.471; p = 0.0001), and a moderate negative significant correlation between the PUT and pH of samples (r = -0.343; p = 0.003) were found.

The main BA in flaxseed and Jerusalem artichoke samples was PHE, and its content ranged from 36.3 ± 1.62 mg/kg (in SSF with P. acidilactici Jerusalem artichoke) to 87.3 ± 11.4 mg/kg (in SMF with P. pentosaceus flaxseed) (Table 3.1.1.3).

The content of PHE was significantly influenced by the fermentation method (p ≤ 0.038), by the type of LAB (p ≤ 0.0001) and by the type of plant product (p ≤ 0.0001). A moderate negative significant correlation between the PHE and PUT (r = -0.464; p = 0.0001), a strong positive significant correlation between the PHE and cadaverine (CAD) (r = 0.749; p = 0.0001), a moderate positive significant correlation between the PHE and TTA of the samples (r = 0.471; p = 0.0001) and a moderate negative significant correlation between the PUT and pH of the samples (r = -0.343; p = 0.003) was found.

The total BAs content in defatted soy ranged from 115.8 ± 15.64 mg/kg (in SMF with L. sakei samples) to 225.8 ± 20.21 mg/kg (in SSF with L.

55

sakei samples), in pea fiber from 94.26 ± 10.08 mg/kg (in SMF with P. acidilactici samples) to 109.07 ± 11.94 mg/kg (in SSF with P. acidilactici samples), in flaxseed from 115.2 ± 20.58 mg/kg (in SSF with P. acidilactici samples) to 218.1 ± 16.74 mg/kg (in SMF with P. pentosaceus samples), and in Jerusalem artichoke from 115.1 ± 9.15 mg/kg (in SMF with L. sakei samples) to 136.9 ± 9 mg/kg (in SSF with P. pentosaceus samples).

Table 3.1.1.3. BAs content in defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke, mg/kg.

BAs L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF

Deffated soy flour PHE 40.2±2.40d 35.1±2.32d 19.2±0.67b 24.1±0.67c 28.2±1.32c 14.1±0.8a

PUT 48.3±7.11c 61.2±5.40d 69.4±11.3e 54.4±9.34d 43.4±3.85c 34.4±8.4a

CAD 8.2±0.97b 15.4±1.11e 8.8±1.68c 7.5±1.29a 8.2±1.02b 7.5±0.97a

HIS 2.1±0.35a 20.8±2.44f 3.8±2.34b 2.7±2.16b 2.8±2.16b 1.7±0.14a

TYR 7.3±1.26b 14.8±1.83e 11.6±7.01d 9.7±1.35c 9.6±1.33c 6.2±1.25a

SPD 47.1±11.5a 58.2±7.36b 56.1±14.6b 49.1±7.81a 61.1±10.1c 50.3±4.2a

SPR 9.7±0.83a 20.3±1.63d 13.8±0.13b 15.3±1.40c 15.3±0.10c 12.1±1.1b

Total 115.8±15a 225.8±20e 182.7±10d 162.8±8.9c 168.6±15c 126.3±3a

Pea fiber PHE 40.6±4.51b 48.7±4.96c 36.4±3.55b 50.6±5.15c 33.1±3.31a 31.3±3.3a PUT 15.5±1.23c 10.3±1.36a 12.6±1.30b 12.5±1.31b 15.6±1.60c 14.8±1.5c CAD 6.4±0.69b 5.5±0.62a 6.4±0.66b 6.4±0.66b 7.4±0.76c 7.0±0.65c HIS 2.4±0.31a 3.5±0.36b 4.7±0.45c 3.9±0.41b 5.4±0.54c 4.2±0.40b TYR 9.8±1.08d 8.4±0.93c 6.6±0.63a 6.0±0.60a 7.6±0.79b 7.3±0.61b SPD 15.5±1.63c 15.3±1.60c 12.1±1.20a 15.4±1.55c 17.2±1.80d 16.8±1.5d SPR 10.2±1.25a 9.9±1.10a 15.4±1.55b 14.3±1.43b 16.5±1.74c 17.1±1.7c Total 100.4±14a 101.6±9a 94.2±10.8a 109.1±12b 102.8±14b 98.5±7a

Linum usitatissimum L. PHE 70.5±11.2c 61.3±6.23b 63.3±14.1b 52.3±7.15a 87.3±11.4d 73.2±5.5c PUT 16.4±3.13c 8.7±1.30a 11.3±3.01b 9.7±2.34b 25.3±3.21d 8.5±2.41a CAD 17.3±6.12b 18.2±2.45c 18.3±6.64c 16.2±3.57b 24.3±7.62d 13.2±2.1a HIS 6.9±0.81c 1.5±0.21a 4.0±0.25b 2.5±0.27a 14.8±2.46d 5.5±0.72b TYR 37.3±1.35d 23.3±1.37a 39.3±1.88d 28.3±1.35b 47.3±1.74e 42.7±1.7c SPD 7.1±0.56c 5.6±0.78b 4.9±0.91b 3.7±0.86a 8.6±0.77c 2.7±0.32a SPR 1.8±0.45a 2.3±0.56a 2.0±0.32a 2.5±0.33a 10.5±0.75d 17.8±1.8e Total 157.3±21c 120.9±14a 143.1±11b 115.2±21a 218.1±17e 163.6±1d

Helianthus tuberosus L. PHE 40.8±1.40b 44.3±2.13c 44.3±3.22c 36.3±1.62a 37.6±2.30a 36.9±3.1a PUT 13.1±1.24b 11.8±1.39a 19.7±2.45c 14.5±1.31b 21.1±1.22d 18.6±1.7c CAD 11.6±0.72b 10.8±1.48a 13.3±1.84c 11.6±0.52b 11.1±0.71a 10.3±0.8a HIS 4.2±0.35a 13.7±0.64d 3.4±0.31a 11.6±0.55c 4.3±0.40a 15.1±0.8d TYR 29.1±1.14d 19.1±0.85b 28.6±1.25d 17.9±1.34a 31.8±2.49a 24.7±1.7c

56

Continue of Table 3.1.1.3. BAs L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF SPD 7.9±1.05a 17.2±0.99c 9.2±0.84a 21.4±2.15d 11.3±1.16d 22.4±1.4d SPR 8.4±0.83c 4.8±0.67a 8.9±0.66c 6.4±0.40b 10.9±1.15b 8.6±1.16c Total 115.1±9.2a 121.7±11 127.4±14c 119.7±14a 128.1±10a 136.6±9 f Data are the mean ± SD (n = 3); SD – standard deviation Mean values within row with different letters are significantly different (p < 0.05) SMF – submerged fermentation; SSF – solid state fermentation; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine

The total BAs content was significantly influenced by the type of LAB (p ≤ 0.018) and the type of plant product (p ≤ 0.0001). A moderate positive significant correlation between PHE, PUT, CAD and HIS and the total BAs content was found (r = 0.263; p = 0.026; r = 0.627; p ≤ 0.0001; r = 0.571; p ≤ 0.0001; r = 0.543; p ≤ 0.0001, respectively). Also, a moderate positive significant correlation between the total BAs content and the moisture of a substrate (r = 0.543; p = 0.044) was found. 3.2. Technological, microbiological and physical chemical parameters of

meat products

3.2.1. Parameters of marinated meat products

Parameters of pork loin marinated with Satureja montana and

Satureja hortensis bioproducts. The quality parameters of pork loin marinated with Sm and Sh bioproducts (MPL) are presented in Figure 3.2.1.1. The moisture content of MPL treated with Sm bioproducts ranged from 22.51 ± 0.02% (MPL treated with SSF with L. sakei Sm) to 29.61 ± 0.03% (MPL treated with SMF with L. sakei Sm), and moisture content of MPL treated with Sh bioproducts ranged from 21.62 ± 0.02% (MPL treated with SSF with L. sakei Sh) to 26.08 ± 0.03% (MPL treated with SMF with P. pentosaceus Sh). MPL treated with SSF savory plants pH in most of the cases was found higher (except MPL treated with SSF with P. pentosaceus Sm) as compared with SMF samples. In MPL samples, pH ranged from 5.86 ± 0.07 (MPL treated with SMF with L. sakei Sm) to 6.59 ± 0.08 (MPL treated with SSF with L. sakei Sm). The pH of MPL was significantly influenced by the savory plant bioproduct fermentation method, by the LAB used for savory plant bioproduct fermentation (p ≤ 0.0001) and by the type of a savory plant (p ≤ 0.0001), and the interaction of these factors on pH of MPL was significant (p ≤ 0.0001). A strong negative significant correlation

57

between the moisture and pH of MPL (r = -0.611; p = 0.0001), a very weak negative significant correlation between the MPL pH and savory plant TTA (r = -0.373; p = 0.019), a very weak negative significant correlation between the MPL pH and the moisture of a savory plant (r = -0.398; p = 0.012), and a very strong negative significant correlation between the MPL pH and the pH of savory plants (r = -0.281; p = 0.083) was found.

The drip loss of MPL treated with Sm ranged from 2.61 ± 0.02% (MBL treated with SSF with L. sakei Sm) to 3.37 ± 0.06% (MPL treated with SSF with P. pentosaceus Sm), and the drip loss of MPL treated with Sh ranged from 2.93 ± 0.03% (MPL treated with SMF with L. sakei Sh) to 3.21 ± 0.05% (MPL treated with SSF with P. pentosaceus Sh). The drip loss of marinated meat samples was significantly influenced by the type of LAB applied for the fermentation of savory plants (p ≤ 0.0001) and by the type of a savory plant (p ≤ 0.0001). The interaction of factors (fermentation method, LAB used for fermentation, and the type of savory plants) significantly influenced the drip loss of MPL (p ≤ 0.004). A very weak negative significant correlation between the drip loss and the pH of MPL (r = -0.464; p = 0.003) was found.

The WHC of most of the MPL sampes treated with Sm and Sh bioproducts was found to be higher (except MPL treated with SMF with P. acidilactici Sm, MPL treated with L. sakei Sm SMF, and MPL treated with P. pentosaceus Sh SMF) compared to control samples (nonmarinated meat). Sm and Sh bioproducts increased the WHC of MPL on average by 0.43% (Sm) and by 1.45% (Sh), respectively, compared with control samples. The WHC of MPL (pork loin marinated with Sm and Sh bioproducts) was significantly influenced by the fermentation method of savory plants (p ≤ 0.0001) and by the type of savory plants (p ≤ 0.0001). The interaction of factors (fermentation method, LAB used for the fermentation of savory plants, and the type of savory plants) have a significant influence on the marinated pork loin WHC (p ≤ 0.0001). A moderate negative significant correlation between the WHC and the moisture content of MPL (r = -0.471; p = 0.002) and a moderate positive significant correlation between the WHC and pH (r = 0.337; p = 0.036) of meat samples was found.

In most of the cases, a higher cooking loss was found of MPL marinated with SMF savory plants as compared with MPL marinated with SSF savory plants (except MPL marinated with SMF with P. acidilactici Sm and MPL marinated with SMF with P. pentosaceus Sm). In meat samples, the cooking loss ranged from 32.56 ± 0.69% to 42.18 ± 0.17% (MPL marinated with SMF with L. sakei Sh and MPL marinated with SMF with P. pentosaceus Sh, respectively). The cooking loss of MPL 24 hours marinated with Sm

58

and Sh bioproducts was significantly influenced by the fermentation method of savory plants (p ≤ 0.0001), by the LAB used for savory plant fermentation (p ≤ 0.0001) and by the type of a savory plant (p ≤ 0.0001). The interaction of these factors significantly influenced the cooking loss of meat samples (p ≤ 0.0001). A very weak negative significant correlation between the cooking loss and moisture of MPL (r = -0.366; p = 0.022), a very weak positive significant correlation between the tenderness and cooking loss of MPL (r = 0.442; p = 0.005), a very weak negative significant correlation between the cooking loss of MPL and TTA of a savory plant (r = -0.323; p = 0.045), a very weak negative significant correlation between the cooking loss of MPL and the moisture content of a savory plant (r = -0.329; p = 0.041) were found.

In most of the cases, Sm and Sh marinades increased the tenderness of meat on average by 55.5% and 35.5% (SMF savory plants) and by 48.7% and 28.5% (SSF savory plants), respectively, compared to control samples. Results of the ANOVA test indicated that the tenderness of pork loin after 24 hours of marination with Sm and Sh bioproducts was significantly influenced by the savory plant fermentation method (p ≤ 0.0001), by the LAB used for marinade production (p ≤ 0.0001), by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was significant for meat sample tenderness (p ≤ 0.0001). A moderate negative significant correlation between the tenderness and pH of MPL (r = -0.458; p = 0.003), a very weak positive significant correlation between the tenderness of MPL and the moisture of savory plants (r = 0.287; p = 0.077), a strong positive significant correlation between the tenderness of MPL and the TTA of savory plants (r = 0.292; p = 0.72) and a very weak positive significant correlation between the tenderness of MPL and the pH of savory plants (r = 0.450; p = 0.004) were found.

The IF of MPL marinated with Sm and Sh bioproducts ranged from 3.03 ± 0.03% to 6.75 ± 0.08%, and was from 15.19% to 48.29% (MPL treated with Sm) and from 14.51% to 45.90% (MPL treated with Sh) lower compared with control meat samples. The IF was significantly influenced by the savory plant bioproduct fermentation method (p ≤ 0.0001), by the LAB used for the fermentation of savory plants (p ≤ 0.0001), by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was significant for the IF content (p ≤ 0.0001). A very weak negative significant correlation between the IF and the tenderness of meat samples (r = -0.352; p = 0.028), a very weak negative significant correlation between the IF of MPL and the TTA of savory plants (r = -0.409; p = 0.01), a very weak negative significant correlation between the IF of MPL and the pH of savory plants (r = -0.415; p = 0.009) were found.

59

Figure 3.2.1.1. Moisture content (%) (a), pH (b), drip loss (%) (c), WHC (%) (d), cooking loss (%) (e), shear force kg/cm2 and IF (%) of marinated with SMF and SSF fermented savory plants bioproducts pork loin (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus; SMF – submerged fermentation; SSF – solid state fermentation, intramuscular fat,WHC – water holding capacity, SF –

shear force).

a)

b)

c)

d)

e)

f)

0

10

20

30

SMF SSF SMF SSF SMF SSF

Pa Pp Ls

%

S. montana S. hortensisControl

0

5

10

SMF SSF SMF SSF SMF SSF

Pa Pp LsS. montana S. hortensisControl

0

2

4

SMF SSF SMF SSF SMF SSF

Pa Pp Ls

%

S. montana S. hortensisControl

0

50

100

55 60 65 70

SMF

SSF

SMF

SSF

SMF

SSF

Pa

Pp

Ls

%

S. hortensis S. montanaControl

0

20

40

60

0 20 40 60

SMF

SSF

SMF

SSF

SMF

SSF

Pa

Pp

Ls

%

S. hortensis S. montanaControl

02468

0123456

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

SMF

SSF

Pa Pp Ls Pa Pp Ls

S. montana S. hortensis

kg/cm2%

SF IF Control

60

Significant differences were observed in the L*, a*, and b* parameters (p < 0.05) of MPL samples (Figure 3.2.1.2). The MPL control samples showed the highest lightness (L* 47.41 ± 0.47), however, the lowest redness (a* 4.62 ± 0.06) as compared with marinated MPL samples. MPL samples marinated with SSF Sm and Sh showed a higher lightness (L*) (between 29.44 ± 0.18 and 39.82 ± 0.37, MPL treated with SSF with P. pentosaceus Sh, and MPL treated with SSF with P. acidilactici Sh, respectively), redness (a*) (ranged between 5.73 ± 0.05 and 12.61 ± 1.10, MPL treated with SSF with P. pentosaceus Sh, and MPL treated with SSF with L. sakei Sm, respectively), and yellowness (b*) (ranged between 2.97 ± 0.04 and 10.80 ± 0.13, MPL treated with SSF with P. pentosaceus Sh and MPL treated with SSF with P. acidilactici Sh, respectively).

Figure 3.2.1.2. Colour parameters (L*, a* and b*) of marinated with SMF and SSF fermented savory plants bioproducts pork loin (Remarks: Ls – L. sakei,

Pa – P. acidilactici, Pp – P. pentosaceus; SMF – submerged fermentation; SSF – solid state fermentation, C – control).

The MPL control samples showed the highest lightness (Figure 3.2.1.2)

(L* 47.41 ± 0.47), however, the lowest redness (a* 4.62 ± 0.06) as compared with marinated MPL samples. MPL samples marinated with SSF Sm and Sh showed a higher lightness (L*) (between 29.44 ± 0.18 and 39.82 ± 0.37, MPL treated with SSF with P. pentosaceus Sh, and MPL treated with SSF with P. acidilactici Sh, respectively), redness (a*) (ranged between 5.73 ± 0.05 and 12.61 ± 1.10, MPL treated with SSF with P. pentosaceus Sh, and MPL treated with SSF with L. sakei Sm, respectively), and yellowness (b*) (ranged between 2.97 ± 0.04 and 10.80 ± 0.13, MPL treated with SSF with P. pentosaceus Sh and MPL treated with SSF with P. acidilactici Sh, respectively). While meat samples marinated with SMF Sm and Sh showed a higher lightness (L*) (ranged between 30.70 ± 0.41 and 39.78 ± 0.30, MPL marinated with SMF with P. pentosaceus Sh

0 10 20 30 40 50 60 70

SMF

SMF

SSF

SSF

SSF

SSF

Ls

Pa

Pp

Ls

Pa

Pp

C

S. m

onta

na

S. h

orte

nsis

L* a* b*

61

and MPL marinated with SMF with P. acidilactici Sh, respectively), and redness (ranged between 5.78 ± 0.04 and 10.62 ± 0.09, MPL marinated with SMF with P. pentosaceus Sh, and MPL marinated with SMF with P. acidilactici Sm, respectively), as well as yellowness (ranged between 1.11 ± 0.09 and 11.37 ± 0.10, MPL marinated with SMF with P. pentosaceus Sh and MPL marinated with SMF with P. acidilactici Sh, respectively). The L*, a*, and b* parameters of meat were significantly influenced by the savory plants fermentation method (p ≤ 0.0001), by the LAB used for savory plants fermentation (p ≤ 0.0001), by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was signifinant on samples L*, b* (p ≤ 0.0001) and a* (p ≤ 0.042). A strong positive significant correlation between the (L*) and (b*) (r = 0.683; p = 0.0001) and a moderate positive significant correlation between the (a*) and (b*) (r = 0.339; p = 0.028) of meat samples were found.

Parameters of beef loin marinated with Satureja montana and

Satureja hortensis bioproducts. The quality parameters of beef loin (MBL) marinated with Sm and Sh bioproducts are presented in Table 3.2.1.1. In all cases, a higher moisture content was found in MBL marinated with Sm as compared with control beef meat samples and ranged from 22.57 ± 0.02% (MBL marinated with SMF with P. acidilactici Sm) to 24.47 ± 0.03% (MBL marinated with SSF with L. sakei Sm). Different tendencies of beef loin marinated with Sh bioproducts were estimated. As compared with control meat samples, meat samples marinated with Sh had a lower moisture content (except in MBL marinated with SMF with P. pentosaceus Sh and MBL marinated with SSF with L. sakei Sh).

In all cases, the pH of MBL marinated with SSF savory plant bioproducts was higher (except MBL samples marinated with SSF with P. pentosaceus Sm). The pH of the samples ranged from 6.02 ± 0.03 (MBL marinated with SMF with P. pentosaceus Sm) to 6.40 ± 0.07 (MBL marinated with SSF P. acidilactici Sm). The pH of beef loin samples 24 hours marinated with Sm and Sh bioproducts was significantly influenced by the fermentation method of marinades produced from savory plants (p ≤ 0.0001). A moderate negative significant correlation between the MBL pH and marinade moisture content (r = -0.426; p = 0.007), a weak negative significant correlation between the MBL pH and marinade TTA (r = -0.379; p = 0.017) were found.

In all the cases, a higher drip loss was found in MBL marinated with SSF plant bioproducts as compared with MBL marinated with SMF plant bioproducts (except MBL fermented with SSF with P. pentosaceus Sh). The drip loss of MBL marinated with Sm bioproducts ranged from 1.96 ± 0.01%

62

(MBL marinated with SSF with P. pentosaceus Sm and MBL marinated with SMF with L. sakei Sm) to 2.23 ± 0.02% (MBL marinated with SSF with P. acidilactici Sm), and in MBL marinated with Sh bioproducts it ranged from 1.89 ± 0.01% (MBL marinated with SMF with L. sakei Sh) to 2.11 ± 0.03% (MBL marinated with SSF with P. acidilactici Sh).

The drip loss of MBL was significantly influenced by the type of the marinade fermentation method (p ≤ 0.0001), by the type of LAB used for the fermentation of savory plants (p ≤ 0.0001), and by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was significant for the drip loss of MBL (p ≤ 0.0001). A moderate negative significant correlation between the drip loss of MBL and the moisture content of savory plant bioproducts (r = -0.434; p = 0.006), a moderate negative significant correlation between the drip loss of MBL and the TTA of savory plant bioproducts and between the drip loss and the cooking loss of MBL (r = -0.385; p = 0.016) were found.

The higher WHC of MBL marinated with Sm and Sh bioproducts, compare to control samples (except MBL marinated with SSF with L. sakei Sh) was found. The Sm and Sh marinades increased the WHC of MBL samples on average by 1.19% (Sm) and by 2.50% (Sh), respectively, as compared to MBL control samples. The WHC of MBL marinated 24 hours was significantly influenced by the fermentation method of savory plant for marinade production used (p ≤ 0.0001), by the type of LAB applied for savory plant fermentation (p ≤ 0.0001), by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was significant on beef loin WHC (p ≤ 0.0001). A weak negative significant correlation between the WHC of MBL and moisture content of savory plant bioproducts (r = -0.327; p = 0.042), a weak negative significant correlation between the WHC of MBL and the pH of MBL (r = -0.338; p = 0.035), a weak negative significant correlation between the WHC of MBL and the cooking loss of MBL (r = -0.333; p = 0.038), a moderate positive significant correlation between the WHC of MBL and the TTA of savory plant bioproducts (r = 0.400; p = 0.012) were found.

In all the cases, a higher cooking loss was found in MBL marinated with SMF savory plants as compared with SSF (except in MBL marinated with SMF with P. pentosaceus Sm and MBL marinated with SMF with P. acidilactici Sh). In MBL samples, the cooking loss ranged from 31.70 ± 0.10% to 40.71 ± 0.15% (MBL marinated with SSF with L. sakei Sm and MBL marinated with SSF with P. pentosaceus Sm, respectively). The MBL cooking loss was significantly influenced by the savory plant marinade fermentation method (p ≤ 0.0001), by the type of LAB applied for savory plant marinade production (p ≤ 0.0001) and by the type of savory plants (p ≤

63

0.0001), and the interaction of these factors was significant for the cooking loss of MBL (p ≤ 0.0001). A weak negative significant correlation between the cooking loss and the moisture content of MBL (r = -0.338; p = 0.035), a weak positive significant correlation between the cooking loss and the drip loss of meat samples (r = 0.385; p = 0.016), a weak negative significant correlation between the cooking loss and the WHC of MBL (r = -0.333; p = 0.038), a weak positive significant correlation between the cooking loss and tenderness of MBL (r = 0.372; p = 0.020), a weak negative significant correlation between the cooking loss of MBL and the TTA of savory plants (r = -0.385; p = 0.016), a moderate negative significant correlation between the cooking loss of MBL and the pH of savory plant bioproducts (r = -0.417; p = 0.008) were found.

In all the cases, beef loin marination with Sm and Sh decreased the tenderness of meat on average by 30.2% and 26.7% (SMF) and by 30.6% and 36.4% (SSF), respectively, as compared with control samples. Meat sample tenderness ranged from 1.90 ± 0.01 (MBL treated with SMF with L. sakei Sh) to 3.86 ± 0.09 (control samples).

The tenderness of beef loin samples after 24 hours of marination with Sm and Sh bioproducts was significantly influenced by the bioproduct fermentation method (p ≤ 0.0001), by the LAB used for bioproduct fermentation (p ≤ 0.0001), by the type of savory plants (p ≤ 0.006), and the interaction of these factors was significant for the tenderness of MBL (p ≤ 0.0001). A weak negative significant correlation between the tenderness and moisture content of MBL (r = -0.381; p = 0.0017), a weak positive significant correlation between the tenderness and pH of MBL (r = 0.352; p = 0.028), a weak positive significant correlation between the tenderness and cooking loss of MBL (r = 0.372; p = 0.020), a weak positive significant correlation between the tenderness and IF of MBL (r = 0.389; p = 0.014), a moderate negative significant correlation between the tenderness of MBL and moisture content of savory plant bioproducts (r = -0.489; p = 0.002), a strong negative significant correlation between the tenderness of MBL and the TTA of savory plants (r = -0.635; p = 0.0001), and a moderate negative significant correlation between the tenderness of MBL and the pH of savory plants (r = -0.556; p = 0.0001) were found.

The IF content in MBL marinated with Sm marinades ranged from 1.65 ± 0.01% (MBL marinated with SMF with L. sakei Sm) to 2.46 ± 0.05% (MBL marinated with SMF with P. pentosaceus Sm) and in MBL marinated with Sh ranged from 0.98 ± 0.01% (MBL marinated with SMF with P. acidilactici Sh) to 4.14 ± 0.05% (MBL marinated with SMF with L. sakei Sh). The IF of MBL was significantly influenced by the LAB used for savory plant bioproducts fermentation (p ≤ 0.0001), by the type of savory

64

plants (p ≤ 0.0001), and the interaction of these factors was significant for the IF content of MBL (p ≤ 0.0001). A weak positive significant correlation between the IF and the tenderness of MBL (r = 0.389; p = 0.014) was found.

Table 3.2.1.1. Quality parameters of the beef loin marinated with Sm and Sh bioproducts.

Sample L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF Control

S. montana MC, % 22.80

±0.02a 24.47 ±0.03c

22.57 ±0.02a

23.41 ±0.01b

24.27 ±0.05c

23.77 ±0.02b

22.46 ±0.02b

pH 6.24 ±0.05b

6.31 ±0.06b

6.05 ±0.03a

6.40 ±0.07c

6.02 ±0.03a

6.30 ±0.05b

6.33 ±0.07c

DL, % 1.96 ±0.01a

1.98 ±0.01a

2.18 ±0.01c

2.23 ±0.02c

1.96 ±0.01a

2.05 ±0.02b

2.17 ±0.02c

WHC, % 62.35 ±0.16b

63.41 ±0.14c

62.46 ±0.12b

63.03 ±0.13c

61.94 ±0.14a

62.57 ±0.11b

61.89 ±0.10a

CL, % 38.04 ±0.12c

31.70 ±0.10a

40.45 ±0.14d

38.72 ±0.13c

39.58 ±0.14c

40.71 ±0.15d

42.75 ±0.16c

SF, kg/cm2

2.54 ±0.02b

2.78 ±0.04c

2.80 ±0.09c

2.88 ±0.05d

2.40 ±0.01a

2.72 ±0.03c

3.86 ±0.09e

IF, % 1.65 ±0.01a

2.44 ±0.04d

2.01 ±0.03b

2.45 ±0.04d

2.46 ±0.05d

2.25 ±0.06c

2.23 ±0.05d

S. hortensis MC, % 20.78

±0.02a 22.58 ±0.03b

22.29 ±0.01b

21.16 ±0.03a

25.11± 0.03c

21.29 ±0.02a

22.46 ±0.02b

pH 6.09 ±0.04a

6.16 ±0.07a

6.32 ±0.05c

6.11 ±0.03a

6.25 ±0.04b

6.29 ±0.06b

6.33 ±0.07c

DL, % 1.89 ±0.01a

2.03 ±0.01b

2.03 ±0.03b

2.11 ±0.03c

2.07 ±0.02b

1.96 ±0.02a

2.17 ±0.02c

WHC, % 63.26 ±0.15b

61.88 ±0.16a

64.55 ±0.12d

63.74 ±0.14c

63.07 ±0.10b

64.12 ±0.09c

61.89 ±0.10a

CL, % 40.39 ±0.16b

37.26 ±0.13a

37.88 ±0.17a

39.61 ±0.13b

38.31 ±0.11b

37.85 ±0.16a

42.75 ±0.16c

SF, kg/cm2

3.16 ±0.03d

1.90 ±0.01a

2.53 ±0.01c

2.59 ±0.02c

2.31 ±0.02b

3.37 ±0.03d

3.86 ±0.09e

IF, % 0.98 ±0.01a

2.27 ±0.03d

1.70 ±0.03c

2.00 ±0.02d

4.14 ±0.05e

1.68 ±0.01c

2.23 ±0.05d

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; WHC – water holding capacity; IF – intramuscular fat; CL – cooking loss, SF – shear force, DL – drip loss, MC – moisture content; SMF – submerged fermentation; SSF – solid state fermentation The colour parameters (L*, a* and b*) of the beef loin marinated with

Sm and Sh bioproducts are presented in Figure 3.2.1.3. It was found that marination with Sm and Sh bioproducts decreased the values of (L*) (from

65

22.64%, MBL marinated with SMF with L. sakei Sm, to 32.97%, MBL marinated with SSF with P. pentosaceus Sm, and from 27.25%, MBL marinated with SSF with P. pentosaceus Sh, to 34.50%, MBL marinated with SSF with Pa Sh, respectively). It was indicated that the value of (L*) was significantly influenced by the fermentation method of savory plant bioproducts (p ≤ 0.0001), by the LAB used for the fermentation of savory plants (p ≤ 0.04), by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was significant on (L*) of MBL (p ≤ 0.0001). A strong positive correlation between the meat (L*) and (a*) parameters (r = 0.720; p = 0.0001), a strong negative correlation between the meat (L*) and the moisture of savory plant bioproducts (r = -0.613; p = 0.0001), a very strong negative correlation between the meat (L*) and the pH of savory plant bioproducts (r = -0.859; p = 0.0001) were found.

Figure 3.2.1.3. Colour parameters (L*, a* and b*) of marinated with SMF and SSF fermented savory plants bioproducts beef loin (Remarks: Ls – L. sakei,

Pa – P. acidilactici, Pp – P. pentosaceus; SMF – submerged fermentation; SSF – solid state fermentation, C – control).

In all the cases, beef loin marination with Sm and Sh decreased the (a*)

values, and they ranged from 5.88 ± 0.07 (MBL marinated with SMF with P. acidilactici Sh) to 15.16 ± 0.15 (control samples). The values of (a*) of beef loin marinated 24 hours were significantly influenced by the savory plant fermentation method (p ≤ 0.0001), by the LAB used for savory plant fermentation (p ≤ 0.0001), by the type of savory plants (p ≤ 0.0001), and the interaction of these factors was significant on the (a*) of MBL (p ≤ 0.0001). A strong positive correlation between the meat (L*) and (a*) parameters (r = 0.720; p = 0.0001), between the meat (a*) and (b*) parameters (r = 0.625; p = 0.0001), a moderate negative correlation between the meat (a*) parameter and the pH of savory plant bioproducts (r = -0.454; p = 0.004) and a weak

0 10 20 30 40 50 60

SMF

SMF

SSF

SSF

SSF

SSF

Ls

Pa

Pp

Ls

Pa

Pp

C

S. m

onta

na

S. h

orte

nsis

L* a* b*

66

negative correlation between the meat (a*) and the TTA of savory plant bioproducts (r = -0.381; p = 0.017) were found.

Marination has an influence on the yelowness (b*) of beef loin samples: the beef meat marination with Sm bioproducts decreased the (b*) values on average by 17.78%, and marination with Sh increased the (b*) values on average by 54.30% as compared with control samples. The (b*) values of meat 24 hours marinated with Sm and Sh were significantly influenced by the marinade fermentation method (p ≤ 0.0001), by the LAB used for marinade fermentation (p ≤ 0.0001), by the type of a savory plant (p ≤ 0.0001), and the interaction of these factors was significant for the (b*) of MBL (p ≤ 0.0001). A strong positive correlation between the meat (b*) and (a*) parameters (r = 0.625; p = 0.0001), a weak positive correlation between the meat (b*) and the TTA of savory plant bioproducts (r = 0.322; p = 0.046) were found.

Parameters of pork neck, shoulder, ham, M. longissimus dorsi and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8, and L. sakei KTU05-6 strains cultivated in potato juice. The parameters of pork meat, marinated with selected LAB strains cultivated in potato juice are presented in Table 3.2.1.2. The quality parameters of pork meat samples after 24 h of marination significantly depended on the LAB strain used for marinade preparation (p ≤ 0.0001) and the part of pork meat (p ≤ 0.0001). The marination of pork neck, ham muscle, M. longissimus dorsi and loin samples with P. acidilactici and P. pentosaceus marinades lowered the meat moisture content by 5.5, 4.5, 2.4% and on average by 14.3%, respectively, as compared with control samples, while the marination effect on the pork shoulder moisture content was not significant. The L. sakei marinade increased moisture content in all tested meat samples (on average by 10.6%), and at a higher level – in loin samples (20.6%).

P. acidilactici and P. pentosaceus marinades lowered the pH of meat on average by 3.9% (in neck and shoulder samples) and by 16.8% (in the muscle M. longissimus dorsi and loin samples) as compared with control samples. The lowest pH has been reached after the marination of pork meat with L. sakei marinade (meat pH reduced by 10% (neck), by 3.6% (shoulder) and on average by 20.7% in other samples). A strong negative significant correlation between the pH of meat samples and drip loss (r = -0.607; p = 0.0001), a moderate positive significant correlation between the pH of meat samples and WHC (r = 0.548; p = 0.0001), a moderate negative significant correlation between the pH of meat samples and cooking loss (r = -0.522; p = 0.0001), a weak negative significant correlation between the pH of meat samples and tenderness (r = -0.290; p = 0.025), and a moderate

67

positive significant correlation between the pH of meat samples and IF (r = 0.513; p = 0.0001) were found. In all the cases, marinated pork meat samples had from 30.3% (M. longissimus dorsi) to 92.9% (pork loin) higher drip loss, and it ranged from 3.20 ± 0.03% (in pork neck samples marinated with P. acidilactici marinade) to 1.52 ± 0.01% (in pork neck control samples). A strong negative significant correlation between the meat sample drip loss and pH (r = -0.607; p = 0.0001), a moderate negative significant correlation between the meat sample drip loss and WHC (r = -0.519; p = 0.0001), a strong positive significant correlation between the meat sample drip loss and the cooking loss (r = 0.642; p = 0.0001), and a weak negative significant correlation between the meat samples drip loss and IF (r = -0.388; p = 0.004) were found.

Marination increased the cooking loss of meat on average by 11.8% (of neck and shoulder) and 30.5% (of M. longissimus dorsi and loin samples), and lowered the WHC of loin by 7.6%, of neck, shoulder and muscle on average by 13%, and of M. longissimus dorsi by 22.2%, as compared with control samples. A moderate positive significant correlation was found between the meat WHC and pH (r = 0.548; p = 0.0001); also, a moderate negative significant correlation between the meat sample WHC and drip loss (r = -0.519; p = 0.0001), a strong negative significant correlation between the meat sample WHC and cooking loss (r = -0.636; p = 0.0001) was found.

Table 3.2.1.2. Quality parameters of pork meat, marinated with selected LAB strains cultivated in potato juice.

Meat samples

Moisture, %

pH Drip loss, %

WHC, %

CL, %

Shear, kg/cm2

IF, %

Neck

Pa 19.90 ±0.11b

6.20 ±0.01b

3.20 ±0.03d

56.73 ±0.10a

44.16 ±0.23d

0.835 ±0.01a

5.46 ±0.03b

Pp 19.51 ±0.10b

6.35 ±0.01b

2.25 ±0.02c

56.85 ±0.10a

44.55 ±0.47d

0.849 ±0.02a

5.21 ±0.01a

Ls 18.38 ±0.09a

6.03 ±0.01a

2.54 ±0.02c

58.94 ±0.12b

40.36 ±0.21b

0.860 ±0.02b

5.42 ±0.04b

Control 20.85 ±0.10c

6.70 ±0.03c

1.52 ±0.01a

67.84 ±0.18c

38.28 ±0.31a

0.878 ±0.01b

5.57 ±0.03c

Shoulder

Pa 23.92 ±0.11b

5.88 ±0.02a

2.85 ±0.03c

56.05 ±0.10b

44.26 ±0.47d

0.846 ±0.03a

3.90 ±0.01b

Pp 24.01 ±0.14b

5.89 ±0.01a

2.60 ±0.03c

53.78 ±0.11a

44.26 ±0.43d

0.857 ±0.04a

3.85 ±0.01a

Ls 21.43 ±0.12a

5.84 ±0.01a

2.55 ±0.02c

60.18 ±0.16c

43.05 ±0.38c

0.876 ±0.02b

3.72 ±0.02a

Control 24.11 ±0.13c

6.06 ±0.01b

1.84 ±0.01a

64.33 ±0.17d

39.40 ±0.34a

0.897 ±0.01b

5.91 ±0.04d

68

Continue of Table 3.2.1.2. Meat samples

Moisture, %

pH Drip loss, %

WHC, %

Cooking loss, %

Shear force,

kg/cm2

IF, %

Ham muscle

Pa 24.75 ±0.11b

5.24 ±0.01b

3.10 ±0.03c

63.56 ±0.16c

43.81 ±0.40c

1.023 ±0.05a

4.66 ±0.04a

Pp 24.13 ±0.12b

5.33 ±0.00b

2.86 ±0.03c

53.08 ±0.10a

44.30 ±0.43c

1.035 ±0.03a

4.90 ±0.02b

Ls 22.83 ±0.15a

5.09 ±0.01a

1.95 ±0.01a

56.71 ±0.14a

44.62 ±0.51d

1.062 ±0.04b

4.89 ±0.03b

Control 25.67 ±0.16c

6.42 ±0.02c

1.74 ±0.01a

65.20 ±0.15c

35.13 ±0.23a

1.098 ±0.02c

5.29 ±0.01c

M. longissimus dorsi

Pa 25.22 ±0.16b

5.30 ±0.01b

2.62 ±0.03c

66.20 ±0.15c

41.18 ±0.31d

0.839 ±0.08a

2.53 ±0.01a

Pp 25.58 ±0.14b

5.52 ±0.01b

2.45 ±0.02b

54.02 ±0.11a

39.12 ±0.30c

0.858 ±0.04a

2.44 ±0.02a

Ls 23.43 ±0.14a

5.14 ±0.01a

2.76 ±0.03c

53.72 ±0.10a

40.74 ±0.36d

0.909 ±0.04c

2.76 ±0.01b

Control 26.02 ±0.11c

6.52 ±0.02c

2.01 ±0.02a

74.46 ±0.18d

29.41 ±0.20a

0.889 ±0.01b

3.24 ±0.01d

Loin

Pa 25.06 ±0.16b

5.30 ±0.01c

3.14 ±0.03d

55.09 ±0.10b

42.80 ±0.41d

1.020 ±0.03a

3.50 ±0.02a

Pp 24.87 ±0.14b

5.41 ±0.01c

3.07 ±0.03d

55.45 ±0.11b

43.37 ±0.44d

1.055 ±0.06b

3.41 ±0.01a

Ls 23.17 ±0.14a

5.06 ±0.00a

2.76 ±0.03c

52.61 ±0.11a

40.82 ±0.38c

1.084 ±0.05b

3.61 ±0.02b

Control 29.17 ±0.19c

6.35 ±0.01d

1.55 ±0.01a

58.86 ±0.12c

32.94 ±0.21a

1.107 ±0.04c

4.88 ±0.02c

Data are the mean ± SD (n = 3); SD – standard deviation; Mean values within column with different letters are significantly different (p < 0.05); Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; WHC – water holding capacity; IF – intramuscular fat, CL – cooking loss

A moderate negative significant correlation between the meat samples cooking loss and moisture content (r = -0.517; p = 0.0001), a moderate negative significant correlation between the meat sample cooking loss and pH (r = -0.522; p = 0.0001), a strong positive significant correlation between the meat samples drip loss and cooking loss (r = 0.642; p = 0.0001) was found.

Marination with LAB-based marinades decreased IF in ham muscle samples by 8.9% and in M. longissimus dorsi and loin samples on average by 25% (Table 3.2.1.2). Also, marination ingreased the drip loss of pork meat samples. A weak negative significant correlation between the IF and

69

moisture content of meat samples (r = -0.364; p = 0.004), a moderate positive significant correlation between the meat IF content and pH (r = 0.513; p = 0.004), and a weak negative significant correlation between the meat sample IF and drip loss (r = -0.388; p = 0.002) were found.

Colour parameters. It was found that lighteness (L*) and redness (a*)

increased in all the five treated groups (p < 0.05) of pork meat (Figure 3.2.1.4). Changes of colour parameters (L*, a* and b*) depended on the LAB strain used for marinade production (p ≤ 0.0001), and on the part of pork meat (p ≤ 0.0001). Marination increased the (L*) of pork samples (from 3.5% in ham muscle to 21.4% in pork loin) as compared with control samples, and it ranged from 45.63 ± 0.19 (control pork loin samples) to 65.99 ± 0.26 (with L. sakei marinated pork ham muscle samples). A strong negative significant correlation between the meat lighteness (L*) and pH (r = -0.602; p = 0.0001) was found.

Marination increased the (a*) of meat (both pedioccoci from 3.2% to 33.8% and L. sakei from 1.4% to 24.0%) as compared to the control samples. The values of (a*) ranged from 3.54 ± 0.05 (control M. longissimus dorsi samples) to 14.01 ± 0.13 (with P. pentosaceus marinated pork shoulder muscle samples). A strong negative significant correlation between meat redness (a*) and meat yelowness (b*) (r = -0.647; p = 0.0001) was found.

Figure 3.2.1.4. Colour parameters (L*, a* and b*) of pork meat, marinated with selected LAB strains cultivated in potato juice (Remarks: Ls – L. sakei, Pa –

P. acidilactici, Pp – P. pentosaceus, C – control).

Marination had an influence on meat sample yellowness (b*). Yellowness was decreased in M. longissimus dorsi (4.4%), neck (7.2%), ham muscle (13.02%), loin (15.2%) and shoulder (24.1%) samples. A moderate positive significant correlation between meat yelowness (b*) and meat pH (r = 0.431; p = 0.001) was found.

0102030405060708090

100

PaPpLs C PaPpLs C PaPpLs C PaPpLs C PaPpLs C

Neck Shoulder Ham Mld Loin

b*

a*

L*

70

The BAs content in pork neck, shoulder, ham, M. longissimus dorsi

and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice. The BAs content in marinated pork meat is presented in Table 3.2.1.3.

The BAs content significantly depended on the LAB strain used for marinade production (p ≤ 0.0001) and on the part of pork meat (p ≤ 0.0001). In many cases marination increased the PHE content in pork (up to 16.47–37.49 mg/kg), except pork shoulder and ham muscle samples and with L. sakei and P. acidilactici marinated M. longissimus dorsi. A weak negative significant correlation between the PHE and meat pH (r = -0.347; p = 0.007) and a weak positive significant correlation between the PHE and histamine (HIS) content (r = 0.313; p = 0.015) were found.

The PUT (14.56–131.29 mg/kg) and CAD (13.15–49.91 mg/kg) were the dominant BAs in all marinated pork meat samples (except pork ham muscle samples marinated with L. sakei and P. acidilactici, and M. longissimus dorsi meat samples marinated with P. acidilactici and P. pentosaceus). The highest content of PUT in M. longissimus dorsi samples marinated with L. sakei (131.29 ± 0.35 mg/kg) was found. The lowest content of PUT in nonmarinated pork ham muscle samples (14.56 ± 0.11 mg/kg) was measured. A weak negative significant correlation between the PUT and HIS (r = 0.381; p = 0.003), a weak negative significant correlation between the PUT and spermidine (SPD) (r = -0.274; p = 0.034), and a weak negative significant correlation between the PUT and pH of meat samples (r = -0.255; p = 0.050) were found.

The concentration of HIS in meat samples ranged from 15.09 ± 0.12 mg/kg and 15.47 ± 0.11 (in pork ham muscle samples with P. acidilactici marinated and in M. longissimus dorsi samples marinated with P. pentosaceus, respectively) to 1.37 ± 0.03 mg/kg (in pork neck samples marinated with P. pentosaceus). HIS was not found in pork shoulder, M. longissimus dorsi, and ham muscle samples marinated with L. sakei and P. pentosaceus. A weak positive significant correlation between the HIS and PHE content (r = 0.313; p = 0.015), a weak negative significant correlation between the HIS and PUT content (r = -0.381; p = 0.003), a moderate negative significant correlation between the HIS and CAD content (r = -0.405; p = 0.001) were found.

The TYR content in meat samples ranged from 1.03 ± 0.01 mg/kg (in M. longissimus dorsi samples marinated with L. sakei) to 11.12 ± 0.1 mg/kg (in ham muscle samples marinated with P. pentosaceus). A moderate positive significant correlation between the TYR and CAD content (r = 0.407; p = 0.001), a weak negative significant correlation between the TYR and spermine (SPR) content (r = -0.362; p = 0.004), and a moderate negative

71

significant correlation between the TYR content and the pH of meat (r = -0.476; p = 0.0001) were found.

Table 3.2.1.3. BAs content (mg/kg) in pork meat, marinated with selected LAB strains cultivated in potato juice.

Samples Pa Ls Pp Control

Neck PHE 21.09±0.14c 16.47±0.12b 12.27±0.10b 7.63±0.07a PUT 27.28±0.14a 31.04±0.15d 61.63±0.31f 23.41±0.12a CAD 22.64±0.12b 20.19±0.11a 19.94±0.10a 18.16±0.09a HIS 3.45±0.05b 7.36±0.07d 1.37±0.03a 5.34±0.07c TYR 2.13±0.02c 4.15±0.03b 1.18±0.01a - SPD 5.17±0.06c - 6.24±0.07c 1.04±0.03a SPR 6.77±0.07b 10.74±0.08c - 2.78±0.03a TRP - 9.10±0.08c 2.74±0.04a 3.21±0.04b Total 88.53b 99.05c 105.37d 61.57a

Shoulder

PHE 7.59±0.07b 4.32±0.01a 10.51±0.08c 8.50±0.08b PUT 30.68±0.20d 19.57±0.13a 32.07±0.20d 44.79±0.25e CAD 33.62±0.15c 28.99±0.13b 19.57±0.10a 31.96±0.17b HIS 2.04±0.01a - 6.11±0.10b 8.71±0.08b TYR 9.74±0.09b 7.44±0.06b 3.11±0.05a 0.55±0.01a SPD 9.71±0.08c 3.21±0.04a - 7.02±0.06b SPR 3.25±0.04a - 15.24±0.10c - TRP - 4.56±0.05b 2.12±0.02a - Total 96.63c 68.09a 88.73b 101.53d

Ham PUT - 37.16±0.20c 75.53±0.34e 14.56±0.11a CAD 13.15±0.09a - 49.91±0.24d 40.14±0.30c HIS 15.09±0.12c 6.02±0.05b 11.2±0.10a 3.15±0.02a TYR 3.38±0.02a 5.26±0.04a 11.12±0.1b - SPD 5.69±0.04b 2.47±0.01a - 3.36±0.04a SPR 1.07±0.01a 2.14±0.02b 0.97±0.01a - TRP - 0.74±0.01a - 1.02±0.01b Total 38.38a 53.79b 148.73e 62.23c

M. longissimus dorsi

PHE 12.35±0.11b - 37.49±0.14e 7.14±0.05a PUT 65.18±0.28d 131.29±0.35f 42.80±0.15c 26.82±0.15a CAD - 20.13±0.12b - 14.19±0.07a HIS 11.24±0.08a - 15.47±0.11b 9.07±0.09a TYR 5.44±0.04b 1.03±0.01a 5.17±0.03b 3.24±0.02a SPD 1.06±0.01a - - 1.22±0.01a SPR 3.12±0.03a 6.59±0.04c 2.33±0.03a 4.17±0.04b TRP - 3.47±0.03b 2.11±0.01a - Total 98.39c 162.52e 105.37c 65.85a

72

Continue of Table 3.2.1.3.

Samples Pa Ls Pp Control Loin

PHE 21.47±0.14c 34.94±0.20d 27.64±0.12c 7.74±0.08a PUT 33.64±0.16d 23.84±0.14b 18.80±0.12a 24.0±0.18b CAD 19.34±0.06a 28.39±0.14b 41.65±0.16c 33.71±0.19b HIS 3.14±0.01a 5.17±0.02a 13.26±0.10c 9.09±0.09b TYR 2.71±0.01a 5.30±0.05b 10.2±0.08c 7.14±0.05b SPD 5.44±0.04b 2.84±0.02a - 3.84±0.02b SPR 8.06±0.07b 2.13±0.01a 3.03±0.02a 0.65±0.01a Total 93.80b 102.51c 114.58d 86.17a Data are the mean ± SD (n = 3); SD – standard deviation; Mean values within column with different letters are significantly different (p < 0.05); Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine

SPD was not found in pork neck samples treated with L. sakei marinade,

in pork shoulder and ham muscle samples treated with P. pentosaceus marinade, in M. longissimus dorsi samples treated with L. sakei and P. pentosaceus marinades, and in loin samples treated with the marinade based on P. pentosaceus (Table 3.2.1.3). The highest concentration of SPD was found in pork shoulder samples marinated with the P. acidilactici marinade (9.71 ± 0.08 mg/kg). A weak negative significant correlation between the SPD and PUT content (r = -0.274; p = 0.034), a moderate negative significant correlation between the SPD and SPR content (r = -0.405; p = 0.001), and a weak negative significant correlation between the SPD and TRP content (r = -0.362; p = 0.004) were found.

The SPR content in pork meat varied from 0 mg/kg (in pork neck samples treated with P. pentosaceus and L. sakei marinade, in pork loin samples treated with P. pentosaceus marinade) to 15.24 ± 0.10 mg/kg (in pork shoulder samples treated with P. pentosaceus marinade). A weak negative significant correlation between the SPR and CAD content (r = -0.261; p = 0.044), a weak negative significant correlation between the SPR and TYR content (r = -0.362; p = 0.004), a moderate negative significant correlation between the SPR and SPD content (r = -0.405; p = 0.001), and a weak negative significant correlation between the SPR and meat moisture content (r = -0.325; p = 0.011) were found.

TRP in pork meat samples marinated with P. acidilactici marinade was not found, and the highest concentration of TRP in pork neck samples treated with L. sakei marinade (9.10 ± 0.08 mg/kg) was established. A weak negative significant correlation between the TRP and SPD content (r = -0.280; p = 0.030), a weak negative significant correlation between the TRP

73

and meat moisture content (r = -0.325; p = 0.011), and a weak negative significant correlation between the TRP and meat pH (r = -0.307; p = 0.017) were found. The total BAs content in marinated pork neck ranged from 61.57 mg/kg to 105.37 mg/kg (in control samples and in meat samples treated with P. pentosaceus marinade, respectively). In pork shoulder, the total BAs content ranged from 68.09 mg/kg to 101.53 mg/kg (in meat marinated with L. sakei marinade and in control samples, respectively), in ham muscle from 38.38 mg/kg to 148.73 mg/kg (respectively, in meat samples marinated with P. acidilactici and with P. pentosaceus), in M. longissimus dorsi from 65.85 mg/kg to 162.52 mg/kg (in control samples and in meat samples treated with P. pentosaceus marinade, respectively) and in loin samples from 86.17 mg/kg to 114.58 mg/kg (in control samples and in meat samples treated with P. pentosaceus marinade, respectively).

The overall acceptability of pork neck, shoulder, ham, M.

longissimus dorsi, and loin marinated with P. acidilactici KTU05-7, P.

pentosaceus KTU05-8, and L. sakei KTU05-6 strains cultivated in potato juice. The overall acceptability of the pork meat, marinated with selected LAB cultivated in potato juice is presented in Figure 3.2.1.5. A significant different (p ≤ 0.05) acceptability between control samples and marinated meat samples was found. The results indicated that the overall acceptability of marinated meat samples depended on the LAB strain used for marinade production (p ≤ 0.0001) and on the part of pork meat (p ≤ 0.0001).

Figure 3.2.1.5. The overall acceptability of the pork neck, shoulder, ham, M. longissimus dorsi, and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato

juice (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, Mld - M. longissimus dorsi).

0

2

4

6

Neck Shoulder Ham Mld LoinScor

es (

min

0;

max

6)

Pa Ls Pp Control

74

Samples treated with marinade based on P. pentosaceus were indicated to have the highest acceptability (5.6–5.9 scores), except pork neck which was indicated to have highest acceptability after treatment with L. sakei marinade (5.6 scores). The lowest acceptability was found in pork meat samples treated with L. sakei marinade (pork shoulder, ham, M. longissimus dorsi, and pork loin, 4.8, 4.9, 5.4 and 4.8 scores, respectively). The overall acceptability of pork neck muscle samples ranged from 4.5 to 5.6 (control samples and samples treated with L. sakei marinade, respectively). Also, the overall acceptability depended on the part of pork: the most acceptable were M. longissimus dorsi samples treated with all tested LAB (scores ranged from 5.5 to 5.8). A weak negative significant correlation was found between the overall acceptability and the IF of meat samples (r = -0.394; p = 0.002).

Parameters of beef neck, shoulder, ham, M. longissimus dorsi and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice. Parameters of beef meat marinated with selected LAB cultivated in potato juice are presented in Table 3.2.1.4. It was found that the quality parameters of 24-hour-marinated beef meat samples significantly depended on the LAB strain used for marinade production (p ≤ 0.0001), and on the part of beef meat (p ≤ 0.0001).

Table 3.2.1.4. Quality parameters of beef meat, marinated with selected LAB strains cultivated in potato juice.

S Moisture, %

pH Drip loss, %

WHC, %

Cooking loss, %

Shear force,

kg/cm2

IF, %

Neck

Pa 18.66 ±0.13a

5.03 ±0.01a

2.14 ±0.3b

61.43 ±0.16a

50.5 ±0.25c

3.82 ±0.1b

3.37 ±0.01a

Pp 19.51 ±0.11b

5.26 ±0.01b

1.86 ±0.2a

61.44 ±0.11a

49.0 ±0.28c

4.64 ±0.4c

3.70 ±0.04b

Ls 19.36 ±0.14b

5.11 ±0.03a

2.07 ±0.1b

62.85 ±0.14b

44.6 ±0.36a

2.68 ±0.2a

3.82 ±0.03b

C 20.79 ±0.12c

5.96 ±0.02c

2.55 ±0.3c

62.92 ±0.15b

46.8 ±0.41b

3.57 ±0.3b

3.37 ±0.03a

Shoulder

Pa 19.29 ±0.15b

5.20 ±0.02b

1.84 ±0.1a

62.78 ±0.12c

47.5 ±0.43b

4.43 ±0.3b

1.39 ±0.02b

Pp 18.75 ±0.11a

4.94 ±0.01a

2.12 ±0.3b

62.13 ±0.11b

46.7 ±0.36a

5.39 ±0.2c

1.23 ±0.02a

Ls 18.45 ±0.12a

5.05 ±0.03a

1.92 ±0.2a

61.36 ±0.14a

49.4 ±0.31d

5.21 ±0.1c

1.37 ±0.01b

C 19.80 ±0.13c

5.72 ±0.05c

2.63 ±0.1c

62.21 ±0.12b

48.4 ±0.22c

2.51 ±0.2a

1.59 ±0.01c

75

Continue of Table 3.2.1.4.

S Moisture, %

pH Drip loss, %

WHC, %

Cooking loss, %

Shear force,

kg/cm2

IF, %

Ham muscle

Pa 21.97 ±0.11b

5.98 ±0.04c

2.17 ±0.3a

61.76 ±0.12a

49.7 ±0.31b

5.91 ±0.1b

1.60 ±0.03b

Pp 21.30 ±0.12b

5.38 ±0.03a

2.30 ±0.1b

62.28 ±0.13b

48.2 ±0.40a

4.05 ±0.2a

1.50 ±0.02a

Ls 20.24 ±0.14a

5.75 ±0.02b

1.94 ±0.1a

63.02 ±0.15c

51.70 ±0.28b

5.85 ±0.4c

1.77 ±0.01c

C 24.32 ±0.13c

6.57 ±0.04a

2.41 ±0.2c

61.89 ±0.11a

53.56 ±0.25c

5.91 ±0.6b

1.48 ±0.04a

M. longissimus dorsi

Pa 20.45 ±0.09a

5.23 ±0.03b

1.76 ±0.1a

63.14 ±0.16c

45.8 ±0.26c

3.87 ±0.2c

1.32 ±0.01c

Pp 21.10 ±0.10b

5.04 ±0.02a

1.96 ±0.2a

62.39 ±0.16b

45.1 ±0.30b

2.59 ±0.1a

0.95 ±0.03a

Ls 20.12 ±0.12a

5.02 ±0.01a

1.88 ±0.4a

62.34 ±0.14b

43.3 ±0.24a

2.60 ±0.1a

0.83 ±0.02a

C 21.25 ±0.11b

5.75 ±0.04c

2.04 ±0.3b

61.94 ±0.11a

42.7 ±0.36a

3.08 ±0.2b

1.13 ±0.02b

Loin

Pa 18.82 ±0.11a

5.91 ±0.02b

1.98 ±0.2a

81.18 ±0.24b

45.6 ±0.28b

3.31 ±0.1a

1.51 ±0.02a

Pp 19.38 ±0.12b

5.44 ±0.04a

1.86 ±0.1a

80.34 ±0.19b

47.4 ±0.33c

5.45 ±0.3d

1.60 ±0.04a

Ls 18.55 ±0.09a

6.02 ±0.02b

2.14 ±0.1b

81.45 ±0.18c

46.5 ±0.29b

4.37 ±0.3c

1.82 ±0.01b

C 22.46 ±0.13c

6.87 ±0.04c

2.25 ±0.5c

77.54 ±0.20a

42.8 ±0.24a

3.86 ±0.1b

2.23 ±0.03c

Data are the mean ± SD (n = 3). Mean values within column with different letters are significantly different (p < 0.05). Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; WHC – water holding capacity; IF – intramuscular fat, S – samples

The treatment of beef neck, shoulder, ham muscle, M. longissimus dorsi

and loin samples with all marinades (P. acidilactici, P. pentosaceus and L. sakei) lowered the moisture content in meat on average by 7.76%, 4.89%, 12.95%, 3.26%, 15.77%, respectively, as compared with control samples.

P. acidilactici, P. pentosaceus, and L. sakei marinades lowered the pH of meat on average by 13.53% (neck and ham muscle samples) and by 11.42% (shoulder and M. longissimus dorsi samples) as compared with the control samples (pH 5.96–5.75). The highest pH decrease (on average by 15.72%) was found after the marination of beef loin (with all marinades), and it was ranged from 5.44 ± 0.04 (beef loin treated with P. pentosaceus marinade) to

76

6.02 ± 0.02 (beef loin treated with L. sakei marinade). A strong positive significant correlation between the meat sample pH and meat sample moisture content (r = 0.635; p = 0.0001), a moderate positive significant correlation between the meat sample pH and meat sample drip loss (r = 0.517; p = 0.0001) and a moderate positive significant correlation between the meat sample pH and meat sample WHC (r = 0.438; p = 0.0001) were found. In all cases, marinated beef meat samples had from 8.49% to 25.47% (in (M. longissimus dorsi and beef shoulder muscle, respectively) higher drip loss, and its ranged from 1.84 ± 0.01% (in M. longissimus dorsi marinated with P. acidilactici) to 2.63 ± 0.01% (in beef shoulder control samples). A weak positive significant correlation between the meat sample drip loss and moisture content (r = 0.393; p = 0.002), a moderate positive significant correlation between the meat sample drip loss and pH (r = 0.517; p = 0.0001) were found.

The treatment of beef meat with all used marinades lowered the WHC of the neck muscle on average by 1.57%, of the shoulder muscle on average by 0.19%, and increased the WHC of the ham muscle on average by 0.75%, of M. longissimus dorsi by 1.10%, and of loin by 4.45% as compared with the control samples. In marinated beef meat, WHC ranged from 61.36 ± 0.14 (in beef shoulder treated with the L. sakei marinade) to 81.45 ± 0.18 (in beef loin treated with the L. sakei marinade). A moderate positive significant correlation between the meat samples pH and WHC (r = 0.438; p = 0.002) and a weak negative significant correlation between the meat samples drip loss and WHC (r = -0.297; p = 0.021) were found.

Treatment with P. acidilactici, P. pentosaceus and L. sakei marinades increased the cooking loss of the neck, M. longissimus dorsi and loin samples on average by 4.42%, 4.64% and 8.73%, respectively. Different tendencies were estimated for the neck and ham muscle cooking loss: in this case, marination decreased the cooking loss on average by 1.11% and 6.87%, respectively. A weak negative significant correlation between the meat sample drip loss and WHC (r = -0.297; p = 0.021) and a strong positive significant correlation between the meat sample cooking loss and tenderness (r = 0.682; p = 0.0001) were found.

The highest tenderness of the beef samples treated with the marinade based on L. sakei was found, except beef shoulder muscle whose shear force decreased on average by 7.08%. Treatment with P. acidilactici and P. pentosaceus marinades increased the shear force of meat (on average by 14.03%), and decreased tenderness as compared with control samples. A strong positive significant correlation between the meat sample tenderness and the cooking loss (r = 0.682; p = 0.0001) was found. The highest IF content in beef neck muscle samples marinated with L. sakei marinade (3.82

77

± 0.03%) and the lowest in M. longissimus dorsi marinated with L. sakei marinade (0.83 ± 0.02%) were found.

Colour parameters. The colour parameters (L*, a* and b*) of beef neck,

shoulder, ham, M. longissimus dorsi and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice are presented in Table 3.2.1.5. It was found that lighteness (L*) increased in all five treated groups (p < 0.05) of beef meat. Changes of meat sample colour parameters (L*, a* and b*) depended on the LAB strain used for marinade production (p ≤ 0.0001) and on the part of pork meat (p ≤ 0.0001).

Table 3.2.1.5. Colour parameters (L*, a* and b*) of beef meat, marinated with selected LAB strains cultivated in potato juice.

Samples L* a* b* Beef neck

Pa 42.06±0.20c 10.46±0.20b 4.45±0.03b Pp 41.02±0.20b 7.26±0.19a 2.84±0.02a Ls 36.86±0.16a 8.31±0.20a 4.71±0.05b Control 38.16±0.17a 18.90±0.29d 4.85±0.05c

Beef shoulder Pa 45.04±0.20b 13.99±0.21a 8.18±0.05c Pp 48.56±0.18c 16.15±0.19b 7.52±0.03b Ls 43.42±0.19a 13.91±0.31a 7.36±0.10b Control 44.61±0.20b 19.48±0.32c 6.47±0.09a

Beef ham muscle Pa 31.05±0.15a 12.84±0.19a 4.96±0.04b Pp 34.32±0.20b 13.17±0.18b 3.85±0.03a Ls 43.06±0.20d 12.57±0.26a 6.08±0.09c Control 30.69±0.15a 15.79±0.31c 3.84±0.05a

M. longissimus dorsi Pa 40.24±0.20c 15.19±0.26b 8.25±0.05d Pp 38.23±0.19b 11.69±0.15a 7.04±0.08c Ls 36.92±0.16a 15.87±0.26c 5.90±0.08b Control 38.03±0.16b 15.16±0.26b 2.25±0.03a

Beef loin Pa 30.79±0.15a 15.31±0.26c 4.81±0.03c Pp 41.02±0.23d 7.26±0.12a 2.84±0.02a Ls 33.62±0.16b 11.12±0.28b 4.17±0.06b Control 29.89±0.14a 14.12±0.26b 2.93±0.01a Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; L* − lightness; a* − redness (or –a* of greenness); and b* − yellowness (or –b* of blueness)

78

Marination increased the (L*) values of beef meat samples (from 1.14% of M. longissimus dorsi to 17.58% of beef loin) compared with control samples, and they ranged from 29.89 ± 0.14 (control beef loin samples) to 48.56 ± 0.18 (beef shoulder muscle samples treated with P. pentosaceus marinade). A moderate positive significant correlation between the meat lighteness (L*) and yelowness (b*) (r = 0.539; p = 0.0001), and a strong negative significant correlation between the meat lighteness (L*) and pH (r = -0.679; p = 0.0001) was found.

Marination decreased the (a*) values of beef meat (marination with P. acidilactici marinade by 16.58%, marination with P. pentosaceus marinade by 33.48% and marination with L. sakei marinade by 24.32%) as compared with the control samples. The values of (a*) ranged from 7.26 ± 0.19 (in P. pentosaceus marinated beef loin and neck muscle samples) to 19.48 ± 0.32 (of control beef shoulder muscle samples). A weak positive significant correlation between the meat redness (a*) and yelowness (b*) (r = 0.370; p = 0.004) and between the meat redness (a*) and meat pH (r = 0.271; p = 0.036) was found.Marination decreased the (b*) values of the beef neck mucle on average by 17.53%: however, it increased the (b*) values of beef shoulder (18.80%), ham muscle (4.96%), M. longissimus dorsi (213.93%), and loin (34.47%) as compared with the control samples. The highest yelowness in M. longissimus dorsi samples (213.93%) was found. A moderate positive significant correlation between the meat yellowness (b*) and lighteness (L*) (r = 0.539; p = 0.0001), a weak positive significant correlation between the meat yellowness (b*) and redness (a*) (r = 0.370; p = 0.004), and a moderate negative significant correlation between the meat yellowness (b*) and pH (r = -0.496; p = 0.0001) was found.

The content of BAs in beef neck, shoulder, ham, M. longissimus dorsi

and loin marinated with the P. acidilactici KTU05-7, P. pentosaceus KTU05-8, and L. sakei KTU05-6 strains cultivated in potato juice. The content of BAs content is presented in Table 3.2.1.5. The LAB strain used for marinade production and the part of beef meat have a significant influence on the BAs formation (p ≤ 0.0001) in meat samples. In all the cases, marination increased the PHE content in beef (up to 11.27–44.94 mg/kg). A weak negative significant correlation between the PHE content and meat pH (r = -0.304; p = 0.018) was found. The PUT and CAD were the dominant BAs in all marinated beef meat samples. The PUT content ranged from 12.27 ± 0.14 mg/kg (in beef neck muscle marinated with P. pentosaceus) to 75.53 ± 0.62 mg/kg (in beef ham muscle marinated with P. pentosaceus), and CAD ranged from 13.15±0.15 mg/kg (in beef ham muscle marinated with P. acidilactici) to 65.71 ± 0.66 mg/kg (in beef ham

79

muscle marinated with L. sakei). A moderate positive significant correlation between the PUT and CAD (r = 0.554; p = 0.0001), between the PUT and HIS (r = 0.504; p = 0.0001), a weak positive significant correlation between the PUT and TYR (r = 0.351; p = 0.006), a strong positive significant correlation between the PUT and SPD (r = 0.618; p = 0.0001) and a weak negative significant correlation between the PUT and meat pH (r = -0.284; p = 0.028), as well as a moderate positive significant correlation between the CAD and SPD (r = 0.591; p = 0.0001) were found.The concentration of HIS in meat samples ranged from 1.75 ± 0.01 mg/kg to11.34 ± 0.15 (in beef neck muscle samples marinated with L. sakei and in M. longissimus dorsi samples marinated with P. pentosaceus, respectively). The HIS in beef ham marinated with P. acidilactici was not found. A moderate positive significant correlation between the HIS and PUT content (r = 0.504; p = 0.0001), between the HIS and TYR content (r = 0.410; p = 0.001) and a moderate negative significant correlation between the HIS and SPR content (r = -0.555; p = 0.0001) were found.

The TYR was the dominant BA in marinated meat, and its content ranged from 0.87 ± 0.02 mg/kg (in control beef shoulder samples) to 5.63 ± 0.09mg/kg (in M. longissimus dorsi samples marinated with P. acidilactici), except beef neck samples marinated with pediococci, beef shoulder marinated with P. pentosaceus, and control M. longissimus dorsi samples. A weak positive significant correlation between the TYR and PUT content (r = 0.351; p = 0.0006), a moderate positive significant correlation between the TYR and HIS content (r = 0.410; p = 0.001), and a weak negative significant correlation between the TYR content and SPD (r = -0.282; p = 0.029) were found.SPD was not found in beef neck samples marinated with P. pentosaceus and in beef shoulder control samples. The highest concentration of SPD was found in beef ham muscle samples marinated with P. pentosaceus marinade (5.47 ± 0.03 mg/kg). A strong positive significant correlation between the SPD and PUT content (r = 0.618; p = 0.0001), between the SPD and CAD content (r = 591; p = 0.0001), and a moderate positive significant correlation between the SPD and SPR content (r = 0.433; p = 0.001) were found.

The SPR content in beef meat samples varied from 0 mg/kg (in control samples of beef neck and loin muscles, in M. longissimus dorsi samples treated with P. acidilactici marinade, and in M. longissimus dorsi samples treated with L. sakei marinade) to 7.61 ± 0.08 mg/kg (in beef shoulder samples treated with L. sakei marinade). A strong negative significant correlation between the SPR and HIS content (r = -0.555; p = 0.0001), a weak negative significant correlation between the SPR and TYR content (r

80

= -0.282; p = 0.029), a moderate positive significant correlation between the SPR and SPD content (r = 0.433; p = 0.001) were found.

Table 3.2.1.5. BAs content (mg/kg) in beef meat, marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice.

Samples Pa Ls Pp Control Neck

PHE 35.09±0.37e 15.78±0.15c 12.27±0.15c 7.63±0.09a PUT 37.09±0.40e 19.51±0.19b 12.27±0.14a 23.41±0.20c CAD 22.64±0.25c 14.30±0.16a 19.94±0.20b 18.16±0.15b HIS 5.17±0.08d 1.75±0.01a 2.95±0.03a 3.22±0.05c TYR - 5.44±0.06b - 1.31±0.02a SPD 1.84±0.03c 1.12±0.01b - 0.58±0.01a SPR 1.44±0.02b 1.36±0.02b 0.91±0.01a - TRP - - - 0.47±0.01a Total 103.27e 59.26b 48.34a 54.78b

Shoulder PHE 40.11±0.45c 29.74±0.35a 38.15±0.39b 25.77±0.30a PUT 33.12±0.35d 28.69±0.29c 31.47±0.33c 13.58±0.15a CAD 20.17±0.19b 18.43±0.19a 22.76±0.22c 18.19±0.20a HIS 3.89±0.05c 2.91±0.03b 2.48±0.03b 1.86±0.02a TYR 1.57±0.03b 2.55±0.05c - 0.87±0.02a SPD 2.79±0.03b 2.33±0.03a 1.86±0.02a - SPR 1.28±0.01a 7.61±0.08c 5.11±0.05b 1.39±0.02a TRP 0.75±0.02b - 0.51±0.01a 0.74±0.01b Total 103.68d 92.26c 102.34d 62.40a

Ham PHE 21.17±0.23c 18.69±0.15b 11.27±0.14a 9.66±0.10a PUT 19.33±0.25a 37.16±0.44c 75.53±0.62e 14.56±0.15a CAD 13.15±0.15a 65.71±0.66d 45.91±0.39c 40.14±0.43c HIS - 5.31±0.05c 4.89±0.06b 2.17±0.02a TYR 1.74±0.01a 2.05±0.02c 1.98±0.02b 1.55±0.01a SPD 3.47±0.04b 4.71±0.03c 5.47±0.03d 2.38±0.01a SPR 6.17±0.05d 2.95±0.04a 4.76±0.05c 3.99±0.05b TRP 1.33±0.01b - - 0.85±0.01a Total 66.36a 136.58c 145.05d 75.30a

M. longissimus dorsi PHE 12.35±0.15a 14.60±0.16b 37.49±0.29d 7.14±0.08a PUT 65.18±0.59d 51.29±0.55c 42.80±0.46b 26.82±0.28a CAD 36.17±0.39d 47.34±0.45e 25.74±0.20c 14.19±0.15a HIS 10.38±0.12b 9.68±0.10a 11.34±0.15c 10.46±0.14b TYR 5.63±0.09c 4.74±0.05b 2.11±0.03a - SPD 2.33±0.03b 1.97±0.04a 2.76±0.03c 1.98±0.02a SPR - 1.12±0.01a 1.45±0.01b 0.85±0.01a TRP 0.96±0.01b 0.87±0.02b - 0.51±0.01a Total 133.0d 131.61e 123.69d 61.95a

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Continue of Table 3.2.1.5. Samples Pa Ls Pp Control

Loin PHE 31.47±0.30c 44.94±0.45e 41.93±0.40d 17.74±0.15a PUT 23.64±0.19a 33.84±0.36c 38.80±0.40d 24.0±0.19a CAD 18.65±0.15a 20.17±0.20b 25.45±0.26c 15.38±0.15a HIS 8.24±0.08b 9.48±0.10c 8.36±0.09b 7.64±0.06a TYR 3.17±0.03a 5.12±0.05c 4.87±0.05b 3.10±0.03a SPD 2.55±0.02b 2.61±0.03c 2.58±0.03b 1.02±0.01a SPR 1.04±0.01b - 0.93±0.01a - TRP - - 1.53±0.02a - Total 88.76b 116.16a 124.45e 66.88a Data are the mean ± SD (n = 3); SD – standard deviation; Mean values within column with different letters are significantly different (p < 0.05); Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine

TRP in beef meat samples marinated with L. sakei marinade (except in M. longissimus dorsi) was not found, and the highest concentration of TRP (1.53 ± 0.02 mg/kg) was established in beef loin samples treated with P. pentosaceus marinade. The total BAs content in marinated beef neck ranged from 48.34 mg/kg to 105.37 mg/kg (in meat samples marinated with P. pentosaceus and with P. acidilactici, respectively), in beef shoulder it ranged from 62.40 mg/kg to 103.68 mg/kg (in control samples and in meat samples marinated with P. acidilactici, respectively), in ham muscle samples from 66.36 mg/kg to 145.05 mg/kg (in meat samples marinated with P. acidilactici and with P. pentosaceus, respectively), in M. longissimus dorsi from 61.95 mg/kg to 133.0 mg/kg (in control samples and in meat samples marinated with P. acidilactici, respectively), and in loin samples from 66.88 mg/kg to 124.45 mg/kg (in control samples and in meat samples marinated with P. pentosaceus, respectively).

The overall acceptability of beef neck, shoulder, ham, M. longissimus

dorsi and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8, and L. sakei KTU05-6 strains cultivated in potato juice. The data on the overall acceptability of the marinated beef meat are presented in Figure 3.2.1.6. Significant differences (p ≤ 0.05) in the overall acceptability were found between the control samples and the marinated meat samples. The results indicated the overall acceptability of marinated beef meat samples to depend on the LAB strain used for marinade production (p ≤ 0.0001) and on the part of beef meat (p ≤ 0.0001). Samples treated with the marinade based on P. acidilactici were indicated to have the highest

82

acceptability which ranged from 5.2 ± 0.05 (beef shoulder muscle samples) to 5.7 ± 0.05 scores (M. longissimus dorsi samples), except M. longissimus dorsi samples which were indicated to have highest overall acceptability after treatment with L. sakei marinade (5.8 ± 0.06 scores).

Figure 3.2.1.6. The overall acceptability of the beef neck, shoulder, ham, M. longissimus dorsi and loin marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato

juice (Remarks: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, M. longissimus dorsi - Mld).

The lowest overall acceptability of control beef neck and shoulder

muscle samples (4.7 ± 0.02 and 4.8 ± 0.04, respectively) and of beef shoulder samples treated with L. sakei marinade (4.8 ± 0.03) was found. The overall acceptability depended on the part of beef meat, and the most acceptable were M. longissimus dorsi samples treated with all tested marinades (scores ranged from 5.4 ± 0.04 to 5.8 ± 0.06). A weak negative significant correlation between the overall acceptability and IF of meat samples (r = -0.325; p = 0.011) was found.

Parameters of organic and conventionally produced chicken meat,

marinated with P. acidilactici KTU05-7 cultivated in potato juice. The technological parameters of organic and conventionally produced chicken breast, drumsticks and thighs are presented in Table 3.2.1.6. The quality parameters of marinated chicken meat samples significantly depended on the part of chicken meat (p ≤ 0.0001) and on the chicken meat production method (organic or conventional) (p ≤ 0.0001). The treatment of organic chicken breast, drumstick, and thigh samples with P. acidilactici marinade increased the meat moisture content (by 0.8% as compared with conventionally produced chicken meat). A strong positive significant correlation between the moisture content and WHC (r = 0.767; p = 0.0001), between the samples content and tenderness (r = 0.787; p = 0.0001), a

0

12

34

56

Neck Shoulder Ham Mld LoinScor

es (

min

. 0;

max

6

Pa Ls Pp Control

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strong negative significant correlation between the moisture content and cooking loss (r = -0.770; p = 0.0001) and a moderate positive significant correlation between the moisture content and IF of the meat samples (r = 0.525; p = 0.001) were found.

Table 3.2.1.6. Quality parameters of organic and conventionally produced chicken breasts, drumsticks and, thighs samples.

Meat sample

Moisture, %

pH Drip loss, %

WHC, %

Cooking loss, %

Shear force,

kg/cm2

IF, %

Breast

O Pa 26.9

±0.02b 5.20

±0.01a 1.03

±0.01c 63.01 ±0.09c

9.25 ±0.04a

2.32 ±0.03c

0.68 ±0.02b

C Pa 26.1

±0.04a 5.17

±0.01a 0.98

±0.02b 62.74 ±0.07b

12.86 ±0.05c

1.86 ±0.01b

0.63 ±0.01b

O 26.7

±0.05b 5.76

±0.02b 0.94

±0.02a 62.93 ±0.12b

9.04 ±0.03a

2.04 ±0.01b

0.60 ±0.01a

C 25.9

±0.08a 5.83

±0.02c 0.92

±0.01a 62.25 ±0.13a

12.21 ±0.07b

1.70 ±0.03a

0.58 ±0.01a

Drumsticks

O Pa 27.4

±0.05b 5.12

±0.01a 0.87

±0.01a 63.25 ±0.02b

9.41 ±0.03a

2.14 ±0.07d

0.76 ±0.03b

C Pa 26.1

±0.01a 5.37

±0.02b 0.93

±0.02b 62.78 ±0.20a

13.62 ±0.03c

1.69 ±0.04b

0.73 ±0.04a

O 27.2

±0.04b 5.84

±0.03c 0.89

±0.01a 63.01 ±0.25b

9.32 ±0.03a

1.98 ±0.02c

0.74 ±0.04a

C 26.0

±0.03a 6.01

±0.04c 0.91

±0.01b 62.69 ±0.12a

13.34 ±0.04b

1.34 ±0.01a

0.69 ±0.02a

Thighs

O Pa 27.6

±0.03b 5.33

±0.02a 1.12

±0.02c 63.04 ±0.17c

9.14 ±0.01a

2.40 ±0.01c

0.91 ±0.05b

C Pa 26.7

±0.02a 5.28

±0.01a 0.93

±0.01b 62.89 ±0.09a

14.31 ±0.03b

1.62 ±0.01a

0.93 ±0.03b

O 27.4

±0.02b 6.14

±0.03c 0.86

±0.01a 62.98 ±0.10b

8.91 ±0.04a

2.21 ±0.01c

0.86 ±0.02a

C 26.4

±0.06a 6.09

±0.02b 0.91

±0.02b 62.83 ±0.11a

14.05 ±0.04b

1.43 ±0.03a

0.89 ±0.02a

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). Pa – P. acidilactici; WHC – water holding capacity; IF – intramuscular fat, O – organic, C – conventionally

In all the cases, marination with the P. acidilactici marinade decreased

the pH of chicken meat samples. A decrease of pH in organic chicken breast (reduced by 9.72%), drumstick (reduced by 12.33%) and thigh (reduced by 13.19%) on average by 11.75% was found. Similar results in conventionally produced chicken breast samples (meat pH reduced by 11.32%) were

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estimated. The analysis of individual chicken meat parts showed the following trends of pH reduction: the drumstick pH reduced by 10.65% and thighs meat pH by 13.30%. A moderate negative significant correlation between the meat sample pH and drip loss (r = -0.469; p = 0.005) and a weak negative significant correlation between the meat sample pH and tenderness (r = -0.380; p = 0.022) were found.

In all the cases, organic and conventionally produced chicken meat samples marinated with P. acidilactici had from 2.20% (conventionally produced thighs and drumsticks) to 30.23% (organic thighs) higher dirp loss, and it ranged from 0.86 ± 0.01% (in organic thighs) to 1.12 ± 0.02% (in with P. acidilactici marinated organic thighs). A moderate negative significant correlation between the meat sample drip loss and pH (r = -0.469; p = 0.004) and a moderate positive significant correlation between the meat sample drip loss and tenderness (r = 0.438; p = 0.008) were found.

Marination with P. acidilactici increased the WHC of organic chicken meat on average by 0.08% and of conventionally produced chicken meat on average by 0.49%. The highest increase of WHC (0.38%) in organic chicken drumstick samples and the lowest increase of WHC (0.1%) in organic and conventionally produced chicken thigh samples were found as compared with control samples. A strong positive significant correlation between the meat samples WHC and moisture content (r = 0.767; p = 0.0001), a moderate negative significant correlation between the meat WHC and cooking loss (r = -0.500; p = 0.002), a moderate positive significant correlation between the meat WHC and tenderness (r = 0.551; p = 0.0001), and a moderate positive significant correlation between the meat WHC and IF (r = 0.487; p = 0.003) were found.

Organic chicken meat samples had a lower cooking loss (on average by 3.17%), as compared with conventionally produced chicken meat samples. Marination increased the cooking loss of organic and conventionally produced chicken meat on average by 0.19% and 0.65%, respectively. A strong negative significant correlation between the meat cooking loss and the moisture content (r = -0.770; p = 0.0001), a moderate negative significant correlation between the meat cooking loss and WHC (r = -0.500; p = 0.002) and a very strong negative significant correlation between the meat cooking loss and tenderness (r = -0.884; p = 0.0001) were found.

Marination with P. acidilactici increased the tenderness of organic and conventionally produced chicken meat (except conventionally produced chicken meat drumstick samples) from 8.08% (organic chicken drumstick samples) to 13.73% (organic chicken meat breast samples) as compared with the control samples. A strong positive significant correlation between the meat sample tenderness and moisture content (r = 0.787; p = 0.0001), a

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weak negative significant correlation between the meat sample tenderness and pH (r = -0.380; p = 0.022), a moderate positive significant correlation between the meat tenderness and WHC (r = 0.551; p = 0.0001) and between the meat sample tenderness and drip loss (r = 0.438; p = 0.008) and a very strong negative significant correlation between the meat tenderness and cooking loss (r = -0.884; p = 0.0001) were found.

Marination increased IF the content in chicken meat samples. The highest increase of IF was found in organic chicken breast samples (13.33%), and the lowest in conventionally produced chicken meat thigs (4.49%) as compared with the control samples, and it ranged from 0.58 ± 0.01% (in conventionally produced chicken breast samples) to 0.93 ± 0.03% (in conventionally produced chicken meat thighs samples). A moderate positive significant correlation between the IF and moisture content in meat samples (r = 0.525; p = 0.001) and a moderate positive significant correlation between the meat IF content and WHC (r = 0.487; p = 0.003) were found.

Colour characteristics. Colour parameters (L*, a* and b*) of organic and conventionally produced chicken breasts, drumsticks and thighs marinated with P. acidilactici KTU05-7 cultivated in potato juice are presented in Figure 3.2.1.7. It was found that the lighteness (L*) increased in all three treated groups (p < 0.05) of chicken meat. Changes of colour parameters (L*, a* and b*) depended on the chicken meat production method (organic or conventional) (p ≤ 0.0001) and on the part of chicken meat (p ≤ 0.0001). Chicken meat treatment with marinade based on P. acidilactici increased the (L*) values of organic (from 1.77% of thighs to 3.38% of breast) and of conventionally produced chicken meat samples (from 1.20% of drumstick to 5.04% of breast) as compared with the control samples. The marination of conventionally produced chicken breast, drumstick and thigh samples increased the chicken meat (L*) values on average by 0.55% as compared with organic chicken meat. A very strong negative significant correlation between the meat lighteness (L*) and redness (a*) (r = -0.819; p = 0.0001) and a very strong negative significant correlation between the meat lighteness (L*) and yelowness (b*) (r = -0.873; p = 0.0001) were found.

Marination decreased the (a*) values of organic and conventionally produced chicken meat on average by 3.35% and 8.99%, respectively (of organic chicken meat samples from 0.78% of breast samples to 6.04% of drumstick samples) and of conventionally produced chicken meat samples from 6.72% of thigh samples to 11.73% of breast samples as compared with the control samples. A strong negative significant correlation between the meat redness (a*) and lighteness (L*) (r = -0.819; p = 0.0001), a strong

86

positive significant correlation between the meat redness (a*) and yelowness (b*) (r = 0.854; p = 0.0001) were found.

Figure 3.2.1.7. Colour parameters (L*, a*, and b*) of organic and

conventionally produced chicken breasts, drumsticks and thighs marinated with P. acidilactici KTU05-7 cultivated in potato juice (Remarks: Pa – P.

acidilactici, L* − lightness, a* − redness (or –a* of greenness), and b* − yellowness (or –b* of blueness, C – control, Conv. – conventionally).

In all the cases, marination with P. acidilactici decreased the (b*) values

of chicken meat samples on average by 6.75% (of organic chicken breast by 10.09%, of drumsticks by 2.39%, of thighs by 7.75%). Similar tendencies of conventionally produced chicken breast samples were estimated, the (b*) values were reduced on average by 11.73%. A very strong negative significant correlation between the meat yelowness (b*) and lighteness (L*) (r = -0.873; p = 0.0001), a very strong positive significant correlation between the meat yelowness (b*) and redness (a*) (r = 0.854; p = 0.0001) were found.

The content of BAs in organic and conventionally produced chicken

breast, drumsticks and thighs marinated with P. acidilactici KTU05-7 cultivated in potato juice. The BAs content in organic and conventionally produced chicken breast, drumsticks and thighs is presented in Table 3.2.1.7. The BAs content significantly depended on the part of chicken meat (p ≤ 0.0001) and on the chicken production method (organic or conventional) (p ≤ 0.0001). In many cases, marination increased the PHE content in chicken meat (up to 7.96–30.55 mg/kg), except in conventionally produced marinated breast and drumstick samples. A moderate negative significant correlation between the PHE and meat pH (r = -0.455; p = 0.005), a moderate positive significant correlation between the PHE and SPR content (r = 0.456; p = 0.005), a weak positive significant correlation

0

20

40

60

80

100

C Pa C Pa C Pa C Pa C Pa C Pa

Organic Conv. Organic Conv. Organic Conv.

Breast Drumsticks Thighs

b*

a*

L*

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between the PHE and meat moisture content (r = 0.359; p = 0.031) were found. Marination decreased the PUT content in all conventionally produced chicken meat samples, and the PUT content ranged from 15.84 ± 0.14 mg/kg (in marinated drumstick samples) to 86.96 ± 0.16 mg/kg (in thigh control samples). Different tendencies of organic chicken meat samples were estimated: marination increased the PUT content by 213.88% in breast samples, by 312.71% in drumsticks samples and by 337.31% in thighs samples as compared with the control samples. A weak positive significant correlation between the PUT and CAD (r = 0.351; p = 0.036) was found. The concentration of CAD in conventionally produced chicken meat samples was found to range from 4.77 ± 0.07 mg/kg (in marinated breast samples) to 50.84 ± 0.13 mg/kg (in drumstick control samples); hovewer, no CAD in all organic chicken meat samples was found. A weak positive significant correlation was found between the CAD and PUT content (r = 0.351; p = 0.036).

The concentration of histamine (HIS) in organic chicken meat samples ranged from 5.33 ± 0.06 mg/kg to 23.76 ± 0.30 mg/kg (in drumstick control samples and in with P. acidilactici marinated thigh samples, respectively) and in conventionally produced meat samples it ranged from 9.12 ± 0.19 mg/kg to 44.85 ± 0.01 mg/kg (in thigh control samples and in marinated drumstick samples, respectively). In most cases marination increased the content of HIS (from 54.03% to 180.31% in organic drumstick samples and in conventionally produced drumstick samples, respectively, compared to control samples), except conventionally produced breast samples. A weak positive significant correlation between the HIS and CAD content (r = 0.354; p = 0.034) was found.

Chicken meat treatment with marinade based on P. acidilactici increased the TYR content in all analysed chicken meat samples (except in organic marinated chicken thigh samples) as compared with control samples. The concentration of TYR in organic and conventionally produced chicken meat samples ranged from 1.31 ± 0.02 mg/kg (in breast control samples) to 11.47 ± 0.13 mg/kg (in marinated drumstick samples) and from 5.35 ± 0.06 mg/kg (in breast control samples) to 11.39 ± 0.13 mg/kg (in marinated thigh samples), respectively.

The concentration of SPD in organic chicken meat ranged from 1.09 ± 0.02 mg/kg (in drumstick control samples) to 4.17 ± 0.04 mg/kg (in thigh control samples), and in conventionally produced chicken meat samples it ranged from 2.17 ± 0.0 mg/kg (in breast control samples) to 8.61 ± 0.10 mg/kg (in marinated drumstick samples). A weak negative significant correlation between the SPD and SPR content (r = -0.385; p = 0.02), and a

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moderate negative significant correlation between the SPD and moisture content (r = -0.475; p = 0.003) were found.

Table 3.2.1.7. BAs content (mg/kg) in organic and conventionally produced chicken meat, marinated with P. acidilactici KTU05-7 cultivated in potato juice.

BAs, mg/kg

Organic Organic Conventionally Conventionally Control P. acidilactici Control P. acidilactici

Breast PHE 6.37±0.11a 8.85±0.21b 16.96±0.14d 16.93±0.17d PUT 16.50±0.15a 51.79±0.10f 29.87±0.13d 19.44±0.20b CAD - - 20.34±0.24d 4.77±0.07a HIS - - 32.59±0.09a - TYR 1.31±0.02a 2.96±0.03b 5.35±0.06c 10.16±0.10e SPD 2.47±0.04b 1.78±0.02a 2.17±0.02a 8.33±0.09c SPR 0.63±0.01a 1.45±0.01a 3.74±0.04b 2.39±0.02b Total 27.28a 66.83c 111.02e 62.02c

Drumsticks PHE - 20.10±0.07e 6.26±0.12b 3.92±0.06a PUT 17.23±0.10a 71.11±0.25e 61.08±0.17d 15.84±0.14a CAD - - 50.84±0.13c 15.94±0.31a HIS 5.33±0.06a 8.21±0.09b 16.00±0.11d 44.85±0.01f TYR 10.24±0.12b 11.47±0.13c 6.34±0.07a 10.22±0.10b SPD 3.11±0.03b 1.09±0.01a 4.20±0.06b 8.61±0.10c SPR - 2.55±0.04b 0.63±0.01a - Total 35.91a 114.53d 145.35e 99.38c

Thighs PHE 12.28±0.12c 30.55±0.23e - 7.96±0.09a PUT 7.13±0.15a 31.18±0.14d 86.96±0.16f 21.87±0.11c CAD - - 18.06±0.18b 13.97±0.35a HIS 15.30±0.18c 23.76±0.30d 9.12±0.19a 18.47±0.20e TYR 6.20±0.06a 5.33±0.06a 10.41±0.12b 11.39±0.13b SPD 4.17±0.04b 2.61±0.03a 8.33±0.10c - SPR - 1.35±0.01a 1.12±0.01a 3.17±0.04b Total 45.08a 94.78c 134.0e 76.83b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine; BAs – biogenic amines

SPR was not found in most of organic meat samples (except in breast

samples), and the highest concentration of SPR in conventionally produced chicken breast control samples (3.74 ± 0.04 mg/kg) was found. A moderate positive significant correlation between the SPR and PHE content (r = 0.456; p = 0.005) and a moderate negative significant correlation between the SPR and meat pH (r = -0.432; p = 0.008) was found.

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3.2.2. Parameters of cold smoked pork sausages

Antimicrobial activity of P. acidilactici KTU05-7, L. sakei KTU05-6, P. pentosaceus KTU05-9 strain supernatants used for cold smoked pork sausages treatment. The antimicrobial activities of the testet LAB are presented in Table 3.2.2.1. As could be seen from the obtained results, LAB supernatants inhibited the growth of all tested bacteria. The diameters of the inhibition zones towards pathogenic and opportunistic strains varied between 8.0 mm and 19.0 mm. The highest antimicrobial activity of L. sakei against Y. pseudotuberculosis (the inhibition zone diameter was 19.0 ± 0.5 mm) was demonstrated. The lowest inhibition zones against B. cereus (8.0 ± 0.5 mm of P. acidilactici and P. pentosaceus supernatants and 8.2 ± 0.4 mm of L. sakei) were observed. The results of the ANOVA test indicated that the antimicrobial activity of testet LAB significantly depended on the type of used LAB (against P. aeruginosa p ≤ 0.0001, S. aureus p ≤ 0.002, L. monocytogenes p ≤ 0.006, E. coli p ≤ 0.003, S. typhimurium p ≤ 0.0001, Y. enterolitica p ≤ 0.001, Y. pseudotuberculosis p ≤ 0.0001).

Table 3.2.2.1. Inhibition of the growth of pathogenic and opportunistic bacteria by selected LAB.

Microorganisms Zone of inhibition/mm* P. acidilactici 7 P. pentosaceus 9 L. sakei

P. aeruginosa 11.0±0.5b 15.0±0.6d 13.0±0.4c S. aureus 12.0±0.6c 13.0±0.5c 10.0±0.6b L. monocytogenes 11.0±0.3b 9.6±0.5a 11.0±0.3b E. coli 11.0±0.3b 11.0±0.6b 8.6± 0.7a S. typhimurium 13.0±0.5b 12.5±0.3b 9.5±0.4a Y. enterolitica 12.5±0.3c 13.0±0.5c 10.9±0.3b Y. pseudotuberculosis 15.0±0.4d 18.0±0.4e 19.0±0.5e B. cereus 8.0±0.5a 8.0±0.5a 8.2±0.4a Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). * The diameter of wells was 6 mm

The content of BAs in cold-smoked pork sausages. The results of BAs

formation in the upper and middle parts of the sausages treated with LAB before and after smoking are presented in Table 3.2.2.2. The main BAs in the upper and middle parts of sausages treated with L. sakei before smoking were SPR (13.5 ± 0.1 mg/kg and 1.73 ± 0.7 mg/kg) and PUT (2.92 ± 0.1 mg/kg and 29.25 ± 0.2 mg/kg), in sausages treated before smoking with P. acidilactici the main BAs were PUT (27.86 ± 0.1 mg/kg) and SPR (46.63 ± 0.2 mg/kg), and in sausages treated before smoking with P. pentosaceus the main BA was PUT (45.7 ± 0.2 mg/kg and 40.56 ± 0.3 mg/kg). The main BA

90

in the upper and middle parts of sausages treated with the L. sakei after smoking was TYR (20.1 ± 0.1 mg/kg and 10.67 ± 0.8 mg/kg, respectively), in sausages treated with P. acidilactici after smoking – TYR (28.68 ± 0.2 mg/kg and 6.33 ± 0.8 mg/kg) and SPR (14.31 ± 0.1 mg/kg and 26.88 ± 0.1 mg/kg), and in sausages treated with P. pentosaceus after smoking – CAD (17.15 ± 0.5 mg/kg and 20.10 ± 0.2 mg/kg), respectively. The total BAs content was lower in control samples and in samples treated with water (after and before smoking) found as compared with samples treated with LAB (from 8.15 to 97.67% and from 4.08 to 99.65%, respectively).

The results of the ANOVA test indicated that there was a significant effect of the type of LAB applied for the fermentation on PHE (p = 0.015), CAD (p = 0.019), TYR (p = 0.0001), and SPR (p = 0.0001) formation in sausages. Also, a significant influence on the TYR content (p = 0.008) had different parts (upper or middle) of the sausages.

No significant differences among the content of other evaluated BAs in the upper and the middle parts of the sausages were found. Sausage treatment with LAB before and after smoking had a significant effect on PHE (p = 0.041), PUT (p = 0.0001), CAD (p = 0.0001) and HIS (p = 0.001) formation. The interaction among all the analysed factors (type of LAB, part of sausages, treatment before and after smoking) was determined as statistically significant (p < 0.0001).

Table 3.2.2.2. BAs content (mg/kg) in cold smoked pork sausages.

BAs L. sakei P. acidilactici 7 P. pentosaceus 9

UP MP UP MP UP MP

Treated with LAB before smoking PHE 5.01±0.1a - - 32.17±0.2c - - PUT 2.92±0.1a 29.25±0.2e 27.86±0.1e 2.33±0.1a 45.7±0.2f 40.56±0.3f CAD - 1.98±0.1a 2.38±0.1a - 2.23±0.1a 2.87±0.1a HIS 10.2±0.7b - - 8.17±0.2a - - TYR 10.3±0.9c 16.20±0.2d 17.4±0.6e 1.09±0.1a 8.25±0.2c 9.34±0.2c SPD 1.38±0.1a 0.34±0.1a 1.43±0.1a 2.87±0.1b 1.10±0.1b 1.70±0.1b SPR 13.5±0.1e 1.73±0.7a 1.92±0.1a 46.63±0.2f 2.37±0.1b 3.30±0.1b Total 43.26a 49.50b 50.99b 93.26e 59.65c 57.77c

Treated with LAB after smoking PHE - - - - - - PUT 5.89±0.7a 8.47±0.8b 10.2±0.1b 4.87±0.6a - - CAD 14.6±0.5c 9.83±0.6a 11.5±0.3b - 17.15±0.5e 20.10±0.2d HIS - 5.23±0.9b - 12.42±0.9d 14.23±0.6d 2.00±0.2a TYR 20.1±0.1d 10.67±0.8c 28.68±0.2e 6.33±0.8b 8.18±0.7b 9.25±0.8c SPD 13.5±0.4d 3.18±0.3b 15.17±0.1e 3.26±0.4b 13.43±0.3d 2.09±0.2a SPR 2.96±0.3b 4.25±0.6b 14.31±0.1d 26.88±0.1f 13.47±0.4d 12.99±0.3d Total 57.31d 41.63a 79.86f 53.76d 65.46e 46.43b

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Continue of Table 3.2.2.2. BAs NT TW NT TW

UP MP UP MP UP MP UP MP Treated with LAB before smoking Treated with LAB after smoking

PUT 17.29 ±0.17d

16.14 ±0.12d

12.31 ±0.10c

11.03 ±0.14c

17.29 ±0.17d

14.16 ±0.09c

10.24 ±0.11b

11.41 ±0.13c

CAD 9.04

±0.11c 7.31

±0.14b 9.02

±0.14c 8.29

±0.12b 9.04

±0.11a 10.23 ±0.11b

7.65 ±0.01a

9.21 ±0.12a

HIS - - - - - 3.44

±0.14a 16.14 ±0.09e

13.29 ±0.17d

TYR 12.91 ±0.14d

11.12 ±0.07c

3.22 ±0.12a

4.21 ±0.14b

12.91 ±0.11c

11.25 ±0.12c

3.67 ±0.04a

2.29 ±0.09a

SPD - 5.09

±0.04d 5.31

±0.09d 11.24 ±0.18f

- - 0.80

±0.01a 2.41

±0.06a

SPR 0.99

±0.05a 1.27

±0.10a 4.82

±0.11c 5.62

±0.09c 0.99

±0.05a 1.21

±0.07a 1.28

±0.02a 2.08

±0.04b Total 40.23a 40.93a 40.00a 40.39a 40.23a 40.29a 39.78a 40.69a Data are the mean ± SD (n = 3); SD – standard deviation; Mean values within column with different letters are significantly different (p < 0.05); LAB – lactic acid bacteria; UP – the upper part of the sausage; MP – the middle part of the sausage; BS – before smoking; AS – after smoking; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine; BAs – biogenic amines; TW – treated with water, NT – nontreated.

Content of PAH in cold-smoked pork sausages. Results of PAHs

formation in the upper and middle parts of the cold-smoked sausages treated with LAB before and after smoking are presented in Table 3.2.2.3.

It was found a significant effect of the type of LAB applied for the fermentation on BaA (p = 0.0001), BbF (p = 0.0001), BaP (p = 0.001) and Chr (p = 0.0001) content in cold-smoked sausages.

Significant changes in different parts (upper or middle) of the sausages depending on BaA (p = 0.0001), BbF (p = 0.035), BaP (p = 0.0001) and Chr (p = 0.0001) content were estimated. Sausage treatment with LAB before and after smoking had a significant effect on the BbF (p = 0.049) and BaP (p = 0.027) content.

BaA and Chr formation in the upper and middle parts of the cold-smoked sausages was similar, and the differences were statistically insignificant. The interaction among all the analysed factors (the type of LAB, different parts of sausages, treatment before and after smoking) on the BaP (p = 0.014) and Chr (p = 0.009) content was determined as statistically significant.

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Table 3.2.2.3. PAH content (ng/g) in cold smoked pork sausages middle and upper part.

PAHs L. sakei P. acidilactici 7 P. pentosaceus 9 BS AS BS AS BS AS

Middle part

BaA, ng/g 0.068

±0.005b 0.054

±0.003a 0.061

±0.004a 0.063

±0.010b 0.065

±0.007b 0.062

±0.005b

Chr, ng/g 0.149

±0.007b 0.168

±0.009c 0.204

±0.006e 0.144

±0.011b 0.168

±0.005c 0.104

±0.010a

BbF, ng/g 0.021

±0.009a 0.033

±0.005b 0.030

±0.004b 0.039

±0.002c 0.031

±0.001b 0.051

±0.004d

BaP, ng/g 0.039

±0.004b 0.030

±0.002a 0.036

±0.007b 0.034

±0.003a 0.034

±0.005a 0.042

±0.003b ∑PAHs 0.227a 0.285c 0.331c 0.280c 0.298d 0.259b

Upper part

BaA, ng/g 0.072

±0.003b 0.065

±0.004a 0.069

±0.005a 0.071

±0.011b 0.069

±0.004a 0.072

±0.003b

Chr, ng/g 0.167

±0.011b 0.184

±0.015d 0.212

±0.014e 0.163

±0.014b 0.184

±0.003d 0.124

±0.012a

BbF, ng/g 0.040

±0.007a 0.051

±0.007b 0.041

±0.005a 0.062

±0.008c 0.054

±0.007b 0.073

±0.005d

BaP, ng/g 0.062

±0.005b 0.054

±0.006a 0.051

±0.009a 0.072

±0.007b 0.068

±0.002b 0.059

±0.004a ∑PAHs 0.341b 0.354b 0.373c 0.368c 0.375c 0.328a PAHs Non treated Treated with water BS Treated with water AS

Middle part BaA, ng/g 0.072±0.006c 0.069±0.005c 0.061±0.003a Chr, ng/g 0.197±0.013d 0.169±0.014c 0.174±0.009c BbF, ng/g 0.033±0.003b 0.021±0.002a 0.024±0.002a BaP, ng/g 0.081±0.007d 0.079±0.004d 0.067±0.005c ∑PAH4, ng/g 0.383f 0.328d 0.326d

Upper part BaA, ng/g 0.086±0.009c 0.074±0.003b 0.068±0.005a Chr, ng/g 0.212±0.011e 0.182±0.013d 0.176±0.012c BbF, ng/g 0.037±0.005a 0.048±0.003b 0.032±0.005a BaP, ng/g 0.106±0.010d 0.098±0.007d 0.081±0.010c ∑PAHs, ng/g 0.441e 0.402d 0.357b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). PAHs – polycyclic aromatic hydrocarbons; BaA – benz[a]anthracene; BbF – benzo-[b]fluoranthene; BaP – benzo[a]pyrene; Chr – chrysene; BS –sausages treated with LAB before smoking; AS – sausages treated with LAB after smoking

Sensory parameters of cold-smoked pork sausages. The parameters of

the sensory evaluation of cold-smoked sausages are presented in Figure 3.2.2.1. The results indicated that the overall acceptability of sausages depended on the LAB strain used for the treatment (p ≤ 0.0001).

93

Figure 3.2.2.1. Cold-smoked sausages sensory analysis by evaluation of panelists facial expression intensity (Remark: LAB – lactic acid bacteria, Pa - P.

acidilactici, Ls - L. sakei, Pp 8 - P. pentosaceus 8, BS – cold smoked pork sausages treated with LAB before smoking, AS – cold smoked pork sausages treated with LAB after

smoking).

Samples treated with P. pentosaceus were indicated having the highest acceptability (5.0–4.5 scores), except control samples (4.8 scores). The lowest acceptability was found in sausages treated with L. sakei (on average 4.35 scores), and this sample acceptability was lower by 9.38% as compared with control samples. The most acceptable were sausages treated with L. sakei, P. acidilactici, P. pentosaceus before smoking as compared with sausages treated with LAB after smoking (by 7.14%, 2.13% and 11.11%, respectively). The higher intensity of the emotion “happy” was fixed for tested samples which were evaluated by panelists for the highest acceptability in the score system. A very strong positive significant correlation was found between the acceptability in the score system and the emotions “happy”, “sad”, and “angry” fixed by the FaceReader software (r = 0.838; p = 0.0001, r = 0.837; p = 0.0001, r = 0.837; p = 0.0001, respectively).

3.2.3. Parameters of ready-to-cook minced pork meat products

3.2.3.1. Parameters of ready-to-cook minced pork meat products produced with biotreated Satureja montana

The overall acceptability of RCMP produced with different quantities of fermented Sm. The overall acceptability of RCMP samples produced with 3%, 5% and 7% of Sm bioproducts was significantly different (p ≤ 0.0001) (Figure 3.2.3.1.1.). The overall acceptability of RCMP was influenced by the type of LAB used for savory plant fermentation, by the type of a plant, and by the quantity of bioproducts used

0123456

00,10,20,30,40,50,6

BS AS BS AS BS AS

Pa Ls Pp C

Sco

res

Em

otio

ns

Happy Sad Angry Points

94

for RCMP production (p = 0.0001). RCMP samples supplemented with 3% of Sm were indicated as to having the highest overall acceptability. The acceptability values of the latter samples in all the cases were higher on average by 25.6% (SMF) and 31% (SSF), respectively, as compared with RCMP samples produced with 5% of Sm.

RCMP samples produced with 7% of Sm were rated as unacceptable to consumers due to the intensive additive taste and flavour. The acceptability of control samples was lower (from 20% to 33%) as compared with samples produced with 3% of fermented Sm.

According to the obtained results, for the further analysis RCMP with 3% of Sm bioproducts were used.

Figure 3.2.3.1.1. The overall acceptability of RCMP samples produced with 3%, 5% and 7% of biotreated Sm (Remark: SMF – submerged fermentation and SSF

– solid state fermentation; Sm – S. montana, RCMP – ready-to-cook minced pork meat products).

The quality parameters of RCMP produced with biotreated Sm. The

quality parameters of RCMP produced with Sm bioproducts are presented in Table 3.2.3.1.1. The fermentation type of Sm additives had a significant (p < 0.05) influence on the quality parameters of the RCMP. The moisture content of RCMP ranged from 27.84 ± 0.2% (RCMP treated with SMF with P. pentosaceus Sm) to 37.06 ± 0.04% (control samples). Savory plant fermentation conditions (SMF or SSF) did not have a significant influence on the RCMP pH (it ranged from 6.02 ± 0.4 to 6.16 ± 0.1, in RCMP produced with SMF with P. pentosaceus Sm and in RCMP produced with SMF with L. sakei Sm, respectively). A moderate negative significant correlation between the RCMP pH and drip loss (r = -0.523; p = 0.015) was found. The supplementation of RCMP with SMF Sm had a positive influence on the WHC and tenderness of meat products. The RCMP produced with SMF with P. acidilactici Sm had the highest WHC (72.4 ± 0.8) and drip loss (3.06 ± 0.4), followed by samples produced with SMF

0

2

4

6

8

SMF SSF SMF SSF SMF SSF

Pa Ls PpRCMP with 3 % RCMP with 5 %

95

with P. pediococcus Sm (69.4 ± 0.2 and 2.16 ± 0.2, respectively), followed by samples produced with SMF with and L. sakei Sm (67.8 ± 0.4 and 2.23 ± 0.3, respectively). The SSF Sm bioproducts decreased the WHC and drip loss of RCMP (by 20% and 17%, respectively) compared to control samples. The drip loss was significantly influenced by the type of LAB used for the fermentation of savory plants (p ≤ 0.008) and by the type of their fermentation (p ≤ 0.0001). A strong negative significant correlation between the RCMP drip loss and WHC (r = -0.636; p = 0.002), a moderate negative significant correlation between the RCMP drip loss and pH (r = -0.523; p = 0.015), and a moderate positive significant correlation between the RCMP drip loss and the moisture content of savory plant bioproducts (r = 0.463; p = 0.035) was found.

Table 3.2.3.1.1. Quality parameters of RCMP produced with fermented Sm.

Sample Moisture, %

pH Drip loss, %

WHC, %

CL, %

Shear kg/cm2

S. montana (3%) Ls SMF 29.67±0.3c 6.16±0.1c 2.23±0.3b 65.9±0.2b 26.36±0.2b 0.27±0.02b Ls SSF 30.60±0.3d 6.08±0.2b 1.20±0.2a 67.8±0.4b 25.32±0.1a 0.29±0.02c Pa SMF 29.56±0.4c 6.08±0.1b 3.06±0.4c 63.5±0.5a 26.57±0.1b 0.32±0.03c Pa SSF 26.94±0.2a 6.04±0.3a 1.21±0.2a 72.4±0.8e 25.25±0.2a 0.20±0.02a Pp SMF 27.84±0.2b 6.02±0.4a 2.16±0.2b 64.0±0.3a 32.28±0.3d 0.20±0.01a Pp SSF 30.18±0.3c 6.10±0.2b 1.19±0.1a 69.4±0.2c 29.32±0.2c 0.25±0.02b

S. montana (5%) Ls SMF 36.82±0.5d 6.03±0.3b 2.07±0.2c 61.8±0.4a 26.97±0.2b 0.28±0.03b Ls SSF 30.62±0.3b 6.03±0.5b 1.55±0.2a 65.7±0.5c 21.46±0.3a 0.42±0.02c Pa SMF 29.65±0.2a 6.02±0.4b 1.96±0.2b 67.4±0.6d 25.49±0.2b 0.31±0.03b Pa SSF 30.80±0.3b 5.85±0.4a 1.68±0.2a 63.4±0.5b 28.52±0.3d 0.22±0.01a Pp SMF 32.24±0.4c 6.00±0.3b 2.04±0.2c 64.1±0.5b 19.69±0.1a 0.21±0.01a Pp SSF 30.08±0.3a 6.04±0.4c 1.88±0.2b 65.1±0.5c 27.03±0.2c 0.31±0.02b

S. montana (7%) Ls SMF 28.35±0.3a 5.99±0.2a 2.43±0.29d 67.6±0.63c 22.72±0.19b 0.26±0.02a Ls SSF 28.63±0.3a 6.30±0.3c 2.06±0.23b 66.2±0.66b 23.90±0.17b 0.33±0.03b Pa SMF 30.82±0.4c 6.02±0.2a 2.31±0.20c 67.1±0.48c 20.29±0.22a 0.27±0.02a Pa SSF 30.59±0.4c 6.14±0.4c 1.85±0.18a 64.3±0.63b 25.17±0.29c 0.33±0.03b Pp SMF 30.00±0.3b 6.05±0.3b 2.20±0.19c 59.3±0.57a 27.17±0.23d 0.23±0.01a Pp SSF 29.40±0.2b 6.17±0.4c 1.69±0.14a 56.0±0.43a 26.45±0.25c 0.30±0.02b Control 37.06±0.04e 6.07±0.3d 1.96±0.15c 60.1±0.45b 24.46±0.22c 0.42±0.03d Data are the mean ± SD (n = 3); Mean values within column with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana; WHC – Water holding capacity; Ls – L. sakei; Pa – P. acidilactici; Pp – P. pentosaceus

96

The WHC of RCMP was significantly influenced by the type of LAB applied for savory plant bioproduct fermentation and by the type of the savory plant fermentation method (p ≤ 0.0001 and p ≤ 0.001, respectively).

A strong negative significant correlation between the WHC and the drip loss of RCMP (r = -0.636; p = 0.002), a strong negative significant correlation between the WHC and the moisture content of RCMP (r = -0.672; p = 0.001), a moderate negative significant correlation between the WHC and tenderness of RCMP (r = -0.560; p = 0.008), a strong positive significant correlation between the WHC of RCMP and the TTA of savory plant bioproducts (r = 0.628; p = 0.002) and a strong positive significant correlation between the WHC of RCMP and the pH of savory plant bioproducts (r = 0.729; p = 0.0001) was found.

In all the cases, RCMP produced with plant bioproducts had a higher tenderness than the control samples (the shear force of RCMP supplemented with fermented Sm was lower on average by 16.7% (SSF Sm) and by 9% (SMF Sm)). A strong positive significant correlation between the tenderness and pH of RCMP (r = 0.788; p = 0.0001), a moderate negative significant correlation between the tenderness and WHC of RCMP (r = -0.560; p = 0.008), a moderate negative significant correlation between the tenderness of RCMP and the moisture content of savory plant bioproducts (r = -0.572; p = 0.007), a strong negative significant correlation between the tenderness of RCMP and the TTA of savory plant bioproducts (r = -0.734; p = 0.0001) and a strong negative significant correlation between the tenderness of RCMP and the pH of savory plant bioproducts (r = -0.742; p = 0.0001) were found.

The influence of Sm bioproducts on the RCMP cooking loss was not significant, with the exception of samples produced with Sm P. pentosaceus bioproducts. RCMP with savory plant additives had by 16.3% (SSF) and 21.9% (SMF) higher cooking loss as compared with control samples. A moderate negative significant correlation between the cooking loss and moisture content of RCMP (r = -0.446; p = 0.043) and a moderate positive significant correlation between the cooking loss of RCMP and moisture content of savory plant bioproducts (r = 0.565; p = 0.008) were found.

Colour characteristics. Significant differences among the RCMP colour parameters L*, a*, and b* (p < 0.05) were observed (Table 3.2.3.1.2.). Control meat samples showed the highest lightness (L*) (65.84 ± 1.78) and yellowness (b*) (15.09 ± 0.71) but lowest redness (a*) (3.91 ± 0.21) as compared with the RCMP produced with Sm bioproducts. The RCMP produced with SSF Sm showed a lower lightness (L*) (ranged between 58.50 ± 1.07 and 58.80±2.31), redness (a*) (between 5.09 ± 0.53 and 5.67 ±

97

0.29), and yellowness (b*) (ranged between 12.54 ± 0.26 and 13.23 ± 0.22), while RCMP produced with SMF Sm showed a higher lightness (L*) (between 59.36 ± 0.97 and 61.01 ± 1.02) and redness (between 5.39 ± 0.88 and 6.22 ± 0.74), as well as yellowness (between 13.59 ± 0.27 and 14.57 ± 0.16). Thus, RCMP samples produced with SSF Sm were darker as compared with RCMP samples produced with SMF Sm.

The yellowness (b*) of RCMP was significantly influenced by the fermentation method of savory plants (p ≤ 0.0001) and by the type of LAB used for savory plant fermentation (p ≤ 0.001). A strong positive significant correlation between the (L*) and (b*) values of RCMP (r = 0.768; p = 0.0001) was found.

Table 3.2.3.1.2. Colour parameters (L*, a* and b*) of the RCMP produced with biotreated Sm.

RCMP L* a* b* S. montana (3%)

Ls SMF 61.01±1.02c 6.22±0.74d 13.62±0.54c SSF 58.80±2.31a 5.29±0.69b 12.54±0.26a

Pa SMF 59.61±1.89b 5.39±0.88b 13.59±0.27b SSF 58.76±1.26a 5.09±0.53a 13.17±0.19b

Pp SMF 59.36±0.97b 6.21±0.42d 14.57±0.16c SSF 58.50±1.07a 5.67±0.29c 13.23±0.22b

S. montana (5%) Ls SMF 58.88±1.12b 5.20±0.60c 14.71±0.43c

SSF 55.05±0.99a 5.09±0.55b 11.58±0.18a Pa SMF 60.30±1.28c 5.33±0.64c 14.37±0.15b

SSF 54.95±1.17a 5.17±0.53b 12.09±0.31a Pp SMF 57.93±1.22b 5.58±0.48d 14.14±0.18b

SSF 56.23±0.99b 4.71±0.51a 12.36±0.25a S. montana (7%)

Ls SMF 57.75±1.38c 4.85±0.52c 13.64±0.18c SSF 55.73±1.27b 3.65±0.43a 11.30±0.17a

Pa SMF 56.74±1.30b 4.54±0.47c 14.07±0.20c SSF 53.22±1.25a 3.89±0.36b 10.96±0.14a

Pp SMF 55.65±1.31b 5.35±0.61d 14.19±0.28c SSF 52.96±1.53a 4.29±0.53c 11.92±0.25b

Control (with semolina)

70.61±2.17e 4.11±0.43c 15.17±0.17d

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p <0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana; L* – lightness; a* – redness; b* – yellowness

Microbiological parameters of RCMP produced with biotreated Sm.

The microbiological analysis showed satisfactory sanitary conditions for all

98

of the RCMP samples produced with 3% of fermented Sm (Figure 3.2.3.1.2), although the average counts of microorganisms observed in all samples increased during 5 days of storage under refrigeration. The inhibition effect on the growth of undesirable microorganisms significantly (p < 0.05) depended on the treatment (SMF or SSF) of Sm bioproducts and the LAB used for their fermentation.

The SSF processing allowed the reduction of mesophilic bacteria growth during 120 hours of storage by up to 33–34% as compared with the control samples, while the RCMP samples containing SMF Sm bioproducts had a lower impact (the reduction of mesophilic bacteria growth during 120 hours of storage reached 13.2–17.4%). The highest antimicrobial activity against pathogens was shown by SSF Sm bioproducts fermented with P. acidilactici. The count of coliform bacteria in RCMP samples containing Sm bioproducts fermented with P. acidilactici was found to be lower by 28.2% (SSF) and by 21.5% (SMF), respectively, as compared with the control (5.03 log10 CFU/g). The SSF with P. acidilactici and L. sakei Sm bioproducts decreased the yeast and mold growth by 53.2% and 48.4%, respectively, compared with the control samples. A moderate negative significant correlations was found between mesophilic bacteria and coliforms in RCMP and the pH of savory plant bioproducts (r = -0.485; p = 0.0001; r = -0.444; p = 0.0001, respectively) and a strong negative significant correlation between mesophilic bacteria in RCMP and the pH of savory plant bioproducts (r = -0.763; p = 0.0001).

Figure 3.2.3.1.2. Microbiological parameters of RCMP produced with

biotreated Sm (3%, m/m) (Remark: SMF – submerged fermentation and SSF – solid state fermentation, Ls – L. sakei; Pa – P. acidilactici; Pp – P. pentosaceus, RCMP – ready-to-cook minced pork meat products, MB - mesophilic bacteria, C – coliforms, Y/M – yeasts

and mold).

A weak negative significant correlation between the coliform in RCMP

and the moisture content in the savory plant bioproducts (r = -0.259; p =

0

10

20

30

40

50

MB MB MB MB C C C C Y/M Y/M Y/M Y/M

0 h 24 h 72 h 120 h 0 h 24 h 72 h 120 h 0 h 24 h 72 h 120 h

CF

U/g

log1

0

Ls SMF Ls SSF Pa SMF Pa SSFPp SMF Pp SSF Control

99

0.018), as well as a moderate negative significant correlation between yeasts and mold in RCMP and moisture content of savory plant bioproducts (r = -0.514; p = 0.0001) were found. Also, a moderate positive significant correlation between mesophilic bacteria and coliforms in RCMP and the moisture content of RCMP (r = 0.534 ; p = 0.0001; r = 0.470; p = 0.0001, respectively), a strong positive significant correlation between yeasts and mold in RCMP and the moisture content of RCMP (r = 0.699; p = 0.0001) were found.

The BAs content in RCMP produced with biotreated Sm. The BAs content (mg/kg) in RCMP produced with Sm bioproducts is presented in Table 3.2.3.1.3. The BAs contents in RCMP samples depended on the LAB strain used for savory plant fermentation, as well as on the moisture content of the fermentable substrate (Sm). The PUT, HIS, SPR and SPD were the main BAs in control meat samples (produced without Sm bioproducts), while CAD, TYR and TRP were most abundant in RCMP produced with Sm bioproducts. Both SSF and SMF Sm bioproducts significant by (p < 0.05) reduced the PUT, HIS, SPR and SPD content in RCMP and increased the PHE, CAD, TYR and TRP as compared with control RCMP samples.

In most of the cases, Sm bioproducts increased the PHE content in RCMP (up to 0.07–5.18 mg/kg), except in RCMP produced with SSF with P. pentosaceus Sm. A strong positive significant correlation between the PHE and TRP (r = 0.788; p = 0.0001) and a moderate negative significant correlation between the PHE and meat pH (r = -0.451; p = 0.040) were found. SMF Sm bioproducts increased the PUT content in RCMP, except in RCMP produced with SSF with L. sakei Sm, and its content ranged from 1.58 ± 0.14 mg/kg to 24.15 ± 0.53 (in RCMP produced with SMF with L. sakei Sm and in RCMP produced with SMF with P. pentosaceus Sm, respectively).

A strong positive significant correlation between the PUT and the CAD (r = 0.742; p = 0.0001), and a strong positive significant correlation between the PUT and the TYR (r = -0.688; p = 0.001) in RCMP were found. Sm increased the CAD content in RCMP from 242.86% (in RCMP produced with SMF with L. sakei Sm) to 1452.68% (in RCMP produced with SMF with P. pentosaceus Sm), except in RCMP produced with SSF with L. sakei Sm (reduced by 50%). A strong positive significant correlation between the CAD and the PUT content in RCMP (r = 0.742; p = 0.0001), a strong negative significant correlation between the CAD and the TYR content in RCMP (r = -0.756; p = 0.0001), a moderate negative significant correlation between the CAD and the SPR content in RCMP (r = -0.469; p = 0.032), a strong negative significant correlation between the CAD and the moisture of

100

RCMP (r = -0.641; p = 0.002), and a strong positive significant correlation between the CAD content in RCMP and the moisture of savory plant bioproducts (r = 0.600; p = 0.004) were found.

Table 3.2.3.1.3. BAs content in RCMP produced with biotreated Sm. Samples PHE PUT CAD HIS TYR SPD SPR TRP Total

S. montana (3%) Ls SMF 0.22

±0.02b 1.58

±0.14a 3.84

±0.23b 2.34

±0.31b 4.75

±0.19c - -

0.92 ±0.13a

13.65b

Ls SSF 3.46 ±0.31d

2.93 ±0.42a

0.56 ±0.11a

0.28 ±0.13a

3.12 ±0.09b

0.32 ±0.07a

- 0.77 ±0.24a

11.44a

Pa SMF - 10.69 ±0.18c

16.31 ±0.29e

6.74 ±0.12d

5.18 ±0.10c

3.35 ±0.24b

- 2.04 ±0.09c

44.31f

Pa SSF 0.19 ±0.11b

4.27 ±0.37b

11.59 ±0.71d

0.18 ±0.09a

2.13 ±0.22b

0.89 ±0.16a

1.16 ±0.11a

-0.23 ±0.06a

20.41d

Pp SMF 0.07 ±0.02a

24.15 ±0.53f

17.39 ±0.41e

- 1.17 ±0.20a

- - 0.84 ±0.10a

43.01d

Pp SSF - 15.34 ±0.40d

10.17 ±0.38d

5.15 ±0.21c

0.56 ±0.12a

- 2.44 ±0.18a

- 34.50e

Control - 7.81 ±0.14c

0.70 ±0.02a

4.71 ±0.12c

- 5.54

±0.18c 15.2

±0.62d - 33.96e

S. montana (5%) Ls SMF 5.17

±0.07a 28.36 ±0.21c

14.94 ±0.14c

11.28 ±0.15d

- 4.39

±0.21b 23.07 ±0.31f

- 87.21e

Ls SSF 10.24 ±0.11b

33.17 ±0.38d

5.63 ±0.32b

8.17 ±0.10c

2.36 ±0.18a

- 8.31 ±0.10c

2.54 ±0.09b

70.42d

Pa SMF 17.33 ±0.20d

25.90 ±0.35c

10.74 ±0.14c

9.34 ±0.11c

6.18 ±0.08b

2.11 ±0.17a

- 1.10 ±0.10a

72.70d

Pa SSF - 38.12 ±0.41e

12.10 ±0.18c

4.71 ±0.21a

13.25 ±0.15d

1.11 ±0.10a

0.74 ±0.08a

2.06 ±0.05b

72.09d

Pp SMF 9.11 ±0.10b

28.64 ±0.25c

29.31 ±0.35e

8.44 ±0.10c

3.05 ±0.05a

6.13 ±0.06c

1.15 ±0.02a

- 85.83f

Pp SSF 11.80 ±0.13b

33.04 ±0.35d

20.15 ±0.21d

10.32 ±0.12d

5.15 ±0.09b

- 0.62 ±0.02a

0.30 ±0.01a

81.38a

Control -

7.81 ±0.14a

0.70 ±0.02a

4.71 ±0.12a

- 5.54

±0.18b 15.2

±0.62e - 33.96a

S. montana (7%) Ls SMF 2.17

±0.02a 30.47 ±0.31d

19.22 ±0.20d

8.61 ±0.09c

5.12 ±0.06b

5.54 ±0.18c

3.17 ±0.01b

0.86 ±0.01a

69.62b

Ls SSF 10.17 ±0.12c

26.31 ±0.30d

25.68 ±0.23f

1.22 ±0.10a

3.76 ±0.05a

0.17 ±0.01a

2.31 ±0.03a

1.28 ±0.02b

70.90b

Pa SMF 19.30 ±0.20d

36.92 ±0.40e

15.16 ±0.16d

14.37 ±0.12d

8.96 ±0.07c

3.20 ±0.03b

- 0.58 ±0.01a

98.49d

Pa SSF 5.29 ±0.06b

23.07 ±0.22d

20.45 ±0.26e

12.06 ±0.13d

1.22 ±0.02a

5.37 ±0.06c

0.86 ±0.01a

0.30 ±0.01a

68.62b

Pp SMF 14.82 ±0.16c

36.94 ±0.41e

18.63 ±0.23d

9.37 ±0.10c

5.24 ±0.06b

- 3.19 ±0.03b

- 88.22c

101

Continue of Table 3.2.3.1.3. Samples PHE PUT CAD HIS TYR SPD SPR TRP Total Pp SSF 13.20

±0.09c 28.11 ±0.30d

10.19 ±0.10c

17.36 ±0.15e

8.61 ±0.04c

- 2.13

±0.02a - 82.80c

Control -

7.81 ±0.14a

0.70 ±0.02a

4.71 ±0.12b

- 3.20

±0.02b 15.2

±0.62d - 33.96a

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine

The concentration of HIS in RCMP produced with SSF Sm bioproducts

decreased from 88.03% to 97.33% (in RCMP produced with SSF with L. sakei Sm and in RCMP produced with SSF with P. acidilactici Sm, respectively) as compared with RCMP produced with SMF Sm. The SSF with P. pentosaceus Sm bioproducts increased the HIS content in RCMP samples.

The concentration of TYR in RCMP samples ranged from 0.56 ± 0.12 mg/kg (in RCMP produced with SSF with P. pentosaceus Sm) to 4.75 ± 0.19 mg/kg (in RCMP produced with SMF with L. sakei Sm). In all the cases Sm bioproducts decreased the SPD and SPR content in RCMP (up to 0.32–3.35 mg/kg and up to 1.16–2.44 mg/kg, respectively). The total BAs content was significantly (p < 0.05) reduced in the RCMP produced with Sm bioproducts, except in RCMP produced with SMF with P. acidilactici Sm, and in RCMP produced with SMF with P. pentosaceus Sm. The BAs content in RCMP samples produced with SSF with L. sakei and P. acidilactici Sm was by 71.5% and 53.6% lower, respectively, as compared with the control samples (43.96 mg/kg).

VC in RCMP produced with biotreated Sm The VC (g/100 g) in

RCMP produced with 3% of Sm bioproducts are presented in Table 3.2.3.1.4. The VC content in RCMP enriched with 3% of Sm bioproducts significantly (p < 0.05) depended on the LAB strain used for the fermentation of savory plants and the fermentation technology (SSF or SMF).

The highest content of ρ-cimene (46.7 and 43.6 g/100 g) and γ-terpinene (11.6 and 10.5 g/100 g) was found in RCMP produced with SMF with P. pentosaceus and P. acidilactici Sm, respectively. The highest content of carvacrol was found in RCMP samples produced with SSF with P. pentosaceus Sm (9.0 g/100 g), followed by RCMP samples produced

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with SMF and SSF with P. acidilactici Sm (6.15 and 6.81 g/100 g, respectively). Table 3.2.3.1.4. The VC in RCMP produced with 3% of fermented Sm.

Compounds Time, min.

L. sakei P. acidilactici

7 P. pentosaceus

8 Contr. SMF SSF SMF SSF SMF SSF

isovaleric acid 4.56 6.91c 0.00 4.73b 5.28b 0.00 3.26a 0.00 n-hexanol 5.21 0.00 3.76b 1.34a 11.6d 0.00 5.40c 0.00 isoamyl acetate 5.38 0.00 2.09a 6.75c 0.00 7.09c 4.20b 0.00 α-toluene 6.62 2.78b 2.20a 4.51c 2.12a 4.88c 2.83b 0.00 α-pinene 6.81 0.00 0.00 1.92a 0.00 1.93a 0.00 0.00 vinyl amyl carbinol 8.08 0.00 6.48d 2.32a 3.00b 2.65a 5.75d 0.00 oktan-3-one 8.29 5.39d 3.37c 1.96a 3.62c 4.04c 0.00 0.00 mircene 8.41 0.00 0.00 3.47c 0.00 3.98c 0.00 0.51a α-terpinene 9.16 0.00 0.00 1.22a 0.00 1.52a 0.00 0.00 ρ-cimene 9.39 31.93a 34.8b 43.6d 30.9a 46.72d 31.53a 0.00 γ-terpinene 10.43 4.74a 6.54b 10.5d 4.22a 11.56d 5.63b 0.00 trans-sabinene hidrate 10.68 0.00 3.67b 0.00 2.92a 0.00 2.73a 0.00 n-dekane 11.66 6.42c 7.99d 3.17b 6.27c 2.67a 5.49c 3.40b thymol methyl ether 15.98 0.00 2.56b 1.52a 2.31b 1.46a 2.61b 0.00 carvacrol 17.68 2.28a 6.89d 6.15c 6.81d 4.00b 9.00e 0.00 Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana; VC – volatile compound

3.2.3.2. Parameters of ready-to-cook minced pork meat products

produced with biotreated Satureja hortensis

The overall acceptability of RCMP produced with different quantities of fermented Sh. Sh have a significant influence (p ≤ 0.0001) on the overall acceptability of RCMP produced with 3%, 5% and 7% of plant bioproducts (Table 3.2.3.2.1.).

The overall acceptability of RCMP was influenced by the type of LAB used for the fermentation of savory plants, by the type of the used plant material, and by the quantity of bioproducts used for RCMP production (p = 0.0001). The RCMP supplemented with 3% of Sh so as to have the highest overall acceptability were indicated. The acceptability values of the latter samples in all the cases were higher on average by 23.6% (SMF) and 23.5% (SSF), respectively, as compared with the RCMP produced with 5% of Sh. The RCMP produced with 7% of Sh was rated as unacceptable for

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consumers, due to an intensive additive taste and flavour. The acceptability of control samples was even lower (from 17.6% to 30%) as compared with samples produced with 3% of fermented Sh.

According to the received results, for the further analysis the RCMP with 3% of Sh bioproducts were used. Table 3.2.3.2.1. The overall acceptability of RCMP samples prepared with 3%, 5% and 7% of biotreated Sh.

RCMP L. sakei P. acidilactici P. pentosaceus Control

SMF SSF SMF SSF SMF SSF RCMP with 3 %

5.1 ±0.03b

6.0 ±0.03d

5.8 ±0.03c

5.7 ±0.04c

5.8 ±0.05c

5.8 ±0.05c

4.2 ±0.02a

RCMP with 5 %

4.2 ±0.02a

5.1 ±0.05c

4.6 ±0.02b

4.5 ±0.03b

4.7 ±0.04b

4.6 ±0.05b

4.2 ±0.03a

RCMP with 7 %

1.3 ±0.01b

1.0 ±0.01a

0.8 ±0.01a

1.1 ±0.01b

0.9 ±0.01a

1.3 ±0.01b

4.2 ±0.03c

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sh – S. hortensis; Control – RCMP with semolina

The quality parameters of RCMP produced with fermented Sh. The

quality parameters of RCMP produced with Sh bioproducts are presented in Table 3.2.3.2.2.

The fermentation type (SMF or SSF) of Sh has a significant (p < 0.05) influence on the quality parameters of the RCMP. The moisture content of RCMP produced with Sh bioproducts ranged from 26.59 ± 0.02% (RCMP produced with SSF with P. pentosaceus Sh) to 37.06 ± 0.04% (control samples). The savory plant fermentation conditions have no significant influence on the RCMP pH. The pH of the RCMP samples ranged from 6.01 ± 0.02 to 6.05 ± 0.04 (RCMP produced with SMF with P. pentosaceus Sh and RCMP produced with SSF with L. sakei Sh, respectively). A moderate positive significant correlation between the pH and moisture content of RCMP (r = 0.482; p = 0.027), a moderate negative significant correlation between the pH and WHC of RCMP (r = -0.528; p = 0.014), a very strong negative significant correlation between the pH of RCMP and the moisture of savory plant bioproducts (r = -0.733; p = 0.0001) and a very strong negative significant correlation between the pH of RCMP and the pH of savory plant bioproducts (r = -0.841; p = 0.0001) were found.

The drip loss of the RCMP produced with Sh bioproducts ranged from 1.79 ± 0.15% (RCMP produced with SSF with P. pentosaceus Sh) to 2.19 ± 0.26% (RCMP produced with SMF with L. sakei Sh).

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Table 3.2.3.2.2. Quality parameters of RCMP produced with biotreated Sh. RCMP Moisture,

% pH Drip

loss,% WHC,

% Cooking loss,%

Shear, kg/cm2

S. hortensis (3 %)

Ls SMF

28.67 ±0.03b

6.01 ±0.03a

2.19 ±0.26b

59.10 ±0.61a

22.60 ±0.30a

0.25 ±0.01a

SSF 30.55 ±0.03c

6.05 ±0.04b

1.86 ±0.18a

65.99 ±0.66b

25.88 ±0.21b

0.27 ±0.02a

Pa SMF

29.84 ±0.04c

6.00 ±0.03a

2.11 ±0.20b

61.80 ±0.61a

30.42 ±0.33c

0.31 ±0.03b

SSF 31.40 ±0.04d

6.03 ±0.02b

1.74 ±0.19a

65.23 ±0.68b

19.98 ±0.23a

0.26 ±0.02a

Pp SMF

29.20 ±0.03b

6.01 ±0.02a

2.03 ±0.20b

65.11 ±0.67b

28.45 ±0.25b

0.24 ±0.02a

SSF 26.59 ±0.02a

6.03 ±0.04b

1.79 ±0.15a

71.28 ±0.70d

21.25 ±0.18a

0.26 ±0.03a

S. hortensis (5 %)

Ls SMF 30.77 ±0.04b

6.00 ±0.03a

2.36 ±0.24d

65.25 ±0.61c

25.84 ±0.26c

0.29 ±0.02a

SSF 29.60 ±0.03a

6.20 ±0.03b

1.90 ±0.20b

68.03 ±0.67a

19.24 ±0.20a

0.53 ±0.03b

Pa SMF 32.90 ±0.04b

5.97 ±0.04a

2.21 ±0.21c

64.47 ±0.60b

24.58 ±0.23b

0.25 ±0.02a

SSF 29.05 ±0.03a

6.03 ±0.04a

1.82 ±0.15a

70.47 ±0.64c

25.95 ±0.21c

0.36 ±0.04a

Pp SMF 28.98 ±0.02a

6.07 ±0.05a

2.08 ±0.20b

64.82 ±0.65c

26.11 ±0.22c

0.31 ±0.02a

SSF 29.80 ±0.02a

6.08 ±0.05a

1.64 ±0.17a

64.25 ±0.65c

25.69 ±0.27c

0.25 ±0.02a

S. hortensis (7 %)

Ls SMF 28.54 ±0.03b

6.01 ±0.04a

2.44 ±0.25c

61.32 ±0.61a

21.56 ±0.21a

0.24 ±0.01d

SSF 28.41 ±0.02b

6.21 ±0.05b

1.79 ±0.15a

61.43 ±0.62b

23.93 ±0.26b

0.30 ±0.03a

Pa SMF 29.83 ±0.03c

5.96 ±0.04a

2.19 ±0.23b

61.31 ±0.58d

30.99 ±0.27d

0.42 ±0.03b

SSF 31.12 ±0.04d

6.08 ±0.06a

1.63 ±0.14a

67.75 ±0.70a

21.85 ±0.17a

0.22 ±0.02a

Pp SMF 29.60 ±0.03c

6.02 ±0.04a

2.11 ±0.24b

65.14 ±0.63c

26.63 ±0.20c

0.32 ±0.04a

SSF 26.83 ±0.02a

6.18 ±0.04b

1.79 ±0.19a

72.76 ±0.77a

21.86 ±0.18a

0.25 ±0.02a

Control 37.06 ±0.04e

6.07 ±0.03a

1.96 ±0.15b

60.13 ±0.45b

24.46 ±0.22b

0.42 ±0.03b

Data are the mean ± SD (n = 3).Mean values within column with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged, SSF – solid state fermentation; Sh – S. hortensis; WHC – water holding capacity; Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei

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The supplementation of RCMP with SMF and SSF Sh in all cases increased the WHC of RCMP (from 2.77% to 18.54% of RCMP produced with SMF with P. acidilactici Sh and RCMP produced with SSF with P. pentosaceus Sh, respectively). The highest WHC of RCMP produced with SSF with P. pentosaceus Sh (71.28 ± 0.70) was established. The RCMP WHC was significantly influenced by the type of LAB applied for savory plant bioproduct fermentation (p ≤ 0.001) and by the savory plant fermentation method (p ≤ 0.0001). A moderate negative significant correlation between the WHC and the moisture content of RCMP (r = -0.528; p = 0.014) and a moderate positive significant correlation between the WHC of RCMP and the pH of savory plant bioproducts (r = 0.441; p = 0.004) was found.

In all the cases, RCMP produced with plant bioproducts has a lower tenderness as compared with control samples: the shear force of RCMP supplemented with fermented Sh was higher on average by 37.31% (RCMP produced with SSF Sh) and by 36.51% (RCMP produced with SMF Sh). The tenderness of RCMP was significantly influenced by the type of LAB applied for savory plant bioproduct fermentation (p ≤ 0.036). A moderate positive significant correlation between the tenderness and drip loss of RCMP (r = 0.461; p = 0.035) was found.

RCMP produced with SMF savory plants has a higher cooking loss as compared with the RCMP produced with SSF bioproducts (on average by 8.54%). SSF savory plant bioproducts reduced the cooking loss of RCMP by 11.02%. The RCMP cooking loss ranged from 19.98 ± 0.23% to 30.42 ± 0.33% (RCMP produced with SSF with P. acidilactici Sh and RCMP produced with SMF with P. acidilactici Sh, respectively).

Colour characteristics. Significant differences (p ≤ 0.05) of the RCMP

produced with different plant bioproduct colour parameters L*, a*, and b* were observed (Table 3.2.3.2.3.). In all the cases, the RCMP control samples showed the highest lightness (L* = 70.61 ± 2.17) and yellowness (b* = 15.17 ± 0.17), but the lowest redness (a* = 4.11 ± 0.43) as compared with RCMP samples produced with fermented Sh bioproducts.

RCMP produced with SSF Sh showed a lower lightness (L*) (ranged between 55.71 ± 1.33 and 56.84 ± 1.32), redness (a*) (ranged between 5.03 ± 0.48 and 5.16 ± 0.54) and yellowness (b*) (ranged between 13.58 ± 0.14 and 13.81 ± 0.14), while the RCMP produced with SMF Sh showed a higher lightness (L*) (ranged between 59.34 ± 1.41 and 60.27 ± 1.23) and redness (ranged between 5.60 ± 0.72 and 6.94 ± 0.72) as well as yellowness (ranged between 14.11 ± 0.17 and 14.39 ± 0.16). Thus, RCMP samples produced with Sh bioproducts fermented with L. sakei in SMF conditions had lower

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(a*) and (b*) values as compared to RCMP produced with SMF with P. acidilactici and P. pentosaceus Sh. The (L*), (a*) and (b*) values of RCMP were significantly influenced by the savory plant fermentation method (p ≤ 0.005). A very strong positive significant correlation between the (L*) and (b*) values of RCMP (r = 0.946; p = 0.0001) and a moderate positive significant correlation between the (L*) value and pH of RCMP (r = 0.514; p = 0.017) was found. Table 3.2.3.2.3. Colour parameters (L*, a* and b*) of RCMP produced with fermented Sh.

RCMP L* a* b* S. hortensis (3%)

Ls SMF 60.27±1.23b 5.60±0.61c 14.11±0.17b SSF 56.78±1.51a 5.16±0.54b 13.80±0.13a

Pa SMF 59.47±1.46b 5.94±0.60c 14.19±0.10b SSF 56.84±1.32a 5.03±0.48a 13.58±0.14a

Pp SMF 59.34±1.41b 6.94±0.72d 14.39±0.16b SSF 55.71±1.33a 5.13±0.55b 13.81±0.14a

S. hortensis (5%)

Ls SMF 58.58±1.32c 5.99±0.63c 13.48±0.13a SSF 54.76±1.39a 4.23±0.50a 13.75±0.14b

Pa SMF 56.03±1.57b 5.83±0.55c 13.77±0.18b SSF 55.48±1.40a 4.73±0.43b 13.96±0.16c

Pp SMF 58.36±1.23c 5.91±0.63c 13.69±0.15a SSF 54.58±1.08a 5.03±0.50b 13.74±0.13b

S. hortensis (7%)

Ls SMF 57.06±1.19c 5.08±0.43b 13.88±0.13b SSF 51.13±1.07a 3.58±0.40a 12.99±0.10a

Pa SMF 55.51±1.14b 5.26±0.53b 13.92±0.17b SSF 53.99±1.28b 4.10±0.39a 12.87±0.16a

Pp SMF 57.22±1.36c 5.14±0.47b 14.02±0.16b SSF 51.96±1.19a 3.53±0.41a 13.08±0.17b

Control 70.61±2.17e 4.11±0.43a 15.17±0.17c Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sh – S. hortensis; Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; L* – lightness; a* – redness (or –a* of greenness); b* – yellowness (or – b* of blueness)

The content of BAs in RCMP produced with fermented Sh. The BAs

content in RCMP produced with Sh bioproducts depended on the LAB strain used for savory plant fermentation, as well as on the fermentation technology (SSF or SMF) (Table 3.2.3.2.4.).

SPR and SPD were the main BAs in control meat samples (produced without Sh bioproducts), while PHE, PUT, CAD, and HIS were most

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abundant in RCMP produced with Sh bioproducts. Both SSF and SMF Sh bioproducts significantly (p < 0.05) reduced the SPR and SPD content in RCMP and increased the PHE, PUT, CAD, and HIS content as compared with control RCMP samples.

In most cases, Sh bioproducts increased the PHE content in RCMP (up to 0.83–3.22 mg/kg), except in RCMP produced with SMF with P. pentosaceus Sh and in RCMP produced with SSF with L. sakei Sh). A moderate positive significant correlation between the PHE and CAD content in RCMP (r = 0.457; p = 0.037) and a strong positive significant correlation between the PHE and SPD content in RCMP (r = 0.619; p = 0.003) were found. The PUT content increased in all RCMP samples produced with SMF and SSF with L. sakei, P. acidilactici, and P. pentosaceus Sh (from 0.81 to 1.56 times, from 2.66 to 2.69 times, from 2.58 to 4.49 times, respectively) as compared with control samples. The Sh bioproducts increased the CAD content in RCMP from 3514.29% (in RCMP produced with SMF with L. sakei Sh) to 5855.71% (in RCMP produced with SMF with P. acidilactici Sh). A moderate positive significant correlation between the CAD and PHE content in RCMP (r = 0.457; p = 0.037), a moderate positive significant correlation between the CAD and HIS content in RCMP (r = 0.457; p = 0.037), a strong positive significant correlation between the CAD and TYR content in RCMP (r = 0.588; p = 0.005), a strong negative significant correlation between the CAD and SPR content in RCMP (r = -0.890; p = 0.0001), a moderate negative significant correlation between the CAD content and the pH of RCMP (r = -0.512; p = 0.018) were found.

The concentration of HIS in RCMP produced with SSF Sh decreased from 9.41% to 37.46% (in RCMP produced with SSF with P. pentosaceus Sh and in RCMP produced with SSF with P. acidilactici Sh, respectively) as compared with the RCMP produced with SMF Sm. A moderate positive significant correlation between the HIS and CAD content in RCMP (r = 0.457; p = 0.037), a strong negative significant correlation between the HIS and SPR content in RCMP (r = -0.637; p = 0.002), a moderate positive significant correlation between the HIS and TRP content in RCMP (r = 0.590; p = 0.005) were found. The concentration of TYR in RCMP ranged from 0 mg/kg (in RCMP control samples) to 13.36 ± 0.12 mg/kg (in RCMP produced with SSF with L. sakei Sh). In all the cases, Sh bioproducts decreased the SPD and SPR content in RCMP (up to 0.89–3.02 mg/kg, and up to 0.17 mg/kg, respectively). A moderate positive significant correlation between the TYR and SPR content in RCMP (r = -0.564; p = 0.008), a strong positive significant correlation between the SPD and PHE content in RCMP (r = 0.619; p = 0.003), a very strong negative significant correlation between the SPD and CAD content in RCMP (r = -0.890; p = 0.0001), a

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strong negative significant correlation between the SPD and HIS content in RCMP (r = -0.564; p = 0.008), a moderate positive significant correlation between the SPD content and the pH of RCMP (r = 0.505; p = 0.020) and a moderate positive significant correlation between the TRP and HIS content in RCMP (r = 0.590; p = 0.005) were found.

The total BAs content significantly (p < 0.05) increased in RCMP produced with Sh bioproducts. The BAs content in RCMP produced with SSF with L. sakei, P. acidilactici and P. pentosaceus Sh was by 19.03%, 15.44% and 5.35% higher, respectively, compared to the RCMP produced with SSF Sh. Table 3.2.3.2.4. BAs content in RCMP produced with fermented Sh. Sample PHE PUT CAD HIS TYR SPD SPR TRP Total

S. hortensis (3%) Ls SMF 0.83

±0.06a 6.30

±0.12a 25.30 ±0.23c

40.10 ±0.52f

2.12 ±0.10a

0.93 ±0.10a

- 1.03

±0.10a 76.61c

Ls SSF - 12.18 ±0.18b

31.09 ±0.38d

29.39 ±0.40d

13.36 ±0.12c

3.02 ±0.02b

- 2.15 ±0.20b

91.19c

Pa SMF 3.22 ±0.12c

20.83 ±0.18c

41.69 ±0.39e

26.53 ±0.30d

3.22 ±0.04a

13.38 ±0.24e

- 0.94 ±0.01a

109.81d

Pa SSF 5.81 ±0.10d

21.01 ±0.28c

35.91 ±0.45d

19.30 ±0.20c

12.20 ±0.22c

0.89 ±0.11a

- - 95.12d

Pp SMF - 20.17 ±0.33e

38.62 ±0.41d

15.35 ±0.19c

11.03 ±0.10c

2.14 ±0.01b

0.17 ±0.01a

0.10 ±0.01a

87.58c

Pp SSF - 35.06 ±0.39d

25.53 ±0.30c

14.03 ±0.19c

6.14 ±0.09b

1.08 ±0.03a

0.17 ±0.01a

1.12 ±0.10a

83.13c

Control - 7.81 ±0.14a

0.70 ±0.02a

4.71 ±0.12a

- 5.54 ±0.18c

15.2 ±0.62c

- 33.96a

S. hortensis (5%) Ls SMF 2.11

±0.10b 15.39 ±0.20b

30.17 ±0.30e

10.36 ±0.13c

5.01 ±0.15b

0.16 ±0.01a

1.33 ±0.10b

0.28 ±0.01a

64.81c

Ls SSF 3.08 ±0.04c

22.05 ±0.25c

39.17 ±0.42f

15.30 ±0.19d

2.13 ±0.09a

3.11 ±0.02b

- 0.51 ±0.01a

85.35e

Pa SMF 1.45 ±0.01a

16.33 ±0.19b

25.63 ±0.23d

20.16 ±0.29e

5.31 ±0.15b

1.20 ±0.10a

0.34 ±0.01a

- 50.26c

Pa SSF 2.11 ±0.06b

20.58 ±0.29c

33.28 ±0.41e

10.34 ±0.10c

8.63 ±0.10c

2.08 ±0.04b

- - 77.02d

Pp SMF - 30.10 ±0.31d

26.19 ±0.28d

5.63 ±0.14b

7.64 ±0.18c

- 3.08 ±0.10b

0.27 ±0.01a

72.91d

Pp SSF 0.36 ±0.01d

35.17 ±0.39c

22.43 ±0.15c

2.06 ±0.07b

6.59 ±0.09a

0.85 ±0.01a

3.47 ±0.06a

1.09 ±0.17c

72.02d

Control - 7.81 ±0.14a

0.70 ±0.02a

4.71 ±0.12b

- 5.54 ±0.18c

15.2 ±0.62d

- 33.96a

S. hortensis (7%) Ls SMF - 39.62

±0.30e 21.37 ±0.25d

5.69 ±0.10a

- - 3.15 ±0.16b

0.39 ±0.01a

71.52c

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Continue of Table 3.2.3.2.4. Sample PHE PUT CAD HIS TYR SPD SPR TRP Total Ls SSF 1.30

±0.12a 35.42 ±0.38d

10.30 ±0.17c

13.15 ±0.15c

8.66 ±0.19c

3.17 ±0.10b

- 2.10 ±0.14c

72.80c

Pa SMF -

46.81 ±0.48f

25.33 ±0.28d

14.07 ±0.15c

2.13 ±0.08a

3.24 ±0.08b

0.81 ±0.01a

0.36 ±0.01a

96.0e

Pa SSF 3.25 ±0.13b

31.04 ±0.29d

30.25 ±0.30e

10.74 ±0.19b

5.69 ±0.10b

1.26 ±0.05a

- 0.41 ±0.01a

80.61d

Pp SMF 1.22 ±0.14a

20.86 ±0.26c

28.68 ±0.30e

7.48 ±0.10b

- 6.37

±0.10c 2.12

±0.08a 1.10

±0.03b 66.61c

Pp SSF -

22.17 ±0.25c

31.20 ±0.30e

17.30 ±0.17d

5.63 ±0.05b

4.21 ±0.02b

- 0.57 ±0.01a

82.44d

Control 1.36 ±0.12a

7.81 ±0.14a

0.70 ±0.02a

4.71 ±0.12a

- 5.54 ±0.18c

15.2 ±0.62e -

33.96a

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). RCMP – ready-to-cook minced pork meat products; SMF – submerged fermentation; SSF – solid state fermentation; Sh – S. hortensis; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine

3.2.3.3. Parameters of ready-to-cook minced pork meat products

produced with pea fiber and semolina

Quality parameters. The quality parameters of RCMP produced with pea fiber and semolina are presented in Figure 3.2.3.3.1. (a) Pea fiber increased the moisture content and drip loss of RCMP (by 13.9% and 4.3%, respectively) compared with the RCMP produced with semolina.

a) b) Figure 3.2.3.3.1. The quality parameters (a) and L*, a* and b*coordinates (b) of RCMP produced with pea fiber and semolina (Remark: RCMP – ready-to-cook minced pork meat products, L* – lightness, a* – redness (or – a* of greenness), and

b* – yellowness (or – b* of blueness). In RCMP samples, the pH ranged from 6.01 ± 0.04 to 6.07 ± 0.03

(RCMP produced with semolina and RCMP produced with pea fiber,

0

20

40

60

M pH DL WHC CL SF

RCMP with pea fiberRCMP with semolina

0

20

40

60

80

L* a* b*RCMP with pea fiber RCMP with semolina

110

respectively). Pea fiber has a negative influence on the WHC of RCMP (decreased it by 8.9%). The higher cooking loss was found in RCMP with semolina (25.19 ± 0.26) as compared with the RCMP produced with pea fiber (24.46 ± 0.22). The shear force of RCMP produced with pea fiber was 1.68 times higher than of RCMP produced with semolina.

RCMP with semolina showed a higher lightness (L*) (by 7.2%), redness (by 5.1%) and yellowness (by 0.5%) as compared with the RCMP produced with pea fiber (Figure 3.2.3.1 (b)).

The content of BAs in RCMP produced with pea fiber and semolina.

The BAs content (mg/kg) in RCMP produced with pea fiber and semolina is presented in Figure 3.2.3.3.2. SPR and SPD were the main BAs in the RCMP produced with pea fiber (10.29 ± 0.47 and 18.66 ± 0.53, respectively), while PUT and SPR were the main BAs in the RCMP produced with semolina (7.81 ± 0.14 and 15.20 ± 0.62, respectively). No PHE and TRP in RCMP samples were found. The CAD content in the RCMP produced with pea fiber was by 60% higher, and the HIS content by 17.4% lower as compared with the RCMP produced with semolina. The total BAs content in the RCMP produced with pea fiber was by 23.9% higher than in the RCMP produced with semolina.

Figure 3.2.3.3.2. The content of BAs (mg/kg) in RCMP produced with pea fiber and semolina. (Remark: BAs - biogenic amines, RCMP – ready-to-cook minced

pork meat products, PHE – phenylethylamine, PUT – putrescine, CAD – cadaverine, HIS – histamine, TYR – tyramine, SPD – spermidine, SPR – spermine, TRP – tryptamine).

3.3. The parameters of unripened curd cheese produced with Satureja

montana and Rhaponticum carthamoides bioproducts

Acidity parameters of UCC. The pH, TTA (T°) and the content of L(+) and D(-)-lactic acid isomers (g/100 g) in the UCC produced with fermented

0

5

10

15

20

PHE PUT CAD HIS TYR SPD SPR TRP

mg/

kg

RCMP with pea fiber RCMP with semolina

111

and nonfermented Sm and Rc bioproducts are presented in Table 3.3.1. In most of cases, the pH of the UCC produced with Sm bioproducts was higher than of the UCC produced with Rc bioproducts (except samples produced with SSF with P. acidilactici savory plants). The UCC pH ranged from 4.15 ± 0.02 to 4.76 ± 0.05 (UCC samples produced with SSF with P. acidilactici Rc and UCC samples produced with SMF with P. pentosaceus Sm, respectively). The pH of UCC was significantly influenced by the savory plant fermentation method (p ≤ 0.0001), the type of LAB applied for the savory plant fermentation (p ≤ 0.0001), and the type of plants (p ≤ 0.0001), and the action of these factors on the pH was significant (p ≤ 0.0001). A moderate negative significant correlation between the pH of UCC and of savory plant bioproducts (r = -0.430; p = 0.002) was found.

Table 3.3.1. Acidity parameters of UCC produced with Sm and Rc.

Param.

With fermented plants

L. sakei P. acidilactici 7 P. pentosaceus 8

SMF SSF SMF SSF SMF SSF

UCC produced with Sm

pH 24 4.68±0.05c 4.38±0.02a 4.54±0.04b 4.41±0.03a 4.76±0.05d 4.51±0.04b pH 48 4.21±0.03a 4.13±0.04a 4.38±0.03b 4.08±0.02a 4.52±0.04c 4.22±0.03a TTA 0.20±0.01b 0.10±0.01a 0.20±0.01b 0.10±0.03a 0.10±0.01a 0.10±0.02a L(+) 5.61±0.07c 7.44±0.09f 5.36±0.07b 6.74±0.01d 5.14±0.07b 5.95±0.08c D(-) 1.39±0.03a 1.17±0.02a 2.35±0.04c 1.94±0.03b 3.28±0.04d 2.47±0.03c

UCC produced with Rc

pH 24 4.38±0.03b 4.21±0.02a 4.42±0.04b 4.15±0.02a 4.44±0.04b 4.40±0.04b pH 48 4.18±0.02b 4.09±0.02a 4.32±0.03c 4.11±0.02a 4.34±0.03c 4.11±0.02a TTA 0.19±0.02b 0.11±0.01a 0.21±0.02b 0.12±0.01a 0.10±0.01a 0.10±0.01a L(+) 2.18±0.06a 5.44±0.07c 2.94±0.09b 5.44±0.07c 5.94±0.03d 4.94±0.03c D(-) 3.47±0.03c 1.30±0.02a 2.76±0.04b 2.45±0.03b 3.45±0.04c 2.68±0.04b

Control With NFP Control With NFP

UCC produced with Sm UCC produced with Rc

pH 24 4.79±0.06d 4.69±0.05c 4.54±0.05c 4.39±0.03b pH 48 4.53±0.05c 4.38±0.04b 4.39±0.03c 4.25±0.03b TTA 0.10±0.01a 0.10±0.01a 0.11±0.01a 0.12±0.01a L(+) 4.68±0.05a 5.24±0.06b 3.17±0.03b 2.00±0.04a D(-) 1.78±0.02b 2.14±0.03b 2.34±0.03b 2.79±0.04b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). SMF – traditional submerged fermentation; SSF – solid state fermentation; Control – UCC without plant bioproducts; UCC – unrippened curd cheese; Sm – S. montana; Rc – R. carthamoides; TTA – total titratable acidity, pH 24 – pH ater 24 h fermentation; pH 48 – pH after 48 h fermentation

112

Similar tendencies of the TTA of UCC were found. The TTA of UCC produced with Sm and Rc bioproducts ranged from 0.1 to 0.2 ºT. The TTA of UCC was significantly influenced by the savory plant fermentation method (p ≤ 0.0001), and the type of LAB applied for the savory plant fermentation (p ≤ 0.0001), and the interaction of these factors on the TTA of UCC was significant (p ≤ 0.0001).

The ratio of L(+)/D(-)-lactic acid isomers in the UCC produced with Sm bioproducts ranged from 4.0 to 6.3 (in UCC samples produced with SSF and SMF with L. sakei Sm), from 2.3 to 3.5 (in UCC samples produced with SSF and SMF with P. acidilactici Sm), and from 1.6 to 2.4 (in UCC samples produced with SSF and SMF with P. pentosaceus Sm), and in UCC with Rc bioproducts it ranged from 0.6 to 4.2 (in UCC samples produced with SSF and SMF with L. sakei Rc), from 1.1 to 2.2 (in UCC samples produced with SSF and SMF with P. acidilactici Rc) and from 1.7 to 1.8 (in UCC samples produced with SSF and SMF with P. pentosaceus Rc) with the L(+)-lactic acid as the predominant isomer. The L(+)/D(-)-lactic acid content was significantly influenced by the savory plant fermentation method (p ≤ 0.0001), the type of LAB applied for savory plant fermentation (p ≤ 0.0001), and the type of plant (p ≤ 0.0001).

Interaction of these factors had a significant influence (p ≤ 0.0001) on the L(+) and D(-) isomer content in the UCC. A weak negative significant correlation between the TTA and L(+) lactic acid content in the UCC (r = -0.360; p = 0.012), a moderate negative significant correlation between the L(+)-lactic acid and D(-)-lactic acid content in the UCC (r = 0.505; p = 0.0001) and a moderate positive significant correlation between the pH of savory plants and the L(+)-lactic acid content in UCC (r = 0.529; p = 0.0001) were found.

The content of VC in the UCC produced with Sm and Rc

bioproducts. VC in the UCC produced with 3% of Sm and Rc bioproducts are presented in Table 3.3.2.

VC in UCC enriched with 3% of Sm and Rc bioproducts significantly (p < 0.05) depended on the type of savory plant bioproducts used for the UCC production.

The results showed that thymol and carvacrol were dominant compounds in UCC samples produced with Sm bioproducts, and its content ranged from 11.12 g/100 g to 25.03 g/100 g (in UCC produced with SSF with L. sakei Sm and in UCC produced with SMF with P. acidilactici Sm, respectively), and from 12.10 to 34.02 g/100 g (in UCC produced with SSF with L. sakei Sm and in UCC produced with SMF with P. acidilactici Sm, respectively).

113

Table 3.3.2. VC (g/100 g) in UCC.

Compound

With fermented plants Control

With NFP L. sakei P. acidilactici

7 P. pentosaceus

8 SMF SSF SMF SSF SMF SSF

S. montana Propyl phenylacet.

0.02 ±0.01a

- 0.51

±0.07b - - -

0.19 ±0.01a

-

Isoamyl acetate

0.08 ±0.02a

0.25 ±0.03a

- 0.74

±0.01c 0.38

±0.01b 0.50

±0.01c 0.05

±0.01a -

Heptanone 0.43

±0.02a 0.36

±0.02a 0.88

±0.08c 0.72

±0.10b 0.38

±0.01a 0.50

±0.01b 0.79

±0.11b 0.77

±0.09b Phellandrene - - - - - - - -

α-pinene 0.05

±0.01a -

0.42 ±0.05b

- 0.09

±0.02a -

0.64 ±0.21b

0.49 ±0.01b

Mircene 0.43

±0.18b - - - -

0.04 ±0.01a

3.59 ±0.10d

2.81 ±0.20c

d-limonene 30.64 ±0.25f

4.87 ±0.10d

0.40 ±0.05a

0.17 ±0.02a

0.22 ±0.01a

5.21 ±0.09d

0.77 ±0.13a

0.63 ±0.11a

n-dekan 0.06

±0.01a -

0.07 ±0.02a

- - - 0.19

±0.01b 0.14

±0.03b

Thymol 14.02 ±0.09b

11.12 ±0.18a

25.03 ±0.19d

17.06 ±0.26b

11.75 ±0.14a

13.14 ±0.17a

0.15 ±0.02a

29.09 ±0.27e

Carvacrol 26.01 ±0.15c

12.10 ±0.22a

34.02 ±0.30e

14.21 ±0.21a

21.72 ±0.22c

23.19 ±0.34c

- 34.23 ±0.35e

Tetrahydro-furyl alcohol

1.09 ±0.23d

1.46 ±0.10e

0.62 ±0.01b

0.79 ±0.03b

0.33 ±0.01a

0.86 ±0.04b

0.24 ±0.05a

0.32 ±0.09a

R. carthamoides

Isoamyl acet. 0.06

±0.02a 0.39

±0.10b 0.04

±0.01a 0.63

±0.05c 0.21

±0.15b 0.44

±0.15b 0.05

±0.01a -

Heptanone 0.02

±0.01a 0.31

±0.09c 0.17

±0.01b 0.12

±0.01b 0.03

±0.01a 0.11

±0.02b 0.79

±0.09d

Phellandrene 6.59

±0.10b 6.46

±0.10b -

15.15 ±0.05e

2.18 ±0.01a

7.62 ±0.06b

0.64 ±0.08a

-

8.53 ±0.20c

ρ-cymene 18.26 ±0.20d

10.22 ±0.05c

14.66 ±0.09c

26.59 ±0.19e

36.31 ±0.31f

25.38 ±0.32e

0.15 ±0.01a

35.15 ±0.18f

d-limonene 5.02

±0.10c 3.54

±0.03b 1.57

±0.02a 12.03 ±0.09d

- 5.98

±0.08c 0.77

±0.06a 7.49

±0.09c

Thymol 29.25 ±0.29c

14.17 ±0.14a

39.28 ±0.38d

15.24 ±0.14a

24.98 ±0.24c

28.51 ±0.20c

- 32.56 ±0.18c

Tetrahydro-furyl alcohol

0.94 ±0.10b

1.10 ±0.09b

0.43 ±0.26a

0.62 ±0.13a

0.27 ±0.01a

0.51 ±0.05a

0.24 ±0.05a

0.32 ±0.01a

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05).; SMF – traditional submerged fermentation; SSF – solid state fermentation; Control – UCC without plant bioproducts; UCC – unrippened curd cheese; NFP – nonfermented plants

114

Thymol, p-cymene and phellandrene in the UCC produced with Rc bioproducts ranged from 14.17 to 39.28 g/100 g (in the UCC produced with SSF with L. sakei Rc and in the UCC produced with SMF with P. acidilactici Rc), from 10.22 to 36.31 g/100 g (in the UCC produced with SSF with L. sakei Rc, and in the UCC produced with SMF with P. pentosaceus Rc), from 0 to 15.15 g/100 g (in the UCC produced with SMF with P. acidilactici Rc and in the UCC produced with SSF with P. acidilactici Rc).

The sensory properties. The sensory properties of the UCC produced

with Sm and Rc bioproducts, determined by the evaluation of the facial expression intensity of panelists, are presented in Table 3.3.3. It was found that the emotion “happy” was intensively expressed in case of the tested samples which were evaluated by panelists as the samples of the highest acceptability according to the score system. Table 3.3.3. The results of the sensory properties of UCC produced with Sm and Rc bioproducts determined with FaceReader software.

Samples Facial expressions recorded during curd cheese tasting time Happy Sad Angry Surprised Scared Disgusted

S. montana UCC 4.27×102d 6.60×102c 3.59×102b 6.88×103c 1.54×104a 3.43×104b NF Sm 4.28×102d 6.16×102c 6.80×102e 2.01×102b 2.5×103b 7.16×104d SMF Ls 3.69×102c 3.84×102b 4.88×102c 1.01×102a 5.03×103c 4.34×104c SSF Ls 2.70×102b 7.89×102d 1.05×102a 1.06×102a 3.30×103b 3.98×104b SMF Pa 2.80×102b 6.81×102c 1.17×102a 2.46×102b 7.51×103d 8.31×104e SSF Pa 4,44×102e 7.52×102d 6.15×102e 9.67×102d 7.47×103d 3.98×104b SMF Pp 4.42×102e 8.57×102d 5.04×102d 5.03×102c 1.93×104a 2.16×104a SSF Pp 2.18×102a 1.18×102a 4.31×102c 8.73×103d 1.44×104a 3.40×104b

R. carthamoides UCC 4.25×102b 5.79×102c 3.41×102b 6.24×103c 1.23×104a 2.43×104a NF Rc 4.33×102b 6.43×102c 5.46×102c 7.34×102c 2.21×104b 6.44×104c SMF Ls 3.64×102b 2.94×102b 5.26×102c 2.14×103a 6.04×103c 3.67×104b SSF Ls 2.61×102a 3.54×102b 2.03×102a 1.17×103a 2.14×104b 4.33×104b SMF Pa 3.17×102b 7.79×102d 1.33×102a 9.78×102d 8.36×103d 6.79×104c SSF Pa 5.02×102c 6.87×102c 5.89×102c 8.33×102d 7.11×103d 4.05×104b SMF Pp 4.18×102b 7.62×102d 5.33×102c 4.94×102b 1.59×104a 1.98×104a SSF Pp 2.31×102a 1.12×102a 3.91×102b 6.55×103c 1.27×104a 3.86×104b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). SMF – traditional submerged fermentation; SSF – solid state fermentation; Control – UCC without plant bioproducts; UCC – unrippened curd cheese; Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei, NF – nonfermented plant

115

The UCC produced with SMF and SSF with P. acidilactici Sm (4.3–4.8 scores), and the UCC produced with SMF and SSF with L. sakei Rc (4.3–4.8 scores) were indicated as having the highest overall acceptability (Figure 3.3.1). The lowest overall acceptability of the UCC control samples (3.8 scores) was found. A higher overall acceptability had the UCC produced with SMF and SSF with L. sakei, P. acidilactici, P. pentosaceus Sm and Rc bioproducts on average by 7.1%, 14.4%, 9.1% and 15.4%, respectively, as compared with control samples. A strong negative significant correlation was found between the overall acceptability of the UCC and its pH (r = -0.738; p = 0.0001). Also, significant positive and negative correlations between the score system of the sensory analysis and the emotions “happy” and “sad” by the FaceReader software (r = 0.744; p = 0.001 and r = 0.585; p = 0.002, respectively) were determined.

Figure 3.3.1. The overall acceptability of UCC produced with Sm and Rc bioproducts determined in 5 point system (Remark: Pa – P. acidilactici, Ls – L. sakei, Pp – P. pentosaceus, SMF – traditional submerged fermentation, SSF – solid state

fermentation, UCC – unripened curd cheese, Sm – S. montana; Rc – R. carthamoides, NF – UCC with nonfermented plant).

The BAs content in UCC produced with Sm and Rc bioproducts. The

BAs content (mg/kg) in UCC samples is presented in Table 3.3.4. The fermentation conditions of the used savory plants have an influence on the formation of the BAs in UCC samples. The BAs content significantly depended on savory plant fermentation method, the type of LAB applied for the fermentation, and the type of the plant (p ≤ 0.0001).

The main BA in the UCC produced with Sm and Rc bioproducts was CAD. The concentration of CAD in UCC varied from 3.55 to 137.6 mg/kg (in the UCC produced with SMF with L. sakei Rc and in the UCC produced with SSF with L. sakei Rc, respectively), and it significantly depended on the LAB strain used for savory plant fermentation and the fermentation

0123456

SMF SSF SMF SSF SMF SSF

Ls Pa Pp NF

Scor

e(m

in 0

; m

ax 6

)

S. montana R. carthamoides UCC

116

technology (SSF or SMF) (p ≤ 0.0001). A weak positive significant correlation between the CAD and PHE content in the UCC (r = 0.311; p = 0.031), a moderate negative significant correlation between the CAD and TYR content in the UCC (r = -0.444; p = 0.002), a strong positive significant correlation between the CAD and SPD content in the UCC (r = 0.738; p = 0.0001) and a weak positive significant correlation between the CAD content in the UCC and the pH of savory plant bioproducts (r = 0.371; p = 0.009) were found.

The concentration of HIS in the UCC produced with Sm and Rc bioproducts ranged from 0.54 ± 0.01 mg/kg to 9.36 ± 0.06 (with SMF with L. sakei Sm and with SMF with P. acidilactici Sm, respectively). In most of the cases, Sm and Rc bioproducts increase the HIS content in the UCC.

The HIS content in the UCC ranged from 4.5% to 78% (in the UCC produced with SMF with L. sakei Sm and in the UCC produced with SMF with P. acidilactici Sm, respectively) and from 12.9% to 51.3% (in the UCC produced with SMF with L. sakei Rc and in the UCC produced with SSF with P. acidilactici Rc) higher as compared with control samples (except the UCC produced with SSF with L. sakei Sm and Rc and in the UCC produced with SMF with P. pentosaceus Sm and Rc). A weak negative significant correlation between the HIS content in the UCC samples and the pH of savory plant bioproducts (r = -0.329; p = 0.023) was found.

Table 3.3.4. The BAs content (mg/kg) in UCC. Param.

With fermented plants Control With

NFP L. sakei P. acidilactici 7 P. pentosaceus 8

SMF SSF SMF SSF SMF SSF UCC produced with S. montana

PHE 34.10 ±0.14f

10.17 ±0.08d

1.36 ±0.02a

5.38 ±0.16c

3.18 ±0.03b

2.14 ±0.03a

3.55 ±0.04b

3.14 ±0.04b

PUT 13.09 ±0.09c

5.41 ±0.09b

0.14 ±0.01a

15.5 ±0.17d

13.33 ±0.09c

15.86 ±0.09d

- 1.87

±0.02a CAD 95.6

±0.34e 12.3

±0.07a 117.8 ±0.37f

40.6 ±0.23c

114.0 ±0.36f

90.21 ±0.30e

- -

HIS 0.54 ±0.01a

- 9.36

±0.06c 4.62

±0.22b -

2.47 ±0.18b

0.12 ±0.01a

25.17 ±0.13e

TYR -

0.17 ±0.01a

- - - - 1.10

±0.02b -

SPD 2.03 ±0.09b

- 1.64

±0.02a -

2.29 ±0.02b

0.93 ±0.01a

- -

SPR - - - - - - - - TRP

- - - 1.21

±0.02a 2.61

±0.02b - - -

Total 145.37f 28.07b 130.3f 67.31c 135.42f 111.61e 4.77a 30.18b

117

Continue of Table 3.3.4.

Param. With fermented plants

Contr. With NFP L. sakei P. acidilactici 7 P. pentosaceus 8

SMF SSF SMF SSF SMF SSF UCC produced with S. montana

PHE 27.3 ±0.15f

0.14 ±0.06a

2.04 ±0.11b

5.38 ±0.16c

2.22 ±0.10b

0.11 ±0.03a

- 0.07

±0.03a PUT 3.09

±0.07a - -

15.5 ±0.17d

3.11 ±0.09a

7.36 ±0.13b

- 2.61

±0.11a CAD 137.6

±0.23f 3.55±0.14a

94.3 ±0.16e

40.6 ±0.23d

98.16 ±0.27e

82.9 ±0.14e

- 100.3 ±0.25e

HIS 1.16 ±0.11b

- 1.46

±0.13b 4.62

±0.22d -

3.54 ±0.18c

0.09 ±0.02a

3.17 ±0.24c

TYR - - - - - - - - SPD 2.03

±0.09b -

2.31 ±0.11b

- - 1.38

±0.11a 0.25

±0.04a -

SPR - - - - - - - - TRP 0.09

±0.03a -

0.17 ±0.05a

- 5.66

±0.11c 0.17

±0.10a - -

Total 171.27f 3.69a 100.28d 66.10c 109.15d 95.46c 0.34a 106.16d Data values were expressed as mean ± SD (n = 3). SMF – traditional submerged fermentation; SSF – solid state fermentation; Control – UCC without plant bioproducts; UCC – unrippened curd cheese; Sm – S. montana; Rc – R. carthamoide; PHE – phenylethylamine; PUT – putrescine; CAD – cadaverine; HIS – histamine; TYR – tyramine; SPD – spermidine; SPR – spermine; TRP – tryptamine; NFP – nonfermented

The Sm and Rc bioproducts increased the PUT, CAD, SPD and TRP

content in the UCC on average by 100% (except the SPD content in the UCC produced with Rc bioproducts) as compared with control samples. A weak negative significant correlation between the PUT and TYR content in the UCC (r = -0.294; p = 0.043) and a moderate positive significant correlation between the PUT content in the UCC and the pH of savory plant bioproducts (r = 0.505; p = 0.0001), a moderate positive significant correlation between the SPD and PHE content in the UCC (r = 0.422; p = 0.003), a strong positive significant correlation between the SPD and CAD content in the UCC (r = 0.738; p = 0.0001) and a weak positive significant correlation between the SPD content in the UCC and the pH of savory plant bioproducts (r = 0.371; p = 0.009) were found.

No SPR was detected in all UCC samples. The total BAs content in the UCC produced with SSF with L. sakei, P. acidilactici and P. pentosaceus Sm bioproducts was 5.2, 1.9 and 1.2 times lower, respectively as compared with the UCC produced with the SMF Sm. Similar tendencies of the UCC produced with SSF with L. sakei, P. acidilactici and P. pentosaceus Rc (the

118

total BAs content was 46.4, 1.5 and 1.1 times lower, respectively, than in the UCC produced with SMF Rc bioproducts) were found.

Antimold activity of Sm and Rc bioproducts. The effect of Sm and Rc

bioproducts on the growth of mold on the UCC surface during 11 days of storage is presented in Table 3.3.5. It was found that the Sm and Rc bioproducts derived from a combination of LAB and savory plants increased the shelf life of the UCC. The longest shelf life was found for the UCC produced with L. sakei fermented Sm bioproducts, and it was the shortest for the control UCC samples.

Table 3.3.5. The effect of Sm and Rc bioproducts on the growth of mold on the UCC surface during 11 days of storage.

Storage time, days

L. sakei and plant bioproducts

Dried plants Control

Sm Rc Sm Rc 4 - - - - - 5 - - - + + 7 - - + + + 9 - + + + + 11 + + + + ++ Data values were expressed as mean ± SD (n = 3). Control – UCC without plant bioproducts; UCC – unrippened curd cheese, Sm – S. montana; Rc – R. carthamoides; (-) no mold; (+) the first traces of mold; (++) large mold colonies.

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4. DISCUSSION

4.1. Lactic acid bacteria – plant bioproducts – alternative preservatives for a higher value food of animal origin production

The nutritional strategies to improve the quality of food products of

animal origin are a relatively new approach which emerged at the interface of animal nutrition, food science and human nutrition. This approach has been effectively used to alter the animal product composition to be more consistent with human dietary guidelines [202]. The natural way to improve food quality is the use of LAB. Fermentation as a process for manufacturing foods has traditionally been used to preserve perishable products and to enhance their nutritional value. In contrast to the western countrie, SSF has been developed in eastern countries over many centuries, and is widely used in these regions ([384]. SSF is involving microorganisms grown on solid or semi-solid substrates or supports and is more effective than the liquid phase SMF, because lower contents of water and energy are used for product stabilisation by using dehydration. LAB including bacteriocin-like inhibitory substances (BLIS) producing strains are GRAS and have received significant attention as a novel approach to the control of pathogens in foods [247, 266]. They are generally accepted as beneficial to the host, and their presence is directly influenced by the ingestion of fermented food or probiotics [10]. However, LAB are very sensitive to the chemical composition of the fermentable substrate. In our study, the microbiological analysis of 48 h fermented Sm, Sh, Rc, defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke has shown that the plants used in experiment are a suitable substrate for selected LAB SMF and SSF fermentation. The total count of LAB in fermented plant products depended on the LAB strain, the type of fermentation (SMF and SSF) and the plant species used, and varied from 6.21 log10 CFU/g (in with P. acidilactici SMF Jerusalem artichoke) to 9.98 log10 CFU/g (in with P. pentosaceus SSF defatted soy flour). There are some disagreements as to the effect of the moisture content of fermentation medium on the growth of LAB. Wang et al. [397] have reported that LAB survive better at a low water activity, although [139] have reported that a medium with a high water content is more suitable for the propagation of LAB due to the solubility of nutrients in the medium.

LAB play the key role in several food fermentations producing lactic acid besides other metabolites [315]. High counts of viable LAB are necessary to get the desired reduction in the pH, which not only affects the product

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organoleptic properties but also prolongs shelf-life and prevents product contamination [314]. The antimicrobial activity of metabolites, such as organic acids, bacteriocins, cyclic dipeptides and TPC produced by LAB, which belong to L. sakei, P. pentosaceus and P. acidilactici species, were reported by other researches as well [237, 324]. The results of this study are in agreement with the results of other studies. This proves that LAB initiate a rapid and adequate acidification in a fermentable substrate during the production of various organic acids from carbohydrates, decrease the pH, and increase the TTA [293, 332]. John et al. [172] proved that the pH set to 6.5 was the optimum for the lactic acid production. In this study, the pH of SSF savory plants, defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke was lower than 6.27. The pH of plant bioproducts ranged from 4.30 (of SMF with P. acidilactici Jerusalem artichoke) to 6.46 (of SMF with P. pentosaceus defatted soy flour). The decline in pH during fermentation was known to act as a preservative factor against bacteria associated with spoilage and against non-desirable and pathogenic microorganisms producing foodborne toxins which are not able to proliferate under low pH conditions [42, 378]. In contrast to the pH, the TTA of fermented SMF Sm, Sh, Rc, defatted soy flour, pea fiber and Jerusalem artichoke was higher on average by 6.0%, 12.7%, 13.8%, 38%, 11.1%, and 146.6% (except flaxseed) as compared with SSF.

Microorganisms growing under extreme conditions (SSF) produce higher amounts of lactic acid, yet LAB with the amylolytic effect, applied in the industry, tend to produce stereospecific lactic acid isomers [316]. During fermentation, LAB may produce a mixture of L(+) and D(-) depending on the LAB strain [114]. The Lactobacillus species have an excellent potential as biocatalysts for L(+) lactic acid production [42], whereas in our experiment the LAB used for the fermentation of selected plants produced a mixture of L(+) and D(-)-lactic acid, and the main isomer was L(+). SSF savory plants, defatted soy flour, pea fiber, flaxseed, and Jerusalem artichoke had a higher (on average by 4.0%, 20.2%, 18.8%, 10.8%, 5.2%, 16.8%, and 1148.9%, respectively) content of L(+) lactic acid as compared with SMF bioproducts. Generally, the tested L. sakei, P. acidilactici and P. pentosaceus strains form L(+) lactic acid higher by 18.2–108.9% (in Sm), by 4.8–78.7% (in Sh), by 3.5–99.7% (in Rc), by 15.9–137.9% (in defatted soy flour), by 62.8–164.5 (in pea fiber), by 44.3–893.6% (in flaxseed), and by 53.4–1461.1% (in Jerusalem artichoke) as compared with the D(-)-lactic acid. The LAB production of L(+) and D(-)-lactic acid depends on the microorganism, the substrate and the growth conditions [373]. The concentration of the D(-)-lactic acid in all of the analysed plants and plant products was far below the levels causing a health risk, as was noted by the

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[101]. The FAO/WHO Experts Committee on Food Additives recommends limiting the daily intake of D(-)-lactate to 100 mg per kilogram of body weight and attempts to favour the use of the L(+)-isomer in fermented food [101].

Another way to improve the food of animal origin quality and safety parameters is the use of plants having functional or/and antimicrobial properties, which are potential sources of natural antioxidants: TPC, lignans, prebiotics, etc. In food systems, they can improve the flavour, suppress lipid oxidation-induced food deteriorations, inhibit the growth of microorganisms and a play role in decreasing the risk of some diseases [350]. The EOs compounds of S. montana L. and R. carthamoides CD. show a different biological activity [249, 355]. The antimicrobial and antioxidant activity of EOs is due to their chemical composition which includes volatile bioactive compounds able to control pathogenic and spoilage microorganisms [124, 130, 359]. Generally, the high antimicrobial activity of Satureja montana L. EOs can be attributed to major compounds such as carvacrol, thymol, terpinen-4-ol and linalool [87, 250]. R. carthamoides CD. is considered to be highly promising in developing new classes of biologically active food additives [44].

This research revealed the antimicrobial activity of Sm, Sh and Rc fermented with selected LAB. Plants of the Satureja species possess strong antibacterial activities of different extents against microorganisms responsible for food spoilage, such as Salmonella, Listeria, Staphylococcus E. coli, P. fluorescens, and B. subtilis. The positive effect of LAB was achieved due to the synergetic activity of bacteriocins producing LAB and antimicrobial plant compounds. The highest antimicrobial activity against P fluorescens was shownby Sm bioproducts fermented with P. acidilactici (with the inhibition zone up to 18.5 mm), which was achieved through the synergetic activity between the savory plants and LAB. The results are in agreement with other studies demonstrating that LAB can be employed as bioprotective cultures to improve the microbial safety of foods [73] with no major detrimental effect on textural or physicochemical properties [297]. The volatile fractions of the Rhaponticum and Satureja genera exhibited antibacterial and antifungal properties [39, 84, 194, 375] and could serve as a source for antimicrobial agents against food pathogens as well [293].

4.2. Changes of bioactive compounds in plants during lactic acid

fermentation process

The functional plant materials come from a wide variety of plant sources which provide important nutraceutical compounds that may be used in food

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systems [216]. Bran, a byproduct of the milling industry, is to be efficiently utilized for human consumption [187]. Numerous epidemiological studies have shown that diets low in fat and rich in complex carbohydrates from vegetables, fruits, and grains are associated with a decreased risk of chronic diseases [321]. International epidemiological comparisons have linked the semi-vegetarian diet in some Asian countries with a reduced incidence of these diseases (i.e. the major hormone-dependent cancers, colon cancer, and coronary heart disease), indicating that some non-nutrient compounds in this diet may contribute to homeostasis and thus have a role in the maintenance of health. One of these non-nutrient groups of compounds are lignans detected and identified in human body fluids. Lignans are diphenolic compounds in plant foods, which belong to the group of phytoestrogens. Their molecular weight and structure are similar to those of steroids, implying that they could be important dietary modulators of the human hormonal system [6, 80]. The health effects of lignans depend both on the amount consumed and the bioavailability [29]. Our study showed that after fermentation the content of MAT increased (from 7.9 to 35.4%) in all pea fiber samples. In all the cases, a higher content of MAT was found in SSF samples as compared with SMF. The highest SECO concentration was found in SSF with the P. acidilactici pea fiber (140.3 μg/100g). Numerous lignans have been identified, and it is not known which of them are converted to enterolignans [356, 357]. For about two decades, only SECO and MAT have been to be the precursors of enterolignans. Lignans are closely associated with the dietary fibre matrix of the plant-derived food; thus, it is possible that their composition might influence the availability of lignans. The use of LAB for the fermentation of plant material could improve the digestibility [274] and conversion of lignans into the enterolignans which have a positive influence on human health.

Another group of biologically active compounds in plants, which was investigated in our study, is ARs. ARs are phenolic lipids present in the outer layer of mainly whole grain (> 500 μg/g) [80]. Cereals constitute a major source of dietary carbohydrates in Western countries, however, cereals are often consumed as refined products, thereby lacking ARs and other compounds associated with the bran. ARs are among the bioactive compounds in whole grain, which could play a role in the protective effect of whole grain products regarding the diabetes risk [125]. Also, the high concentration of plasma ARs was associated with a lower incidence of distal colon cancer [193]. In most cases, fermentation decreased the ARs content in plant samples, and the highest ARs content was found in nonfermented pea fiber (267 μg/g). The use of LAB fermentation reduces the ARs content by 40% (in SSF with P. acidilactici) to 73% (in SSF with P. pentosaceus).

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Also, fermentation reduces the content of β-glucans (SMF and SSF decreased the cotent of β-glucans in the pea fiber on average by 15.1% and 17.0%, respectively, as compared with nonfermented samples) in a fermentable substrate. Results of numerous studies showed different beneficial effects of β-glucans on humans and animals and recommended the intake of this compound with food at the level of at least 3 g/d [269].

β-glucans possess the free RSA [88, 248, 295, 336, 386]. The TPC content in SSF pea fiber increased from 0.6% to 10.7% compared with SMF samples. Our results are in agreement with the data of [58, 313] who have stated that SSF increase the TPC content in certain food products, thus enhancing their antioxidant activity. The TPC content and the RSA of pea fiber were significantly correlated and provided a strong evidence that the antioxidant activity is derived from the TPC content. SSF with selected LAB increase the RSA of plant bioproducts (from 3.4% to 7.2%) as compared with SMF. The increased inhibition percentage of free radical formation in fermented samples than in the nonfermented ones was supported by many other findings [89, 173]. Also, fermented plant products have a big potential as a technological source of amylolytic and proteolytic enzymes for food production. SFF plant bioproducts have a higher activity of amylolytic enzymes than SMF (with L. sakei fermented pea fiber 1.06 times higher, with L. sakei fermented defatted soy flour 3.90 times higher; with P. acidilactici fermented flaxseed and defatted soy flour 2.10 and 5.98, respectively, times higher; with P. pentosaceus fermented Jerusalem artichokes and flaxseed 1.13 and 2.57, respectively, times higher). Higher amylolytic activity of Jerusalem artichokes fermented with P. pentosaceus, flaxseed fermented with P. acidilactici and P. pentosaceus, pea fiber fermented with L. sakei and defatted soy flour fermented with L. sakei and P. acidilactici was estimated by applying SSF (1.1; 2.1; 2.6; 1.1; 3.6 and 6.9 times higher, respectively) as compared with SMF samples. In all SSF plant bioproducts a lower proteolytic enzymes activity as compared with SMF was estimated, and it varied from 309.1 AU/g to 1178.3 AU/g in flaxseeds fermented with L. sakei and in Jerusalem artichokes fermented with P acidilactici, respectively. It has been estimated that SSF technological schemes enable the output of products with a less proteolytic activity, whereas the amylolytic activity depended more on the fermented substrate that on its moisture. Microorganisms with a high proteolytic activity may be improper for the material fermentation of albuminous plants (defatted soy flour, pea fiber), as it can initiate BAs formation. According to the obtained results, it could be stated that SMF is more appropriate for carbohydrate substrate fermentation and SSF for albuminaceous plants. The development of food additives, whose technology is based on fermentation with LAB,

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demands to ensure the safety of new products in accordance with the formation of BAs. BAs formation through the microbial decarboxylation of amino acids is dependent on the specific bacterial strain(s) present, the level of decarboxylase activity, and the availability of the amino acid [361]. SSF increase the total BAs content in defatted soy flour, pea fiber, and Jerusalem artichoke on average by 19.7%, 4.2%, 2.1%, respectively, as compared with SMF. A lower BAs content in the SSF flaxseed as compared with SMF was estimated. The amount and types of BAs formed in fermented food products are strongly influenced by the food composition, microbial flora, environment and the other parameters that allow bacterial growth during food processing and storage [47]. The most important BAs, HIS, TYR, TRP, PUT, and CAD are formed from free amino acids, namely histidine, tyrosine, tryptophane, ornithine and lysine, respectively. SPD and SPR arise from putrescine [40, 104, 408]. The toxicological level of BAs is very difficult to establish because it depends on individual characteristics and the presence of other amines [61, 157, 206], etc. PUT and CAD can enhance HIS toxicity by inhibiting the intestinal HIS metabolizing enzyme, including diamine oxidase [17]. For this reason, the BAs content in the final product should be controlled.

4.3. Marinades based on lactic acid bacteria cultivated in an alternative substrate for improving meat quality parameters indifferent

part of pork, beef and chicken

The use of LAB as technological ingredients and biological preservatives

in meat products could confer the health benefits to consumers. The preservative ability of LAB in foods is attributed to the production of antimicrobial metabolites including organic acids and bacteriocins [98]. The LAB fermentative metabolism prevents the development of spoilage and pathogenic microflora by acidification of the product, also contributing to its colour stabilization and texture improvement [276]. The previous research confirmed that the metabolites of L. sakei KTU05-6, P. acidilactici KTU05-7, P. pentosaceus KTU05-8, KTU05-9 and KTU05-10 inhibited the growth of pathogenic bacteria belonging to the genera Bacillus, Pseudomonas, Listeria and Escherichia [59].

Trends in meat industry are focalizing in products with high organoleptic standards, textures, long shelf-life, and containing specific nutrients to cover special consumer requirements [98]. In the last decade, marinating of fresh meat has increased in popularity and has now become a standard practice for some product lines [298]. Marination is a common technique used to

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improve meat quality attributes such as WHC, tenderness and flavor. The colour, WHC and tenderness of meat are primary determinants of its visual and sensory appeal. WHC is important because of its role in moulding muscle structure and the consequent effects on quality. There is a correlation between the WHC of raw meat, the water loss during cooking, and the juiciness of the final product. During cooking, the loss of water from the structure has an inverse relationship to juiciness, and there appears to be a positive correlation which exists between sensory juiciness and tenderness [163]. The treatment of pork and beef neck, ham muscle, M. longissimus dorsi, and loin samples with P. acidilactici, P. pentosaceus and L. sakei marinades lowered the moisture content of meat on average by 5.3%, 5.6%, 26.6%, and on average by 11.3%, 15.3%, 12.7%, respectively, as compared with control samples. In contrast, the treatment of organic produced chicken breast, drumstick, and thighs with P. acidilactici marinade increased the moisture content of meat by 3.1%, 0.7% and 0.7% and of conventionally produced chicken breast, drumstick and thighs by 3.0%, 0.4% and 1.1%, respectively, as compared with control samples.

The pH value is a fundamental datum to be monitored during marination. Previous studies have reported that the use of organic acids, including citric and lactic acids, within marinades leads to a decrease in the pH value of marinated meat [188]. Marinated pork and beef meat with all marinades lowered the pH of meat on average by 9.1%, 11.0%, 15.2% and on average by 16.36%, 13.7%, 10.2%, respectively, as compared with the control samples. The treatment of organic and conventionally produced chicken meat with the P. acidilactici marinade lowered the pH of meat on average by 11.7% and increased drip loss on average by 12.5% and 3.6%, respectively, as compared with the control samples.

The evaluation of drip loss and pH have raised great attention of both producers and consumers. The occurrence of increased drip loss as well as pH variation to a large degree result in the degradation of other quality attributes such as tenderness, juiciness, and flavour thereby influencing the consumers’ acceptance and willingness. Drip loss is of importance in meat production due to the fact that it directly relates to sensory and other physicochemical attributes such as WHC. Drip loss can cause sensory loss including texture and flavour. The increase of drip loss is closely consistent with a lower WHC which is mainly caused by fibre shrinkage, cell damage, lower protein solubility, protein denaturation and aggregation taking place during freezing and thawing. Increasing drip loss also leads to the loss of some water-soluble nutrients such as proteins and amino acids [147]. In our study marination increased the drip loss of all pork and beef meat samples, and it ranged from 4.9% (in beef loin marinated with L. sakei) to 110.5% (in

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pork neck marinated with P. acidilactici). P. acidilactici, P. pentosaceus and L. sakei marinades increased the cooking loss of pork and beef meat on average by 24.5%, 23.9%, 20.8% and on average by 2.6%, 1.5%, 0.81%, respectively, as compared with control samples. The WHC of pork and beef meat was reduced on average by 9.8%, 16.9%, 14.21% and on average by 1%, 0.5% and 1.2%, respectively, as compared with control samples. A slight increase in WHC (0.2% and 0.3%) and cooking loss (2.0% and 3.0%) was found of organic and conventionally produced chicken meat after marination with P. acidilactici.

It is important to predict the increase of meat WHC because it is responsible for weight loss in raw, cooked and processed meats [328]. The main cause of the quality defect is the denaturation of sarcoplasmic proteins and myosin, leading to a decrease in the water-binding capacity of the protein sand and, as a result,to a decrease of the WHC of the meat. Several studies have suggested that tenderness is directly influenced by WHC [26]. Ke et al. [188] have suggested that tenderness is related to the pH of the muscle. The low muscle pH induced by marination resulted in the increased binding of water, as the percentage of bound water decreased when the muscle pH was lower than normal (pH 5.26). The continual increase in tenderness after the initial benefits of acid marination is attributed to an enhanced release of cathepsin enzymes. This allows the enzymes to increase the degradation of the myofibrillar proteins. Tenderness has been identified as one of the most important characteristics that determine the consumer’s eating satisfaction [144].

At a lower pH, meat tends to have a better textural quality. An adequate decrease in pH is directly related to the colour, tenderness and capacity of the muscle to retain water [224]. The results have suggested that substantial changes in meat tenderness can be achieved by altering the pH of the meat. A decrease in shear force can be achieved after 24 hours of pork and beef meat marination with LAB-based marinades: the increase of meat tenderness was found on average by 6.4% and 18.9% (P. acidilactici marinade), 4.6% and 27.7% (P. pentosaceus marinade), and by 2.2% and 15.9% (L. sakei marinade). Marination decreased shear force in all chicken meat samples and ranged from 8.1% (in marinated organic produced drumstic) to 26.1% (in marinated conventionally produced drumstic). It is clear that the lipid content is not the single source of variation in determining the pork sensory quality. Lonergan et al. [230] have reported that a high pH results in a greater sensory tenderness and juiciness scores and lower the star probe values and sensory chewiness scores. Lonergan et al. [230] suggest that a high pH product (above pH 5.80) can be expected to be superior to a lower pH product with regard to sensory quality, texture,

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and cook loss. In general, at a high pH, addition of lipid does not improve sensory tenderness, sensory chewiness, sensory juiciness, or star probe values. Importantly, there is a significant correlation between lipid content and sensory quality. Besides, IF contents were found lower in pork and beef meat samples after marination (except beef shoulder and beef M. longissimus dorsi samples). The highest IF loss was found in a pork shoulder sample (35.3%), and the lowest effect on IF in pork neck samples (3.1%) was fixed as compared to controls.

Also, colour and pH are important indicators of the overall meat quality and must be monitored during the marination process [228]. This study showed that marination increased the (L*) values of all meat samples: pork from 3.5% (ham muscle) to 21.4% (loin), beef from 0.5% (M. longissimus dorsi) to 40.3% (ham), and chicken from 1.2% (conventionally produced drumstick) to 5.0% (conventionally produced breast) as compared to controls. The marination of pork meat samples increased the (a*) values from 3.2% to 33.8% (pediococci) and from 1.4% to 24.0% (L. sakei) as compared to controls and decreased the (b*) values by 4.4% (M. longissimus dorsi), 7.2% (neck), 13.02% (ham muscle), 15.2% (loin) and 24.1% (shoulder) as compared with controls.

Different tendencies were estimated for beef meat samples: marination decreased the (a*) values on average by 16.6% (P. acidilactici), 33.34% (P. pentosaceus) and 24.3% (L. sakei),and increased the yellowness (b*) of beef samples on average by 96.6% (P. acidilactici), 56.6% (P. pentosaceus), and 69.4% (L. sakei) as compared with control samples. Marination of organic and conventionally produced chicken meat decreased the (a*) values on average by 3.3% and 8.9%, and the (b*) values on average by 6.2% and 12.1% (except thighs samples), respectively, compared with the control samples.

The colour compounds of processed meat can change during processing due to the presence of microorganisms and the inactivation of enzymes, which can result in undesired chemical reactions in the food matrix [25]. As the pH increases and moves further from the isoelectric point, meat proteins bind more water on their negatively charged side chains. Because of binding more water, there is less water to reflect light, and the meat appears darker in colour. It has been shown that as the pH decreases, myoglobin is more easily oxidized to metmyoglobin, and the colour of the meat is lighter and lower in colour intensity [106]. Nevertheless, meat acidification by LAB has positive technological aspects including the activation of muscle proteases and increased reddening through the formation of nitric oxide and nitrosylmyoglobin [142].

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Marination in weakly acidic media is one of the best ways to give meat some more flavor and tenderize it at the same time. On the other hand, during marination, free polyunsaturated fatty acids are to some extent oxidized, first to the primary oxidation products oxylipins and further to the secondary oxidation products that all may have detrimental effects on both the organoleptic properties and safety of the meat; besides, marination can increase the content of BAs in the processed food [207].

HIS, TYR, PUT, CAD and PHE are reported as the most important BAs in foods [126, 207], which may exert either psychoactive or vasoactive effects on sensitive humans: synaptic transmission, blood pressure control, allergic response and cellular growth control. Marination with marinades based on selected LAB was found to increase the PHE content from 15.4% (in beef shoulder treated with L. sakei) to 425.1% (in M. longissimus dorsi treated with P. pentosaceus). The high doses of PHE could induce behavioral responses in rodents, similar to those observed following amphetamine administration, and it also shows the ability to induce neurodegeneration in animals [170]. The PUT (12.27–131.29 mg/kg) and CAD (13.15–65.71 mg/kg) were dominant BAs in all pork and beef meat samples, except a pork ham muscle marinated with P. acidilactici. The PUT and CAD production is frequently found in Enterobacteria, and the TYR production is reported in the majority of Enterococci [67].

The highest concentrations of HIS were found in the pork ham muscle treated with P. acidilactici (15.09 mg/kg) and with P. pentosaceus (15.47 mg/kg) marinades, and the lowest – in pork neck (1.37 mg/kg) marinated with the P. pentosaceus marinade. HIS in the beef ham muscle treated with P. acidilactici was not detected. In many cases, marination increased the PHE content in chicken meat (up to 7.96–30.55 mg/kg), except in conventionally produced marinated breast and drumstick samples, and decreased the PUT content in all conventionally produced chicken meat samples by 34.9%, 74.1% and 74.9%. In organic produced chicken breast samples no HIS was found, and the highest content of HIS was 44.85 mg/kg (in conventionally produced marinated chicken drumsticks).

According to [35], slight responses are possible at concentrations in the food of 10–40 mg HIS, 5–10 mg TYR, 25 mg TRP, or 5 mg PHE, while for a toxic response 80–100 mg HIS or 25–250 mg TYR are necessary. Therefore, BAs accumulation in foods is a complex process affected by multiple factors and their interactions, the combinations of which are variable and product-specific [35], and the control of the end products is needed.

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4.4. New by developed plant bioproducts for improving ready-to-cook minced pork, pork and beef loin quality and safety

parameters

One of the modern approaches to improve the safety and functional value of food products is the application of the combined bioproducts consisting of savory plants having antimicrobial properties and bacteriocins producing LAB. Aromatic herbs can enhance the shelf-life of food because of their antimicrobial nature: they are primarily added to change or improve the taste [235]. In our study, 3% (from meat mass, m/m) of Sm and Sh bioproducts for improving RCMP quality were selected. The high content of savory plants is unacceptable for consumers, probably because of a strong, warm, spicy, herbaceous taste and flavor [135]. The use of savory plant bioproducts is a suitable solution for the improving RCMP microbial safety parameters. All of the RCMP samples produced with 3% of fermented Sm had satisfactory sanitary conditions. The average count of microorganisms observed in all samples increased during 5 days of storage under refrigeration; however, the tolerant coliforms and fungi were below the limits prescribed by the Commission Regulation (EC) No 852/2004 of the European Parliament. The inhibition effect on the growth of undesirable microorganisms was significantly (p < 0.05) influenced by the Sm fermentation technology (SMF or SSF) and the type of LAB used for fermentation. Microbial growth and metabolism contribute to the limitation of the shelf-life of meat products. SSF allowed the reduction of mesophilic bacteria growth during 120 h of the RCMP storage (up to 33.0–34.0% compared to control samples), while RCMP samples containing SMF Sm had a lower impact (the reduction of mesophilic bacteria growth during 120 h of storage was 13.2–17.4%). The highest antimicrobial activity against pathogens was shown by SSF with P. acidilactici Sm bioproducts. The results show that the antimicrobial activity is due to the LAB and the Satureja montana active compounds as well as their synergetic activity. Also, the antimicrobial activity of fermented Sm against pathogens could be due to the lactic acid [21] and antimicrobial compounds such as BLIS produced by the tested LAB [59]. Additionally, the Satureja species contains compounds with antimicrobial properties [333], and such a complex bioproduct (Sm and Sh, and LAB) could be promising for the preservation of meat products.

The microbial ecology of meat fermentation is a complex process in which LAB play a major role [98]. The processing of meat causes a number of physical, biochemical and microbiological changes which eventually result in the functional characteristics of the end product [202]. Lipid

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oxidation products, free fatty acids play a preponderant role in the sensorial properties of meat products as they act as VC precursors and also a as solvent of aroma compounds [192]. On the other hand, protein degradation leads to the formation of aromatic compounds [297]. According to our results, the Sm and Sh fermentation technology has a significant (p < 0.05) influence on the RCMP, pork and beef loin quality parameters. The pH of all meat products was considered highly acceptable regardless of treatment and ranged from 6.02 to 6.16, from 6.01 to 6.05, from 5.86 to 6.59, from 6.10 to 6.56 in the RCMP, pork and beef loin produced with Sm and Sh bioproducts, respectively. Numerous studies have shown the pH to be an indicator of meat quality. A low pH (< 5.4) corresponds to a propensity toward pale, soft, and exudative pork. The pH values 6.01–6.56 in this study indicate the acceptable quality of the tested meat products. The RCMP produced with SMF and SSF Sm and Sh had the dual benefit of increasing WHC (on average by 7.2% and 16.2%, by 3.1% and 12.3%, respectively) and tenderness (on average by 37.3% and 41.3% and by 36.5% and 36.3%, respectively) compared to control samples. RCMP produced with SSF with P. acidilactici Sm have the highest WHC (72.44 ± 0.79). In pork and beef loin samples marinated with SMF and SSF Sm and Sh bioproducts, similar tendencies were estimated (except pork samples produced with SMF with L. sakei Sm ): bioproducts increased the WHC of meat on average by 1.4%, 1.6% and 1.3%, by 0.6% and 1.8%, and by 2.8% and 2.2%, respectively. SSF Sm and Sh lowered the drip loss of minced meat products, marinated pork and beef loin by 38.8%, 8.3%, 0.4%, 0.4%, 3.8% and 6.3%, respectively as compared to control samples. No significant impact of fermented Sm and Sh was found on RCMP cooking loss (exept RCMP produced with SSF with P. pentosaceus and P. acidilactici Sh). The SMF Sm increased the cooking loss of RCMP on average by 16.1%, and the SMF Sh decreased the cooking loss on average by 11.0% as compared with control samples. A decrease of cooking loss was estimated in pork and beef loin marinated with SSF and SMF Sm and Sh bioproducts (except pork samples produced with SMF and SSF with L. sakei, P. pentosaceus and P. acidilactici Sm). The SMF and SSF Sh decreased the cooking loss of marinated beef loin on average by 14.3% and 5.3%, the SMF and SSF Sm and Sh decreased the cooking loss of marinated beef loin on average by 7.9%, 13.34%, 9.1% and 10.5%, respectively, as compared with control samples. It is well known that myofibrillar proteins are mainly responsible for the WHC and textural properties of processed meat products. Plant ingredients rich in dietary fibres have been used in cooked meat products to improve their textural properties and impart health benefits [212]. Ke [189] indicated that the use of lactic acid fermentation for the production of meat

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products inhibits the lipid oxidation and also could increase their WHC and tenderness. In addition, the muscle membrane protection from lipid oxidation by applying lipid-soluble antioxidants of the plant origin can maintain the membrane integrity of muscle fibres and reduce moisture loss [254]. However, because high amounts of plant dietary fibre could increase the hardness of cooked meat products, their application should be kept at low levels (< 2%).

Also, savory plant bioproducts influence the meat colour parameters. In fresh meat and meat products, colour is a strong indicator of quality. Myoglobin is the principal protein responsible for meat colour, although other heme proteins such as hemoglobin and cytochrome C may also play a role in beef, pork and poultry colour [156, 236]. Discolouration results from the oxidation of both ferrous myoglobin derivatives to ferric iron. The metmyoglobin formation depends on numerous factors including oxygen partial pressure, temperature, pH and, in some cases, microbial growth [32]. Lipid oxidation products induce the redox instability of muscle oxymyoglobin [11]. The authors have suggested that this covalent modification of myoglobin plays a role in the interrelationship between pigment and lipid oxidation. The rate of lipid oxidation in meat products can be effectively retarded by the use of antioxidants. Although chemical additives have been widely used in the meat industry to inhibit the development of lipid oxidation, their use is tending to decrease because of the growing concern among consumers about such chemical additives. This concern has led to an increased interest in the use of natural antioxidants. For example, the natural products isolated from spices or aromatic herbs can act as antioxidants and stabilize meat colour, thus extending the shelf-life of meat and meat products [181].

Despite the positive effects of fermented savory plant additives on meat technological properties, they can promote the formation of BAs in meat. The degradation of proteins during the fermentation process is one of the key factors involved in the improvement of the functional value of meat products. Furthermore, BAs can occur in fermented meat products at high concentrations since their accumulation is mainly related to the action of decarboxylase-positive bacteria and meat enzymes during fermentation and ripening [76]. The BAs determination is important not only because of their toxicity, but also for their potential use as freshness indicators [24]. PUT, HIS, SPR and SPD were the main BAs in control meat samples, and CAD, TYR were the most abundant in RCMP produced with fermented savory plants. In most of the cases, Sm bioproducts reduced the PUT content from 45.3% to 79.8% (in the RCMP-produced SSF with P. acidilactici Sm and in the RCMP-produced SMF with L. sakei Sm, respectively), HIS from 50.3%

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to 96.2% (in the RCMP-produced SMF with L. sakei Sm, and in RCMP-produced SSF with P. acidilactici Sm, respectively), SPR by 100% in most of the samples, and SPD from 39.5% to 100% (in RCMP-produced with SSF with P. acidilactici Sm and in RCMP-produced with SSF and SMF with P. pentosaceus and L. sakei SMF Sm, respectively). In contrast, the concentrations of PHE, CAD, TYR and TRP in most of the RCMP samples were increased. Different tendencies in RCMP samples produced with Sh bioproducts were estimated: the increased PUT, CAD and HIS content on average by 146.6%, 4617.6%, 412.0%, respectively, and the decreased PHE, TYR and TRP content as compared to control samples.

The TYR, CAD, PUT and HIS were reported as the most prevalent BAs in meat and meat products [327], and only SPD and SPR are present at significant levels in fresh meat [143]. The concentrations of TYR, PUT and CAD normally increase during the processing and storage of meat and meat products, whereas SPD and SPR decrease or remain constant [327]. Therefore, their amounts and ratios have been proposed as an index of the hygienic conditions of raw material and/or manufacturing practices since their amount increases during microbial fermentation or spoilage [214]. The HIS intake higher than 100 mg/kg may cause a slight or intermediate poisoning [290]. Less is known about the toxic doses of other amines. The recommended maximum level of TYR has been proposed variously to be in the range of 100–800 mg/kg of food. The value of 30 mg/kg for β-phenylethylamine has been reported as a toxic dose in food [120]. The BAs contents in RCMP, pork and beef loin treated with bioproducts were found far below those levels associated with a health risk.

4.5. Pea fiber incorporation in the formula of gluten-free meat products

The growing demand of gluten-free (GF) products constitutes areas of

increasing interest to meet cereal-based requirements of celiac (CD) and wheat-intolerant patients [253, 256, 404]. The removal of gluten from food products traditionally based on wheat has a significant impact on their structure and texture. It is not an easy task to adjust a recipe for GF products, which would give a product with sensory attributes, nutritional value and consumer acceptance comparable to traditional food [404]. Because derivates of glutenrich-grains are important sources of nutrients in the general diet, their exclusion from the diet of celiac patients could potentially have a major effect on their nutritional status if such foods are not replaced with balanced alternatives [348]. A proper replacement of gluten-forming cereals by non-gluten-forming systems in food products is still a major challenge, particularly in achieving sensory and nutritionally

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balanced foods, despite the accumulating knowledge of the physical, chemical and technological principles of matrices [341]. To produce GF foods containing rice or corn starches and legume flours, DF is used in order to increase the food nutritional value [252]. DF can effectively be incorporated in the processed meat products as binders, extenders, and fillers, they can significantly replace the unhealthy fat components from the products, increase their acceptability by improving nutritional components, pH, WHC, emulsion stability, shear press value, the sensory characters of finished products [372].

Meat industry has been searching for food additives and ingredients for their products that provide nutritional benefits, such as the use of fibres, minerals or bioactive compounds [342]. The inclusion of DF as ingredients in meat products has been considered of interest for the partial replacement of meat due to their inherent functional and nutritional effects [151, 204]. The consumption of meat products fortified with DF can lead to the prevention of diseases such as the coronary heart disease, diabetes, irritable bowel disease, obesity [372].

It was found that the moisture content and drip loss of RCMP produced with the pea fiber was 1.1 and 1 times higher, respectively, as compared with RCMP produced with semolina. The addition of fibers into meat products usually increases the WHC of the food matrix and consequently the moisture content [95, 301].

RCMP produced with pea fiber had a higher WHC (by 9.7%) as compared with RCMP produced with the semolina. It was found that semolina increased the cooking loss by 3%, as compared with the RCMP produced with the pea fiber. The use of pea fibers increased the cooking yield and minimized the production cost without lowering the sensory properties [34]. A slight increase in the shear force was found in the RCMP produced with semolina (by 40.5%). It is well known that the ultimate pH of the muscle is an important contributing factor to meat quality, expressed as tenderness, colour and storage life [388].

In our study, the RCMP produced with semolina had a higher (L*) value (by 7.2%), (a*) value (by 5.1%) and (b*) value (by 0.5%). Semolina has a light colour, neutral taste/odor [37], and pea fiber can be white to green depending on pea source and the purification that has been done [267]. Colour is one of the most important characteristics of meat substitutes. The green colour of peas can be a barrier to their use in meat products [396].

The content and composition of BAs is associated with the degree of food fermentation or degradation [103]. It was found that in both RCMP (with pea fiber and semolina) the main BA was SPR. No PHE and TRP in

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RCMP samples were found. The addition of semolina reduced the total BAs content by 19.3% as compared with the RCMP produced with pea fiber.

4.6. Lactic acid bacteria for decreasing the polycyclic aromatic

hydrocarbons content in cold smoked pork sausages

According to the EFSA, meat and meat products are one of the food categories contributing most to the dietary PAHs intake per day of the European Union member state consumers [91, 97]. This demonstrates the important role of PAHs studies for smoked food products, with a view to quantifying these compounds and identifying the factors that increase PAHs in foods. Recently, great interest in the biodegradation of chemical compounds using microorganisms has been expressed. Researchers have reported the positive effect of LAB against PAHs and heterocyclic aromatic amines [2, 3, 4]. The PAHs content obtained in sausages even in not pre-treated with LAB suspension samples, was below the maximum value currently allowed by the European Union regulations. According to Ledesma et al. [219], the maximum BaP content found in various studies mainly falls below the maximum value currently allowed by the European Union regulations (2.0 mg/kg); however, a high concentration of BaP is still found in smoked meat products around the world. Therefore, it is still important to control and study their manufacturing conditions in order to minimize the contamination of smoked meat products with PAHs. The application of LAB for sausage treatment before and after smoking has a significant influence on BaP and Chr decreasing. In our study, the pH values of LAB used for sausage treatment were in the range of 4.21–4.42. Zhao with co-authors [411] reported that the maximum values of the BaP binding rate of several LAB were obtained at pH 4.0 and 5.0. The results obtained in our study indicate that during the direct smoking process the greatest amount of PAHs is formed in the outer layers of sausages, as compared with the inner layers. Similar results were reported by [217, 218], as they found that the greatest amount of BaP, the PAHs content indicator, was deposited in the casing of the meat product and not inside the product. Andrée et al. [15] and Santos et al. [339] report that PAHs accumulate on the surface of the smoked meat products during smoking and then migrate into the products being smoked. [2] reported that Bifidobacterium bifidium, Streptococcus thermophilus and Lactobacillus bulgaricus reduced the PAHs content respectively by 46.6%, 87.7% and 91.5%. Zhao et al. [411] reported that several LAB strains together might be beneficial for removing several toxic compounds. Therefore, the mechanism of reducing toxic compounds is still unclear. Some researchers suggest that toxins are converted by specific

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enzymes produced by cells [112], therefore, the PAHs content decrease after sausage treatment with the LAB suspension could be achieved. Other reports revealed that this process was due to the binding of the carcinogen to cell wall components [140].

The use of bacteriocins producing LAB with a wide range of antimicrobial activity can reduce the PAH content in cold smoked pork sausages, increase the shelf life of the products and inhibit the growth of certain pathogenic bacteria [201]. L. sakei demonstrated the highest antimicrobial activity against Y. pseudotuberculosis (the diameter of the inhibition zone was 19 ± 0.5 mm). A very strong inhibition of LAB metabolites was observed against Pseudomonas aeruginosa (the diameter of the inhibition zone varied from 11.0 ± 0.5 to 15.0 ± 0.3 mm). Other researchers reported about the antimicrobial activity of LAB-produced metabolites as well [9, 59, 325]. Organic acids, hydrogen peroxide and bacteriocins produced during fermentation decreased the pH and inhibited the growth of spoilage microorganisms; therefore, LAB species can contribute to the control of the indigenous microbial reproduction during technological processes [9]. [288] have reported that some strains of P. pentosaceus produce antimicrobial peptides identified as a pediocin or a class II bacteriocin which, are heat- and cold-stable peptides with the inhibitory activity against several Gram-positive pathogens. In our study used LAB strains demonstrated good inhibition properties against all tested undesirable microorganisms and could be recommended for the surface treatment of cold-smoked pork sausages, in order to reduce chemical and biological contamination.

However, relatively high levels of BAs can be found in fermented sausages [231]. Food spoilage bacteria such as Enterobacteria contribute to the production of decarboxylases and BAs formation. The diamines PUT and CAD can react with nitrite to form carcinogenic nitrosamines [146]. Polyamines and diamines may be converted into stable carcinogenic N-nitroso compounds and enhance the growth of chemically induced aberrant crypt foci in the intestine [94].

The main BAs in the outer layers and centre of the non-treated sausages were PUT (17.29 ± 0.17 and 16.14 ± 0.12 mg/kg, respectively), TYR (12.91 ± 0.14 and 11.12± 0.07 mg/kg, respectively) and CAD (9.04 ± 0.11 and 7.31 ± 0.14 mg/kg, respectively). A positive effect of the LAB treatment was observed on the content of CAD and SPD. A significant reduction of CAD after treatment with all tested LAB before smoking was observed. In control samples, CAD ranged from 7.31 mg/kg to 9.04 mg/kg, whereas after treatment with L. sakei before smoking the content of CAD in the centre of sausages decreased to 1.98 mg/kg; moreover, in the outer layers of sausages

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no content of CAD was detected. After treatment with P. acidilactici before smoking, the content of CAD in the outer layers decreased to 2.38 mg/kg; moreover, in the centre of sausages the CAD was not detected. P. acidilactici treatment before smoking decreased the content of CAD to 69%. A positive effect of LAB treatment before smoking on the content of SPD was found. Other tendencies were noted when sausages were treated with LAB after smoking. In this case, the content of PUT was significantly decreased (approx. by 53% when L. sakei and P. acidilactici were applied) or totally eliminated (using P. pentosaceus) from the outer layers and the centre of sausages. However, the total content of BAs in control samples was found to be lower, also in samples treated with water (after and before smoking) as compared with samples treated with LAB (from 8.2% to 97.7%, and from 4.1% to 99.7%, respectively). Some species of LAB, however, can produce BAs, whereas [366] reported that Pediococci did not possess amino acid decarboxylase activity. A positive effect on decreasing the content of BAs was shown by P. acidilactici KTU05-7, P. pentosaceus KTU05-9, L. sakei KTU05-6 strains, when these LAB were applied for RCMP production [365]. According to literature data, the BAs production is dependent on the microflora, the availability of precursors and physicochemical factors such as temperature, pH, salt, oxygen and sugar concentration [406]. Furthermore, the bacterial growth also increases the amount of BAs by raising the production of decarboxylase [215].

4.7. Savory plant bioproducts for higher sustainability unrippened curd

cheese production UCC (acid curd cheese, tvorog, cottage cheese) is a traditional Russian

cottage cheese which is also consumed in the neighbouring countries including Lithuania. Short shelf-life is considered the greatest fault of fresh curd cheeses [168]. LAB with antimicrobial properties may be used in cheese production as natural preservatives to extend the shelf-life of the product [64]. Natural food additives deriving from herbs and spices have been recognized and used in food preservation for centuries [291]. Because of the presence of a high content of thymol and carvacrol in the Satureja species, its attractive aroma and simple cultivation these plants are used as a flavouring substance in many foods [52]. The formation of VC in UCC depended on the savory plant bioproducts used for UCC production. In the final products with fermented plants, some VC were reduced, eliminated or their concentration was increased. Similar results as to the changes of VC from medical plants (S. montana L. seeds) after fermentation using LAB were reported by [176].

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Fermented dairy products have a beneficial dietary effect on human health, however, they might be a source of higher alimentary intake of BAs in the human diet [208]. The main source of decarboxylase-positive bacteria in cheese is raw milk. In fermented foods, including cheeses, the production of BAs increases as a result of proteolysis [337]. Among the most frequently consumed food products, cheese is most closely associated with BAs after fish [319]. Several factors influence the BAs content in cheese: production conditions, the time of ripening and storage, milk pasteurization, and the starter culture [363]. BAs production by LAB may be controlled at various levels during fermentation [227]. Therefore, the selection of LAB able to decrease the content of BAs is of outstanding importance. The main BA in UCC produced with savory plant bioproducts was CAD. The concentration of CAD in UCC varied from 3.55 mg/kg to 137.6 mg/kg, and depended on the LAB strain used for savory fermentation and its fermentation technology (SSF or SMF). The BAs content in UCC produced with Rc – L. sakei bioproducts and in SSF conditions, was 28 times lower than the BAs content in UCC produced with nonfermented Rc. The content of BAs in milk ranges around 1 mg/L. After the fermentation process, more than 1000 mg of BAs can be found in 1 kg of cheese, probably as a result of a longer ripening period and its proteolysis [226, 227]. Enterococci have been traditionally regarded as the main TYR formers and heterofermentative lactobacilli have been considered the main HIS producers, but other genera of bacteria are also capable of forming BAs in cheese [303]. It has been shown that bacteriocinogenic strains of LAB are capable of inhibiting decarboxylase-positive bacteria, thus hindering BAs formation [46]. The results of this study are in agreement with [46] statement that a decrease of BAs in the production of UCC could be achieved by applying bacteriocins producing LAB with antimicrobial activity against Enterobacteria able to decarboxylate free amino acids. Some strains of Lactobacillus, Pediococcus and Micrococcus have the ability to degrade BAs, such as TYR and HIS, by means of monoamine oxidases, preferably in aerobic conditions [223].

The fermentation of Sm and Rc with selected LAB significantly reduced the amount of Enterobacteria, yeast, fungi and spores of mesophilic bacteria in the samples and increased the amount of VC, such as thymol, carvacrol and p-cymene, in UCC. The bioproducts derived from a combination of LAB and savory plants increased the overall acceptability and shelf life of curd cheese and, therefore, can be recommended for the higher quality and longer shelf life unrippened curd cheese production.

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CONCLUSIONS

1. Fermentation with Lactobacillus sakei KTU05-6, Pediococcus acidilactici KTU05-7, Pediococcus pentosaceus KTU05-8 is an appropriate technology for Sm, Sh, Rc, defatted flaxseed, Jerusalem artichoke, pea fiber and defatted soy flour biotreatment; however, the fermentation conditions (SSF or SMF) should be selected in accordance with the plant type and the desired properties of the plant bioproduct:

1.1. In most of the cases, a higher TTA and a lower pH could be reached after the SMF of plants, and the interaction of the analysed factors (fermentation method, LAB used for the fermentation, and the type of plants) is significant for the TTA and pH of plant bioproducts (p ≤ 0.05).

1.2. The LAB count in plant bioproducts is significantly influenced by the fermentation method (p ≤ 0.0001), and the highest content of valuable LAB by using SSF conditions could be achieved.

1.3. The activity of amylolytic and proteolytic enzymes is significantly influenced by the fermentation method (p ≤ 0.0001), by the type of LAB used for the fermentation (p ≤ 0.0001), and by the type of plant (p ≤ 0.0001).

2. Fermentation has an influence on the content of bioactive compounds

(ARs, lignans and VC), TPC and RSA of plant bioproducts: 2.1. Fermentation increases the MAT and SECO content, and a

higher content of lignans by using the SSF of plants could be reached.

2.2. Fermentation decreased the ARs content in all samples, and its content was significantly influenced by the type of LAB applied for fermentation: C15:0 (p ≤ 0.004), C19:0 (p ≤ 0.001), C21:0 (p ≤ 0.0001), and C23 (p ≤ 0.001).

2.3. Fermentation increased the TPC content and RSA in plant bioproducts, and TPC was significantly influenced by the fermentation method (p ≤ 0.037) and the type of LAB used for the fermentation (p ≤ 0.001), and RSA – by the type of LAB (p ≤ 0.05).

3. SSF and SMF fermentation with selected LAB has a different influence

on plant bioproduct safety parameters; however, fermentation decreased the count of undesirable microorganisms in plants, and the D(-)-lactic

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acid and BAs content in plant bioproducts was far below those levels associated with a health risk:

3.1. The interaction of factors (fermentation method, LAB used for the fermentation, and the type of plant) has a significant influence on the content of L(+) and D(-) isomers in plant bioproducts (p ≤ 0.0001), with L(+)-lactic acid as the predominant isomer.

3.2. The total BAs content in fermented plants was significantly influenced by the type of LAB (p ≤ 0.05) and by the type of plant product (p ≤ 0.05) and a moderate positive significant correlation between the total BAs content and the moisture of the substrate (r = 0.543; p = 0.044) was found.

3.3. The count of Enterobacteria, yeasts and molds in plant bioproducts was significantly influenced by the fermentation method (p ≤ 0.001 and p ≤ 0.002, respectively) and by the LAB used for the fermentation (p ≤ 0.0001), and in all bioproducts Enterobacteria, yeast and mold counts were lower, as compared with nonfermented samples.

4. Plant bioproducts have a higher antimicrobial activity than its

individual components, and the antimicrobial activity of savory plant extracts and their bioproducts was significantly influenced by the type of LAB (against E.coli p ≤ 0.010, B. subtilis p ≤ 0.0001, P. fluorescens biovar. III p ≤ 0.023, and P. fluorescens biovar. V p ≤ 0.001) and by the type of a savory plant (against E.coli p ≤ 0.002 and against B. subtilis, P. fluorescens biovar. III and P. fluorescens biovar. V p ≤ 0.0001).

5. The application of selected new created plant bioproducts to the food of

animal origin formula allows to produce a higher value safer food: 5.1. The Sm and Sh bioproducts are safe (BAs content in RCMP

produced with plant bioproducts is far below those levels associated with a health risk) and the improve stability, enrich the flavour (with a higher content of ρ-cimene, γ-terpinene and carvacrol in RCMP), prevent meat decolouration, increase the content of phenolic compounds, thus increasing the acceptability and shelf-life of RCMP.

5.2. Marination with the lacto-fermented marinade based on potato juice with both pediococci caused a higher WHC and tenderness of pork meat, significantly increased its redness and overall acceptability and could be recommended for pork meat

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treatment to improve its tenderness and colour, thus increasing the overall acceptability.

5.3. Marination with all used lacto-fermented marinades increased the WHC of the ham muscle, M. longissimus dorsi, and loin. The marinade based on L. sakei caused a higher tenderness, and beef meat treated with P. acidilactici was indicated as having the highest acceptability.

5.4. The marinade based on P. acidilactici increased the WHC and tenderness of organic and conventionally produced chicken meat, and the BAs content was far below the recommended limits.

5.5. The LAB marinade based on potato juice could be applied for the cold smoked pork sausages surface treatment in order to reduce microbial contamination, BAs content (decrease CAD, SPD, and PUT) and PAHs formation (reduces the BaP and Chr content).

5.6. Sm and Sh bioproducts increase the VC (thymol, carvacrol and p-cymene) content in UCC, its overall acceptability and shelf life.

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PUBLICATIONS

Publications based on the results of the dissertation were published in journals with an impact factor refereed by the „Thomson Reuters

Web of Knowledge“

1. Bartkiene E, Mozuriene E, Juodeikiene G, Vidmantiene D, Cizeikiene D, Maruska A, Stankevicius M, Ragazinskiene O. Pork meat products functional value and safety parameters improving by using lactic acid fermentation of savory plants. Journal of Food Science and Technology 2015;52(11):7143-52.

2. Mozuriene E, Bartkiene E, Juodeikiene G, Zadeike D, Basinskiene L, Maruska A, Stankevicius M, Ragazinskiene O, Damasius J, Cizeikiene D. The effect of savoury plants, fermented with lactic acid bacteria, on the microbiological contamination, quality, and acceptability of unripened curd cheese. Journal of Food Science and Technology 2016; 69:161-8.

3. Mozuriene E, Bartkiene E, Krungleviciute V, Zadeike D, Juodeikiene G, Damasius J, Baltusnikiene A. Effect of natural marinade based on lactic acid bacteria on pork meat quality parameters and biogenic amine contents. LWT-Food Science and Technology 2016; 69:319-26.

4. Bartkiene E, Bartkevics V, Mozuriene E, Krungleviciute V, Novoslavskij A, Santini A, Rozentale I, Juodeikiene G, Cizeikiene D. The impact of lactic acid bacteria with antimicrobial properties on biodegradation of polycyclic aromatic hydrocarbons and biogenic amines in cold smoked pork sausages. Food Control 2017; 71:285-92.

Publications based on the results of the dissertation were published

in peer – reviewed journals refereed by other data bases 1. Bartkiene E, Skabeikyte E, Krungleviciute V, Jakobsone I, Bobere N,

Bartkevics V, Juodeikiene G. The influence of fermentation with BLIS producing lactic acid bacteria on the content of alkylresorcinols and lignans in plant products. The Open Biotechnology Journal 2015;9:32-8.

2. Mozūrienė E, Bartkienė E, Krunglevičiūtė V, Juodeikienė G. Skirtingų jautienos skerdenos dalių, fermentuotų pieno rūgšties bakterijomis, kokybės rodikliai. Maisto chemija ir technologija 2014;48(2):64-73.

3. Mozūrienė E, Bartkienė E, Juodeikienė G, Ragažinskienė O, Maruška A. Jautienos nugarinės, apdorotos pieno rūgšties bakterijomis fermentuotu vaistiniu - prieskoniniu augalu, kokybės rodikliai. Maisto chemija ir technologija 2015;49(1):36-45.

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4. Oniščiukas M, Bartkienė E, Skabeikytė E, Krunglevičiūtė V, Juodeikienė G. Pienarūgštės fermentacijos įtaka ekologiškai ir tradiciškai pagamintų vištienos skerdenėlių kokybės rodikliams. Maisto chemija ir technologija 2014;48(2):74-83.

5. Bartkienė E, Juodeikienė G, Zaborskienė G, Krunglevičiūtė V, Rekštytė T, Skabeikytė E. Biogeninių aminų susidarymas pašarams naudojamuose fermentuotuose augaliniuose produktuose. Veterinarija ir zootechnika 2013;63(85):3-11.

6. Bartkiene E, Skabeikyte E, Juodeikiene G, Vidmantiene D, Basinskiene L, Maruska A, Ragazinskiene O, Krungleviciute V. The use of solid state fermentation for food and feed plant material processing. Veterinarija ir zootechnika 2014;66(88):3-11.

Abstracts of international conferences based on the results of the

dissertation 1. Mozuriene E, Bartkiene E, Krungleviciute V, Juodeikiene G,

Vidmantiene D, Baltusnikiene A. The influence of natural marinade based on bacteriocins producing lactic acid bacteria on pork meat quality parameters and biogenic amines content. 4th International Conference for PhD Students Multidirectional Research in Agriculture and Forestry. Poland, Krakow. 2015. March 22-23. P. 96.

2. Mozuriene E, Bartkiene E, Juodeikiene G, Zadeike D, Maruska A, Ragazinskieke O. The influence of fermented with certain lactic acid bacteria Satureja hortensis on the quality and technological parameters of pork and beef loin. The 9th Vital Nature Sign 2015. Lithuania, Kaunas. 2015. May 14-16. P. 66.

3. Mozuriene E, Bartkiene E, Juodeikiene G, Maruska A, Ragazinskieke O. The Influence of fermented with lactobacilli Satureja montana on the pork loin quality and safety parameters. 10th Baltic Conference on Food Science and Technology „Future Food: Innovations, Science and Technology“ – FoodBalt – 2015. Lithuania, Kaunas. 2015. May 21-22. P. 28, OP 14.

4. Krungleviciute V, Bartkiene E, Kantautaite J, Skabeikyte E, Jakobsone I, Gailane N, Bartkevics V, Juodeikiene G. Changes of biologically active dietary fibre compounds during barley bran, pea fiber and lupine seeds fermentation. 6 th International Dietary Fibre Conference. France, Paris. 2015. June 1-3 P. 157-158. 3a.14.

5. Krungleviciute V, Bartkiene E, Kantautaite J, Skabeikyte E, Jakobsone I, Gailane N, Bartkevics V, Juodeikiene G. The influence of lactic acid fermentation on biologically actine compounds (lignans and alkylresorcinols) content in certain plant products. Nutrition, health and

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quality of food and animal origin challenges and opportunities. Lithuanian university of health sciences. Lithuania, Kaunas. 2014. P. 38-41.

6. Skabeikyte E, Lepeskaite E, Bartkiene E, Juodeikiene G, Vidmantiene D, Maruska A, Ragazinskiene O. The influence of solid state fermented savory plants of the genus Satureja (montana and hortensis) with certain lactobacilli on the quality and safety of chicken meat products. 9th Baltic Conference on Food Science and Technology „Food for consumer well-being“. FOODBALT – 2014, Latvija, Jelgava. 2014. May 8-9. P. 68.

7. Skabeikyte E, Jociene G, Bartkiene E, Juodeikiene G, Vidmantiene D, Maruska A, Ragazinskiene O. The influence of the fermented with certain Lactobacilli Rhaponticum carthamoides on the quality and safety of curd cheese. 8th International Scientific Conference THE VITAL NATURE SIGN VNS2014. Lithuania, Kaunas. 2014. May 15-17. P. 50.

8. Bartkienė E, Juodeikienė G, Zaborskienė G, Krunglevičiūtė V, Rekštytė T, Skabeikytė E. Biogenic amines formation in fermented plant products. Tarptautinė mokslinė konferencija „Žemės ūkio gyvulių fiziologija“, LSMU VA. Lietuva, Kaunas. 2012. Rugsėjo 27-28 d. 73 P.

9. Bartkiene E, Krungleviciute V, Juodeikiene G, Vidmantiene D, Skabeikyte E, Maruska A, Ragazinskiene O, Matusevicius P. The influence of spontaneous and lactic acid fermentation with certain lactobacillus on biogenic amines and D-(-) lactate formation in Jerusalem artichoke. 12. Tagung. Institut für Agrar- und Ernährungwissenschaften Professur für Tierernährung, Lutherstadt Wittenberg Martin-Liuter-Universität Halle-Wittenberg. Deutschland, Wittenberg. 2013. Nowember 12–14. P. 210-212.

10. Bartkiene E, Krungleviciute V, Juodeikiene G, Vidmantiene D, Skabeikyte E, Maruska A, Ragazinskiene O, Matusevicius P. The influence of solid state fermentation with certain lactobacillus on biogenic amines formation in flaxseed. 12. Tagung. Schweine- und Geflügernährung, Naturwissenschaftliche Fakultät III, Lutherstadt Wittenberg Martin-Liuter-Universität Halle-Wittenberg. Deutschland, Wittenberg. 2013. Nowember 12–14. P. 213-214.

Abstracts of national conferences based on the results of the

dissertation 1. Skabeikytė E, Bartkienė E, Juodeikienė G, Maruška A, Ragažinskienė

O. Pienarūgštėmis bakterijomis apdorotų vaistinių – prieskoninių augalų panaudojimo galimybės maltos mėsos pusgaminių vertei

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padidinti. VI nacionalinė doktorantų mokslinė konferencija „Mokslas – sveikatai“. Lietuvos sveikatos mokslų universitetas. Lietuva, Kaunas. 2013. Balandžio 5 d. P. 27-29.

2. Mozūrienė E, Bartkienė E, Jočienė G, Juodeikienė G, Bašinskienė L, Vidmantienė D, Maruška A, Stankevičius M, Ragažinskienė O. Pieno rūgšties bakterijomis fermentuotų Satureja montana L. ir Rhaponticum carthamoides produktų įtaka varškės sūrių saugos ir kokybės rodikliams. III jaunųjų mokslininkų konferencija „Jaunieji mokslininkai – žemės ūkio pažangai“. Lietuvos mokslų akademija, Žemės ūkio ir miškų mokslų skyrius. Lietuva, Vilnius. 2014. Lapkričio 5 d. P. 51.

3. Skabeikytė E, Bartkienė E. Biotechnologinės priemonės kiaulienos kokybės ir saugos rodiklių pagerinimui. VII nacionalinė doktorantų mokslinė konferencija „Mokslas – sveikatai“. Lietuvos sveikatos mokslų universitetas. Lietuva, Kaunas. 2014. Balandžio 9 d. P. 47-48.

4. Mozūrienė E, Bartkienė E, Juodeikienė G, Ragažinskienė O, Maruška A. Pieno rūgšties bakterijomis apdorotų Satureja montana augalinių priedų įtaka jautienos nugarinės kokybės rodikliams. VIII nacionalinė doktorantų mokslinė konferencija „Mokslas – sveikatai“. Lietuva, Kaunas, 2015. P. 121-122.

5. Skabeikytė E, Bartkienė E, Juodeikienė G, Vidmantienė D, Eidukonytė D, Bašinskienė L, Maruška A, Ragažinskienė O. Bakteriocinus produkuojančių pieno rūgšties bakterijų taikymas kietafazei augalinių produktų fermentacijai. Konferencija „Jaunieji mokslininkai – žemės ūkio pažangai“. Lietuvos mokslų akademija, Žemės ūkio ir miškų mokslų skyrius. Lietuva, Vilnius. 2012 m. Lapkričio 21 d. 53 P.

6. Skabeikytė E, Bartkienė E, Juodeikienė G, Vidmantienė D, Maruška A, Stankevičius M, Šiugždaitė J, Ragažinskienė O. Pieno rūgšties bakterijomis fermentuotų Satureja montana L. produktų įtaka maltos mėsos pusgaminių saugos ir kokybės rodikliams. Konferencija „Jaunieji mokslininkai – žemės ūkio pažangai“. Lietuvos mokslų akademija, Žemės ūkio ir miškų mokslų skyrius. Lietuva, Vilnius. 2013. 45-46 P.

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SUMMARY IN LITHUANIAN

SANTRUMPOS

AJ – Aromatiniai junginiai ARs – Alkilrezorcinoliai BA – Biogeniniai aminai BaA – Benz[a]antracenas BaP – Benzo[a]pirenas BbF – Benzo-[b]fluorantenas BFJ – Bendras fenolinių junginių kiekis BTR – Bendras titruojamasis rūgštingumas Chr – Chrisenas FEN – Feniletilaminas HIS – Histaminas INR – Ilgiausiasis nugaros raumuo KAD – Kadaverinas KF – Kietafazė fermentacija KTU – Kauno technologijos universitetas LRSGA – Laisvųjų radikalų surišimo gebos aktyvumas LSMU VA – Lietuvos sveikatos mokslų universitetas Veterinarijos akademija MAT – Matairezinolis PAA – Policikliniai aromatiniai angliavandeniliai PRB – Pieno rūgšties bakterijos PSO – Pasaulio Sveikatos Organizacija PUT – Putrescinas Rc – Rhaponticum carthamoides CD. SEKO – Sekoizolaricirezinolis Sh – Satureja hortensis L. Sm – Satureja montana L. SPD – Spermidinas SPR – Sperminas TF – Tradicinė fermentacija TIR – Tiraminas TR – Tarpraumeniniai riebalai VDU – Vytauto Didžiojo universitetas VN – Virimo nuostoliai VR – Vandens rišlumas VS – Varškės sūris

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ĮVADAS

Didėjant vartotojų poreikiui saugesniam ir sveikesniam maistui, atsiranda niša naujų, tvarių ir saugių maisto gamybos biotechnologijų kūrimui [38]. Kietafazė fermentacija (KF) sulaukė biotechnologijos pramonės dėmesio ir manoma, kad tai puiki alternatyva tradicinei fermentacijai (TF) skystoje fazėje. KF procesas yra efektyvesnis, nes mikroorganizmų dauginimasis bei metabolizmas vyksta labai arti fermentuojamojo substrato. Šiam procesui svarbu parinkti tinkamus mikroorganizmus, nes nuo jų priklauso ne tik metabolizmo produktų koncentracija substrate, bet ir galutinio produkto kokybė [319, 353].

Vis dažniau gamintojų ir vartotojų dėmesio sulaukia maisto tvarumo ir saugos užtikrinimas natūraliomis priemonėmis, pvz., bio-konservavimas bakteriocinus produkuojančiomis pieno rūgšties bakterijomis (PRB) [98] ir / ar antimikrobinėmis savybėmis pasižyminčiais augalais [45, 400]. Tačiau iki šiol nėra atlikta tyrimų ir nenustatyta, koks būtų šių priemonių simbiotinis poveikis maisto produktų tvarumui ir kokybės bei saugos rodikliams. Dėl to atsiranda poreikis PRB – augalų bioproduktų, skirtų maisto tvarumo, saugos ir kokybės užtikrinimui sukūrimui.

Maisto pramonėje naudojami natūralūs maisto priedai, gaunami iš savitų cheminių savybių turinčių augalų, pasižyminčių išskirtinėmis juslinėmis, antimikrobinėmis, antioksidacinėmis bei konservuojančiomis savybėmis [249, 291, 345]. Baltymingų ir didelį kiekį skaidulinių bei biologiškai aktyvių medžiagų turinčių augalų fermentacija PRB didina foliatų, tirpių skaidulinių medžiagų, fenolinių junginių kiekį, antioksidacinį aktyvumą, virškinamumą, lignanų ir alkilrezorcinolų kiekį (ARs) [50, 86, 242, 308, 378]. Parinktais mikroorganizmais fermentuoti savitos cheminės sudėties augalai galėtų būti puikus šaltinis gyvūninės kilmės maisto žaliavų ir / ar produktų vertės ir tvarumo padidinimui, bio-saugos užtikrinimui, savitų juslinių savybių suteikimui [383]. Mėsa ir mėsos produktai yra puikus baltymų, mineralinių medžiagų, vitaminų, mikro- ir makro-elementų šaltinis žmonių mityboje, bet ir puiki terpė nepageidaujamiems mikroorganizmams daugintis bei vystytis. Saugių technologijų kūrimas mėsos ir mėsos produktų tvarumo ir saugos užtikrinimui yra ypač aktualus [222].

Kita grupė gyvūninių maisto produktų, kuri užtikrina vartotojams pilnavertišką mitybą yra pieno produktai. Išskirtinio populiarumo susilaukia rauginti pieno produktai, dėl savo maistinės vertės, tvarumo, natūralumo bei teigiamo poveikio sveikatai [23]. Tiek mėsos, tiek pieno pramonėje, siekiant gauti išskirtinių savybių produktus taikomos fermentacijos technologijos, kuriose pagrindiniai mikroorganizmai yra PRB. Išskirtinėmis savybėmis pasižyminčios PRB gali padidinti maisto produktų tvarumą, slopindamos

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nepageidaujamų mikroorganizmų veiklą, išskirti metabolitus (bakteriocinai, pieno ir acto rūgštys, etanolis, acetaldehidai, aromatiniai junginiai ir kt.) užtikrinančius produktų saugą [134, 251], formuoti skonį, spalvą ir tekstūrą, suteikiant išskirtines juslines bei technologines savybes [38, 385].

PRB yra saugūs mikroorganizmai [407], tačiau fermentacijos metu, priklausomai nuo technologinėse schemose naudojamų mikroorganizmų bei substrato savybių, gali susidaryti nepageidaujami, vartotojų sveikatai žalingi junginiai – biogeniniai aminai (BA) [70]. Taip pat, reikėtų akcentuoti, kad pagrindinis PRB metabolitas (pieno rūgštis) gali būti L(+) ir D(-) izomerai. Įvertinant, kad pastarasis nėra metabolizuojamas žinduolių organizme ir gali sukelti acidozes, maisto pramonėje rekomenduojama parinkti ir naudoti mikroorganizmus, kurie produkuotų daugiau L(+) pieno rūgšties [393]. Modeliuojant naujus maisto produktų gamybos bio-technologijų prototipus, išskirtinis dėmesys turėtų būti skiriamas ne tik jų kokybei bei tvarumui, bet ir saugos užtikrinimui BA ir D(-) pieno rūgšties izomerų aspektu.

1. DISERTACINIO DARBO TIKSLAS IR UŽDAVINIAI

Darbo tikslas: taikant TF ir KF schemas, sukurti bakteriocinus produkuojančių PRB ir savita chemine sudėtimi ir savybėmis pasižyminčių augalų (turinčių didelį baltymų, fitoestrogenų, inulino, fenolinių junginių, eterinių aliejų bei skaidulinių medžiagų kiekį) bioproduktus bei pritaikyti juos vertingesnių, saugesnių ir tvaresnių gyvūninės kilmės maisto produktų gamybai.

Uždaviniai:

1. Sumodeliuoti augalinių substratų fermentacijos technologijų schemas bei įvertinti pagrindinius, proceso efektyvumą nusakančius, rodiklius.

2. Įvertinti biologiškai aktyvių junginių (ARs, lignanų, β-gliukanų, aromatinių junginių (AJ), bendro fenolinių junginių kiekio (BFJ)) ir laisvųjų radikalų surišimo gebos aktyvumo (LRSGA) pokyčius augaliniuose substratuose fermentacijos metu.

3. Atlikti KF ir TF bioprocesų palyginamąjį įvertinimą, atrinkti saugius bioproduktus ir įvertinti jų antimikrobinį aktyvumą.

4. Pritaikyti sukurtus bioproduktus didesnės vertės, saugesnių ir tvaresnių gyvūninės kilmės maisto produktų technologijų prototipų sukūrimui bei tradicinių maisto produktų gamybos technologijų saugos rodiklių pagerinimui.

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Darbo aktualumas ir naujumas Sprendžiant Pasaulinės sveikatos organizacijos (PSO) dokumente

„Mityba, fizinis aktyvumas ir sveikata“ suformuotus uždavinius saugoti ir stiprinti žmogaus sveikatą bei mažinti dažniausiai pasitaikančių ligų riziką – svarbus vaidmuo tenka saugiam ir kokybiškam maistui. Siekiant užtikrinant maisto produktų saugą, gamintojai naudoja sintetinius maisto priedus, kurių nuolatinis vartojimas gali sukelti vartotojams sveikatos sutrikimų. Tenkinant augančius vartotojų poreikius natūraliems maisto produktams, būtina ieškoti naujų biotechnologinių sprendimų maisto produktų kokybei, saugai ir tvarumui užtikrinti, išskirtinį dėmesį skiriant padidintos rizikos žmonių grupių mitybai. Disertacijos mokslinis ir praktinis naujumas glūdi dviejų veiksnių – (I) Lietuvoje auginamų augalų kaip biologiškai aktyvių medžiagų šaltinio panaudojimo ir (II) KF bakteriocinus produkuojančiomis PRB pritaikymo, kuriant padidintos vertės, saugesnių ir tvaresnių maisto produktų gamybos technologijų prototipus.

Didesnės vertės, saugesnių ir tvaresnių maisto produktų kūrimas be sintetinių antioksidantų ir konservantų, taikant augalų su gausiu baltymų (sojų miltai), fitoestrogenų (Linum usitatissimum L. su sumažintu riebalų kiekiu), inulino (Helianthus tuberosus L.), fenolinių junginių, eterinių aliejų (Satureja montana L., Satureja hortensis L. ir Rhaponticum carthamoides CD.) bei skaidulinių medžiagų kiekiu (žirnių skaidulos) KF yra naujas ir perspektyvus. Disertacijos tematika atitinka prioritetines mokslinių tyrimų kryptis: biotechnologijos vystymo ir taikymo, atsinaujinančių žaliavų šaltinių panaudojimo, netradicinės žemdirbystės vystymo bei žemės ūkio sektoriuje inovacijų diegimo.

2. METODIKA

2.1. Tyrimų vieta ir laikas

Tyrimai atlikti 2012–2016 m. Lietuvos sveikatos mokslų universitete

Veterinarijos akademijoje (LSMU VA) Maisto saugos ir kokybės katedroje; Gyvūnų auginimo technologijų intitute, Gyvulių mėsinių savybių ir mėsos kokybės įvertinimo laboratorijoje (LSMU VA); Kauno technologijos universitete (KTU) Maisto mokslo ir technologijos katedroje; Vytauto Didžiojo universitete (VDU) Biologijos katedroje / Aplinkos tyrimų centre; Kauno botanikos sode (VDU); Latvijos universitete Maisto chemijos centre (Ryga, Latvija); Maisto saugos, gyvūnų sveikatingumo ir aplinkosaugos institute „BIOR“ (Ryga, Latvija) bei įmonėse ŽŪB „Nematekas“ (Dovainonys, Lietuva) ir UAB „Judex“ (Kaunas, Lietuva).

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2.2. Medžiagos

Mikroorganizmai. Bakteriocinus produkuojančios Lactobacillus sakei KTU05-6, Pediococcus acidilactici KTU05-7, Pediococcus pentosaceus KTU05-8 ir Pediococcus pentosaceus KTU05-9 išskirtos iš savaiminiu būdu fermentuotos grūdinės žaliavos [82] gautos iš KTU, Maisto mokslo ir technologijos katedros, Grūdai ir grūdų produktai mokslo grupės kolekcijos (Kaunas, Lietuva). Bacillus cereus ATCC 10876, Bacillus subtilis, Escherichia coli 1.10, Escherichia coli ATCC25922, Escherichia coli, Listeria monocytogenes 1.1, Pseudomonas fluorescens biovar. V ir Pseudomonas fluorescens biovar. III buvo gautos iš Gamtos tyrimų centro kolekcijos (Vilnius, Lietuva).

Pseudomonas aeruginosa NCTC 6570, Pseudomonas aeruginosa VUL-13, Staphylococcus aureus ATCC 9144, Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 8739, Listeria monocytogenes ATCC 35152, Salmonella enterica serovar typhimurium ATCC 13311, Bacillus cereus ATCC 11778, Yersinia enterolitica DSM 13030 ir Yersinia pseudotuberculosis III HH 146-36/84, išskirtos iš kiaulienos gamybos grandinės, gautos iš LSMU VA Maisto saugos ir kokybės katedros kolekcijos (Kaunas, Lietuva).

Gyvūninių produktų gamybai naudotas PRB, pagausintų alternatyvioje terpėje marinatas (2.2.1 lentelė).

2.2.1 lentelė. Alternatyvaus marinato rūgštingumo rodikliai (pH ir BTR) bei PRB kolonijas sudarančių vienetų skaičius ml (KSV/ml) marinato

Ferm. laikas, h

Bulvių sultys

pH BTR, °N PRB skaičius (KSV/ml

log10) Pa Pp Ls Pa Pp Ls Pa Pp Ls

72 4.10±0.04

4.15±0.0

7

4.24±0.08

8.92± 0.16

8.87± 0.13

8.53± 0.12

9.74± 0.15

9.60± 0.17

9.41± 0.11

Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; BTR – bendras titruojamas rūgštingumas (ºN); h – valandos.

Augalinės žaliavos. Eksperimente naudotos augalinės žaliavos ir jų apibūdinimas pateiktas 2.2.2 lentelėje. Augalai fermentuoti TF ir KF bakteriocinus produkuojačiomis PRB, įvertinti jų saugos ir kokybės rodikliai bei tinkamumas gyvūninės kilmės maisto produktų kokybei, saugai ir tvarumui padidinti.

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2.2.2 lentelė. Eksperimente naudotos augalinės žaliavos ir jų apibūdinimas Augalo sudėties

savitumas Augalas

Gamintojas / augintojas

Pritaikymo sritis

Didelis baltymų kiekis

Sojų miltai su sumažintu riebalų kiekiu

Gamintojas „Maxima“, įsigyti vietiniame prekybos tinkle (Kaunas, Lietuva)

Gyvūninės kilmės maisto produktų praturtinimas augaliniais baltymais

Didelis inulino kiekis

Topinambai (Helianthus tuberosus L.)

2011 metų derliaus, gauti iš Lietuvos sodininkystės ir daržininkystės instituto bandomojo ūkio (Babtai, Lietuva)

MMP (maltos mėsos produktai) praturtinimas [365]

Didelis fenolinių junginių kiekis

Kalninis dašis (Satureja montana L.) (Sm), Daržinis dašis (Satureja hortensis L.) (Sh), Paprastasis rapontikas (Rhaponticum carthamoides CD.) (Rc)

2012 metų derliaus, gauti iš VDU Kauno botanikos sodo kolekcijos

Sm ir Sh panaudoti maltos mėsos pusgaminių (MMP) gamybai bei kiaulienos ir jautienos nugarinės apdorojimui. Sm ir Rc panaudoti varškės sūrių (VS) gamyboje.

Didelis lignanų kiekis

Linų sėmenys su sumažintu riebalų kiekiu (Linum usitatissimum L.)

Gamintojas (Institut Wlokien Naturalnych, Poznanė, Lenkija), įsigyti vietiniame prekybos tinkle (Kaunas, Lietuva)

Gyvūninės kilmės maisto produktų praturtinimas fitoestrogenais

Didelis skaidulinių medžiagų kiekis

Žirnių skaidulos

Gamintojas (M plant Ltd., Hamburgas, Vokietija), įsigyti vaistinėje (Kaunas, Lietuva)

Panaudotos MMP be glitimo gamybai

Sm – S. montana, Sh – S. hortensis, Rc – R. carthamoides, MMP – maltos mėsos produktai, VS – varškės sūris

Maisto produktų gamybai naudotos gyvūninės kilmės žaliavos.

Marinuotos kiaulienos ir jautienos gamybai naudotos skerdenų dalys: sprandinė, mentė, kumpis, ilgiausiasis nugaros raumuo (INR) ir nugarinė, įsigytos vietiniame prekybos tinkle (Kaunas, Lietuva). Mėginiai supjaustyti 2,5 cm storio gabalais, jų marinavimui naudoti bioproduktai: alternatyvioje terpėje pagausintos PRB bei S. montana (Sm) ir S. hortensis (Sh) bioproduktai. Pagamintiems marinuotiems mėsos gaminiams įvertinti šie

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kokybės rodikliai: pH, SM (sausosios medžiagos), virimo nuostoliai (VN), švelnumas, drėgmės nuostoliai, tarpraumeninių riebalų kiekis (TR), vandens rišlumas (VR), spalvų koordinatės, biogeninių aminų kiekis (BA) ir bendras priimtinumas.

Marinuotos P. acidilactici marinatu, ekologiškai (Kaunas, Lietuva) ir tradiciškai pagamintos (įsigytos vietiniame prekybos tinkle, Kaunas, Lietuva) vištienos gamybai naudotos skerdenėlių dalys: krūtinėlės, šlaunelės ir blauzdelės. Marinuotiems vištienos gaminiams įvertinti šie kokybės rodikliai: pH, SM, VR, švelnumas, drėgmės nuostoliai, TR, spalvų koordinatės, bendras priimtinumas, atlikta BA kokybinė ir kiekybinė analizė.

Maltos mėsos produktų (MMP) gamybai naudota kiaulienos nugarinė, įsigyta vietiniame prekybos tinkle (Kaunas, Lietuva). Mėginiai supjaustyti 2,5 cm storio gabalėliais ir sumalti. Jų apdorojimui naudoti: Sm ir Sh bioproduktai, manų kruopos ir žirnių skaidulos. MMP praturtinimui [365] buvo sukurti topinambų bioproduktai. Įvertinti šie MMP kokybės rodikliai: sporas formuojančių aerobinių mezofilinių bakterijų, Enterobakterijų, mielių ir pelėsinių grybų kiekis, pH, SM kiekis, VN, VR, švelnumas, drėgmės nuostoliai, TR, spalvų koordinatės, BA kiekis bei AJ kiekis ir bendras priimtinumas.

Šaltai rūkytos dešros pagamintos ŽŪB „Nematekas“ (Dovainonys, Lietuva). Prieš ir po rūkymo dešros apdorotos alternatyvioje terpėje pagausintos PRB. Įvertinti šie kokybės rodikliai: PRB, naudotų dešrų apdorojimui, antimikrobinis aktyvumas, BA kiekis, policiklinių aromatinių angliavandenilių (PAA) kiekis ir bendras priimtinumas.

Varškės sūriai (VS) pagaminti iš šviežio karvės pieno (Mažeikiai, Lietuva) su Sm ir Sh bioproduktų preidais. Pagamintiems sūriams įvertinti šie kokybės rodikliai: augalinių bioproduktų antipelėsinis aktyvumas, BTR (ºT), pH, L(+) ir D(-)-pieno rūgšties izomerų kiekis, BA kiekis ir AJ kiekis. Sūrių priimtinumas įvertintas standartiniu metodu bei Facereader (Noldus, Olandija) programine įranga, skanuojant ir fiksuojant degustatorių emocines veido išraiškas, ragaujant produktą.

2.3. Augalinių žaliavų fermentacija

Augalinių žaliavų fermentacijai naudotos PRB: L. sakei; P. acidilactici;

P. pentosaceus 8 (2 %, m/m), fermentacijos trukmė 48 valandos, kiekvienai PRB optimalioje augimo temperatūroje. Augalinio substrato drėgnis apskaičiuotas pagal substrato drėgį ir gebėjimą sugerti vandenį (KF – ne daugiau nei ≤ 50 % ir TF ne mažiau nei ≥ 70 %).

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2.4. Augalinių bioproduktų tyrimų metodai SM kiekis nustatytas džiovinant termostate (UNB400, Memmert,

Vokietija) (103 ± 2) °C temperatūroje iki pastovios masės. Bendrasis mezofilinių PRB, sporas formuojančių aerobinių mezofilinių

bakterijų, Enterobakterijų, mielių ir pelėsininių grybų skaičius nustatytas pagal [262] aprašytą metodiką.

Satureja montana L., Satureja hortensis L. ir Rhaponticum carthamoides CD. augalų ir jų bioproduktų eterinių aliejų ekstrakcija atlikta pagal [176] aprašytą metodiką.

Eterinių aliejų antimikrobinis aktyvumas įvertintas, taikant diskų difuzijos metodą pagal [31] aprašytą metodiką.

PRB antimikrobinis aktyvumas įvertintas pagal [28] aprašytą tyrimo metodiką.

Terpės pH išmatuotas pH – metru (PP – 15, Sartorius, Gotingenas, Vokietija).

Bendras titruojamas rūgštingumas (BTR) nustatytas vandeniniuose mėginių tirpaluose, juos titruojant 0,1 N NaOH tirpalu iki terpės pH=8,5. Rūgštingumas įvertintas Neimano laipsniais (°N).

Bendras pieno rūgšties kiekis ir L-(+)/D-(-) pieno rūgšties izomerų kiekis nustatytas fermentiniu metodu, pagal [165] bei [75] metodikas.

BFJ kiekis nustatytas pagal [387] metodiką. Bendras β-gliukanų kiekis nustatytas fermentiniu metodu [243] metodiką

(Megazyme International, Bray, Airija). ARs, lignanų ir LRSGA kokybinė ir kiekybinė analizė atlikta pagal [30] aprašytą metodiką.

BA analizė atlikta pagal [33] metodiką. Amilolitinių fermentų aktyvumas nustatytas pagal [268] metodiką,

proteolitinių fermentų aktyvumas – pagal [69] metodiką.

2.5. Gyvūninės kilmės maisto produktų technologija

Kiaulienos ir jautienos nugarinės apdorojimas Sm ir Sh bioproduktais. Nugarinės mėginiai (masė 100 g) apdoroti Sm ir Sh bioproduktais ir marinuoti 24 valandas +4 ± 1°C temparatūroje.

Skirtingų kiaulienos ir jautienos skerdenos dalių apdorojimas alternatyvioje terpėje pagausintomis PRB. Kiaulienos ir jautienos (sprandinė, mentė, kumpis, INR ir nugarinė) skerdenos dalių mėginiai (masė 100 g) apdoroti alternatyviu marinatu ir marinuoti 24 valandas +4 ± 1 °C temperatūroje.

Tradiciškai ir ekologiškai pagamintos vištienos marinavimas. Tradiciškai ir ekologiškai pagamintos vištienos (krūtinėlės, blauzdelės,

221

šlaunelės) skerdenėlių dalių mėginiai (masė 40 g) apdoroti alernatyviu marinatu (fermentuotu P. acidilactici) 12 valandų +4 ± 1 °C temperatūroje.

Šaltai rūkytų dešrų apdorojimas. Šaltai rūkytų dešrų paviršius prieš rūkymą (60 minučių, 18 ± 2 °C) ir po jo (24 valandas, 18 ± 2 °C) apdorotas PRB, pagausintomis alternatyvioje terpėje.

Maltos mėsos pusgaminių gamybos technologija. MMP pagaminti su 3; 5 ir 7 % Sm ir Sh bioproduktų, MMP su 5 % žirnių skaidulų ir manų kruopų priedais (masė 50 g) laikyti 5 paras +4 ± 1 °C temperatūroje.

VS gamybos technologija. VS pagaminti pagal tradicinę technologiją ir praturtinti 3 % Sm ir Rc bioproduktų priedo (masė 100 g), iki tyrimų 12 valandų laikyti 4 ± 1°C temperatūroje.

2.6. Gyvūninės kilmės maisto produktų tyrimo metodai

pH nustatytas pagal metodiką, pateiktą 2.4. skyriuje. BTR nustatytas vandeniniuose mėginių tirpaluose, juos titruojant 0,1 N

NaOH tirpalu. Rūgštingumas įvertintas Ternerio laipsniais (°T). SM kiekis nustatytas pagal [18] metodą. Mėsos VN nustatyti 100 g mėsos mėginį verdant cirkuliacinėje vandens

vonelėje 15 minučių 100 oC temperatūroje, po to apskaičiuojant procentinį masės pokytį.

Mėsos švelnumas nustatytas Warner – Bratzler tipo prietaisu, kuriuo įvertinama jėga, reikalinga perpjauti mėsos mėginį specialiu peiliu.

Mėsos drėgmės nuostoliai nustatyti pagal [105] metodiką. TR kiekis nustatytas Skoksleto metodu pagal [107] metodiką. VR nustatytas pagal [127] metodiką. Spalvų koordinatės nustatytos trijose skirtingose paviršiaus vietose

CIELab skalėje (CromaMeter CR-400, Conica Minolta, Japonija). L* − šviesumas, nuo visiškai nepermatomo (0) iki visiškai balto (100), a* − rausvumas (arba žalsvumas), b* − gelsvumas (arba mėlynumas) [244].

Mėsos produktų bendras priimtinumas nustatytas pagal [162]. VS juslinė analizė atlikta pagal [160] standartą ir Face Reader

programine įranga (Noldus Information Technology, Vageningenas, Nyderlandai) naudojant 3 pagrindinių emocijų skalę (laimingas, liūdnas ir piktas).

BA, L(+ ) ir D(-) pieno rūgšties izomerai nustatyti pagal metodikas, pateiktas 2.4. skyriuje.

PAA nustatyti pagal metodiką, aprašytą [28]. AJ analizė atlikta pagal [176] aprašytą metodiką.

Sm ir Rc džiovintų augalų ir jų bioproduktų įtaka mielių augimui VS laikymo metu nustatyta pagal [262] aprašytą metodiką.

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2.7. Statistinė analizė

Produktų gamyba kartota tris kartus, tiriant paraleliai tris mėginius. Skirtingų veiksnių (fermentacijos būdo, augalinio bioprodukto ir pasirinkto mikroorganizmo) įtaka produktų rodikliams įvertinta atlikus daugiafaktorinę dispersinę analizę (ANOVA), taikant Tjukio-HSD testą (statistinė programa R3.2.1, Core Team 2015). Rezultatai reikšmingi, kai p ≤ 0,05.

3. TYRIMŲ REZULTATAI

3.1. Augalinių žaliavų ir jų bioproduktų mikrobiologiniai ir fizikiniai cheminiai rodikliai

Augalinių žaliavų ir jų bioproduktų mikrobiologiniai rodikliai.

Augalinių bioproduktų PRB KSV/g kito nuo 6.21 log10 iki 9.98 log10, atitinkamai, P. acidilactici TF apdorotuose topinambuose ir P. pentosaceus KF apdorotuose sojų miltuose (3.1.1 pav.). Fermentacijos metodas ir fermentacijai naudotos PRB turėjo reikšmingos įtakos sporas formuojančių aerobinių mezofilinių bakterijų (fermentacijos metodas p ≤ 0,0001, PRB p ≤ 0,0001, faktorių sąveika p ≤ 0,0001), Enterobakterijų (fermentacijos metodas p ≤ 0,0001, PRB p ≤ 0,0001, faktorių sąveika p ≤ 0,0001) bei mielių ir pelėsinių grybų ( fermentacijos metodas p ≤ 0,0002, PRB p ≤ 0,0001, faktorių sąveika p ≤ 0,0001) kiekiui vaistiniuose – prieskoniniuose augaluose.

3.1.1 pav. Vaistinių – prieskoninių augalų mikrobiologiniai rodikliai.

Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė fermentacija, TF – tradicinė fermentacija, Sm – S. montana, Sh – S. hortensis, Rc – R.

carthamoides, SFAMB – sporas formuojančios mezofilinės bakterijos, Ent – Enterobakterijos, M/P – mielės ir pelėsiai, p – skirtumo tarp rezultatų reikšmių

patikimumas, p – patikimas, kai ≤ 0,05

0

2

4

6

8

10

SmPRB

ShPRB

RcPRB

SmSAMB

ShSAMB

RcSAMB

SmEntB

ShEntB

RcEntB

SmM/P

ShM/P

RcM/P

KSV

/g lo

g 10

Ls TF Ls KF Pa TF Pa KFPp TF Pp KF NF

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Vaistinių – prieskoninių augalų, jų bioproduktų ekstraktų ir fermentacijai naudotų PRB antimikrobinis aktyvumas. Didžiausias antimikrobinis aktyvumas nustatytas Satureja rūšies augalų ekstraktų prieš E. coli, P. fluorescens ir B. subtilis (3.1.1 lentelė). 3.1.1 lentelė. Vaistinių – prieskoninių augalų ir jų bioproduktų antimikrobinis aktyvumas.

Mikroorganizmas Inhibicinė zona/mm Pa7 Pp8 L. sakei Nef.

S. montana E. coli 9,0±0,09b 11,8±0,11d 9,4±0,10b 8,6±0,09a B. subtilis 6,5±0,07a 6,8±0,07b 8,8±0,09d 6,8±0,08b P. fluorescens biovar. III 13,2±0,14c 14,9±0,15d 14,9±0,16d 11,0±0,12a P. fluorescens biovar. V 16,0±0,18d 18,8±0,19e 18,0±0,20e 14,5±0,15a

S. hortensis E. coli 7,5±0,09a 9,8±0,09d 8,6±0,09c 7,3±0,07a B. subtilis 6,2±0,05c 5,8±0,06a 7,5±0,08d 6,6±0,07c P. fluorescens biovar. III 14,6±0,11d 12,4±0,10a 14,5±0,17d 13,8±0,14c P. fluorescens biovar. V 17,2±0,21e 16,4±0,15c 17,8±0,8e 15,6±0,16a

R. carthamoides E. coli 8,6±0,09b 8,6±0,09b 9,0±0,10c 8,0±0,09a B. subtilis - - - - P. fluorescens biovar. III 12,8±0,13e 11,2±0,12c 12,0±0,12e 9,2±0,09a P. fluorescens biovar. V 12,0±0,10c 12,0±0,12c 12,0±0,13c 10,0±0,11a Paaiškinimai: Pa – P. acidilactici, Pp – P. pentosaceus. p – patikimas, kai ≤ 0,05. a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės eilutėse pažymėti skirtingomis raidėmis, tarpusavyje patikimai skiriasi (p < 0,05)

Augalų bioproduktų antimikrobinis aktyvumas nustatytas didesnis nei

atskirų jų komponentų (augalo ar PRB), tačiau priklauso nuo bioproduktų atskirų komponentų ir yra specifiškas patogeninių mikroorganizmų rūšiai (bioproduktuose PRB turėjo reikšmingos įtakos E.coli p ≤ 0,010, B. subtilis p ≤ 0,0001, P. fluorescens biovar. III p ≤ 0,023 ir P. fluorescens biovar. V p ≤ 0,001, augalo rūšis – E.coli p ≤ 0,002, B. subtilis, P. fluorescens biovar. III, P. fluorescens biovar. V p ≤ 0,0001).

Augalinių žaliavų ir jų bioproduktų fizikiniai cheminiai rodikliai. Tirtų augalų (Sm, Sh, Rc, sojų miltų, linų sėmenų, topinambų ir žirnių skaidulų) bioproduktų fizikiniai cheminiai rodikliai (3.1.2 pav.) kito priklausomai nuo fermentacijos technologijos ir augalinės matricos specifikos. Mažesnis augalinių bioproduktų pH ir didesnis BTR nustatytas TF apdorotuose augaluose (atitinkamai, vidutinės vertės TF – 4,76, KF – 5,53 ir TF – 9,98 ºN, KF – 9,03 ºN). KF apdoroti bioproduktai saugesni, nes juose mažesnis D(-) pieno rūgšties izomero kiekis, o L(+) izomero nustatyta nuo 4,0 % iki 1148,9 % daugiau, atitinkamai, Sm ir topinambuose.

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a)

b)

c)

d)

e)

f)

3.1.2 pav. Augalų bioproduktų a) pH, b) bendras titruojamas rūgštingumas, Nº, c) L(+) ir D(-) pieno rūgšties izomerai, g/100 g, d-e) amilolitinis ir

proteolitinis fermentų aktyvumas, AU/g, f) biogeninių aminų kiekis, mg/kg (Paaiškinimai; ŽS – žirnių skaidulos, Lu – linų sėmenys, Ht – topinambai, SM – sojų miltai,

Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė, TF – tradicinė fermentacija)

01234567

TF KF TF KF TF KF

Ls Pa Pp

SM

ŽS

Lu

Ht

Sm

Sh

Rc

0

5

10

15

20

25

TF KF TF KF TF KF

Ls Pa Pp

NºSM

ŽS

Lu

Ht

Sm

Sh

Rc

0

5

10

15

L(+

)

D(-

)

L(+

)

D(-

)

L(+

)

D(-

)

L(+

)

D(-

)

L(+

)

D(-

)

L(+

)

D(-

)

TF KF TF KF TF KF

Ls Pa Pp

g/100 g

SM Ht Lu ŽS Sm Sh Rc

0

500

1000

1500

TF KF TF KF TF KF

Ls Pa Pp

AU/g

SM HtLu ŽS

0

500

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1500

TF KF TF KF TF KF

Ls Pa Pp

AU/g

SM HtLu ŽS

0 100 200

TF

KF

TF

KF

TF

KF

Ls

Pa

Pp

mg/kg

ŽS Lu Ht SM

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Amilolitinių ir proteolitinių fermentų aktyvumas KF ir TF apdorotuose augaluose kito nevienareikšmiai – didesnis amilolitinių fermentų aktyvumas nustatytas KF apdorotuose bioproduktuose (vidutiniškai nuo 0,83 iki 3,69 karto, atitinkamai, topinambuose ir sojų miltuose), o didesnis proteolitinių fermentų aktyvumas – TF apdorotuose bioproduktuose (vidutiniškai nuo 1,08 iki 4,52 karto, atitinkamai, žirnių skaidulose ir linų sėmenyse). Amilolitinių ir proteolitinių fermentų aktyvumui substrate reikšmingos įtakos turėjo fermentacijos metodas, naudotos PRB ir augalo rūšis (p ≤ 0,0001).

Analizuoti veiksniai turėjo reikšmingos įtakos BA formavimuisi substratuose (naudotos PRB (p ≤ 0,018) ir augalo rūšis (p ≤ 0,0001) turėjo reikšmingos įtakos suminiam BA kiekiui) (3.1.2. pav. e) Lyginant KF ir TF procesų įtaką BA formavimuisi nustatyta, kad augalo rūšis buvo reikšmingas faktorius: didesnis suminis BA kiekis nustatytas KF sojų miltuose (19,7 %), žirnių skaidulose (4,2 %) ir topinambuose (2,1 %), tačiau KF mažino suminį BA kiekį linų sėmenyse (22,4 %).

Fermentacijos įtaka BFJ, ARs, lignanų ir β-gliukanų kiekiui bei

LRSGA žirnių skaidulose. Žirnių skaidulų fermentacijai naudotos PRB turėjo reikšmingos įtakos matairezinolio (MAT) ir sekoizolaricirezinolio (SEKO) (p ≤ 0,001) (padidėjo, atitinkamai, 23,8 % ir 57,5 %, nuo vidutinės vertės) ir suminiam lignanų (padidėjo vidutiniškai TF 27,9 %, KF – 41,0 %) kiekiui (3.1.2 lentelė). 3.1.2 lentelė. BFJ, β-gliukanų, ARs ir lignanų kiekis žirnių skaidulose

Žirnių skaidulos

BFJ, mg GAE/100g

LRSGA, % β-gliukanai,

%

ARs kiekis, μg/g

Lignanų kiekis,

μg/100 g

Ls TF 102,13±1,81c 17,61±0,18d 1,40±0,01e 97±0,87c 158,3±2,61b KF 105,98±1,53c 18,88±0,20e 1,32±0,02d 125±0,45d 176,4±2,93d

Pa TF 102,31±2,10c 17,04±0,17c 1,29±0,01c 72±0,36a 218,4±4,01e KF 102,94±2,15c 17,86±0,21d 1,24±0,02c 122±0,67d 238,6±4,85e

Pp TF 107,63±4,12c 18,33±0,20e 0,90±0,01a 161±0,80e 167,8±2,84c KF 119,18±3,51d 18,95±0,19e 0,95±0,01a 74±0,69a 165,5±3,12c

Nefermentuota 22,05±0,25a 5,83±0,07a 1,41±0,03e 267±0,84f 134,6±4,50a Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė fermentacija, TF – tradicinė fermentacija, BFJ – bendras fenolinių junginių kiekis, LRSGA – laisvųjų radikalų surišimo geba; ARs – alkilrezorcinoliai, p – skirtumo tarp rezultatų reikšmių patikimumas, p – patikimas, kai ≤ 0,05. a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės eilutėse pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Didesnis BFJ kiekis ir LRSGA nustatytas KF apdorotuose augaluose

(atitinkamai, nuo 0,6 % P. acidilactici iki 10,7 % P. pentosaceus ir nuo 3,4

226

% P. pentosaceus iki 7,2 % L. sakei fermentuotose žirnių skaidulose), lyginant su TF. Fermentacijai naudotos PRB turėjo reikšmingos įtakos BFJ kiekiui ir LRSGA (atitinkamai, p ≤ 0,001 ir p ≤ 0,002).

Priešingos tendencijos nustatytos analizuojant ARs ir β-gliukanų pokyčius fermentacijos metu, jų kiekis žirnių skaidulose sumažėjo (atitinkamai, nuo 40,0 % P. acidilactici KF iki 73,0 % P. pentosaceus KF ir nuo 17,6 % L. sakei TF iki 77,3 % P. acidilactici KF apdorotuose augaluose). Šiems pokyčiams reikšmingos įtakos turėjo fermentacijos technologija (p ≤ 0,0001; p ≤ 0,021) ir jai naudotos PRB (p ≤ 0,0001; p ≤ 0,014).

3.2. Mėsos produktų technologiniai, mikrobiologiniai ir fizikiniai cheminiai rodikliai

3.2.1. Marinuoti mėsos produktai

Sm ir Sh bioproduktų įtaka kiaulienos nugarinės kokybės rodikliams. Fermentacijos metodas (p ≤ 0,0001), fermentacijai naudotos PRB (p ≤ 0,0001) ir augalo rūšis (p ≤ 0,0001) turėjo reikšmingos įtakos marinuotos kiaulienos nugarinės technologiniams rodikliams (3.2.1.1 lentelė). 3.2.1.1 lentelė. Kiaulienos nugarinės, apdorotos Sm ir Sh augaliniais bioproduktais, technologiniai rodikliai. Mėg. SM, % pH Drėgmės

nuostoliai, %

Vandens rišlumas,

%

Virimo nuostoliai,

%

Švelnumas, kg/cm2

S. montana Pa TF 22,58±0,03a 6,0±0,05a 3,12±0,03c 65,31±0,6b 42,1±0,19c 3,16±0,06d Pa KF 23,36±0,04b 6,27±0,03b 3,16±0,06c 67,52±0,6c 40,8±0,20b 1,77±0,02a Pp TF 24,66±0,05b 5,98±0,02a 3,04±0,04b 68,61±0,5d 42,2±0,17c 2,05±0,02b Pp KF 24,70±0,03b 5,97±0,06a 3,37±0,06d 66,76±0,5b 37,1±0,26a 2,45±0,02c Ls TF 29,61±0,03c 5,86±0,07a 2,96±0,04b 63,58±0,6a 36,4±0,19a 2,14±0,03b Ls KF 22,51±0,02a 6,59±0,08c 2,61±0,02a 67,14±0,2c 40,5±0,23b 2,03±0,04b

S. hortensis Pa TF 22,25±0,04a 6,12±0,04a 3,01±0,03a 68,22±0,3d 34,7±0.17b 1,48±0,01c Pa KF 22,23±0,05a 6,56±0,06b 2,98±0,03a 67,39±0,2c 41,0±0.18d 1,41±0,01c Pp TF 26,08±0,03c 6,10±0,07a 3,15±0,04b 65,13±0,2a 34,2±0.22b 1,16±0,02a Pp KF 25,20±0,03c 6,18±0,06a 3,21±0,05b 67,54±0,5c 34,5±0.19b 1,25±0,03b Ls TF 24,71±0,03b 6,28±0,07a 2,93±0,03a 68,37±0,5d 32,6±0.16a 1,59±0,04c Ls KF 21,62±0,02a 6,30±0,06b 3,03±0,03a 66,31±0,2b 36,5±0.26c 1,87±0,03d Kontr. 22,65±0,02a 6,52±0,08c 3,06±0,02a 66,20±0,4b 39,5±0.18d 1,05±0,02a Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė fermentacija, TF – tradicinė fermentacija, SM – sausosios medžiagos, p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

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KF fermentuoti Sm ir Sh bioproduktai daugeliu atvejų didino mėsos pH (nuo 0,32 % L. sakei Sh iki 12,46 % P. pentosaceus Sm) bei drėgmės nuostolius (vidutiniškai 0,10 % Sm ir 1,44 % Sh). Daugeliu atvejų didesnis VR nustatytas Sm ir Sh bioproduktais marinuotoje mėsoje (Sm KF – 1,42 %, Sh KF – 1.33 %, TF – 1.65 %), o Sh bioproduktai mažino VN (vidutiniškai 9,82 %).

Marinavimas Sm ir Sh bioproduktais mažino mėsos švelnumą (vidutiniškai 55,51 % ir 35,50 % TF ir 48,72 % ir 28,53 % KF fermentuoti augaliniai bioproduktai) ir TR kiekį (Sm – 23,07 %, Sh – 26,25 %), lyginant su kontroliniais mėginiais.

Fermentacijos technologija, naudotos PRB bei augalo rūšis turėjo reikšmingos įtakos mėsos VR, VN, švelnumui ir TR kiekiui (p ≤ 0,0001).

3.2.1.1 pav. Kiaulienos nugarinės, apdorotos Sm ir Sh augaliniais

bioproduktais, spalvų koordinatės (Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė fermentacija, TF – tradicinė fermentacija, SM – sausosios

medžiagos, p – patikimas, kai ≤ 0,05, L* − šviesumas, a* – rausvumas, b* – gelsvumas, K – kontrolė)

Fermentacijos metodas (p ≤ 0,0001), fermentacijai naudotos PRB (p ≤

0,0001) ir augalo rūšis (p ≤ 0,0001) turėjo reikšmingos įtakos Sm ir Sh bioproduktais apdorotos kiaulienos nugarinės spalvų koordinatėms (3.2.1.1 pav.). Augaliniai bioproduktai mažino mėsos (L)* ir (b*) (vidutiniškai Sm (17,24 % TF ir 24,18 % KF), ir Sh (5,48 % TF ir 16,63 % KF)) bei didino (a*) (vidutiniškai Sm – 117,24 % ir Sh – 58.37 %).

Sm ir Sh bioproduktų įtaka jautienos nugarinės kokybės rodikliams.

Augaliniai bioproduktai (jų fermentacijos technologija p ≤ 0,0001, naudotos PRB p ≤ 0,0001 ir augalo rūšis p ≤ 0,0001) turėjo reikšmingos teigiamos įtakos marinuotos jautienos nugarinės technologiniams rodikliams (3.2.1.2 lentelė).

01020304050607080

TF KF TF KF TF KF TF KF TF KF TF KF

Ls Pa Pp Ls Pa Pp K

S. montana S. hortensis

b*

a*

L*

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3.2.1.2 lentelė. Jautienos nugarinės, apdorotos Sm ir Sh augaliniais bioproduktais, technologiniai rodikliai.

Mėg. SM, % pH Drėgmės

nuostoliai, %

Vandens rišlumas,

%

Virimo nuostoliai,

%

Švelnumas, kg/cm2

S. montana Pa TF 22,57±0,2a 6,05±0,3a 2,18±0,1c 62,46±0,1b 40,45±0,1d 2,80±0,09c Pa KF 23,41±0,1b 6,40±0,7c 2,23±0,2c 63,03±0,1c 38,72±0,13c 2,88±0,05d Pp TF 24,27±0,5c 6,02±0,3a 1,96±0,1a 61,94±0,14 39,58±0,14c 2,40±0,01a Pp KF 23,77±0,2b 6,30±0,5b 2,05±0,2b 62,57±0,1b 40,71±0,2d 2,72±0,03c Ls TF 22,80±0,2a 6,24±0,5b 1,96±0,1a 62,35±0,1b 38,04±0,12c 2,54±0,02b Ls KF 24,47±0,3c 6,31±0,6b 1,98±0,1a 63,41±0,1c 31,70±0,10a 2,78±0,04c

S. hortensis Pa TF 22,29±0,1b 6,32±0,5c 2,03±0,3b 64,55±0,1d 37,88±0,17a 2,53±0,01c Pa KF 21,16±0,3a 6,11±0,3a 2,11±0,3c 63,74±0,1c 39,61±0,13b 2,59±0,02c Pp TF 25,11±0,3c 6,25±0,4b 2,07±0,2b 63,07±0,1b 38,31±0,11b 2,31±0,02b Pp KF 21,29±0,2a 6,29±0,6b 1,96±0,2a 64,12±0,1c 37,85±0,16a 3,37±0,03d Ls TF 20,78±0,2a 6,09±0,4a 1,89±0,1a 63,26±0,2b 40,39±0,16b 3,16±0,03d Ls KF 22,58±0,3b 6,16±0,7a 2,03±0,1b 61,88±0,2a 37,26±0,13a 1,90±0,01a Kontr. 22,46±0,02b 6.33±0,07c 2,17±0,02c 61,89±0,1a 42,75±0,16c 3,86±0,09e Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė fermentacija, TF – tradicinė fermentacija, SM – sausosios medžiagos, p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Sm ir Sh bioproduktai sumažino mėsos pH (atitinkamai, vidutiniškai 1,74

% ir 2,0 %) ir drėgmės nuostolius (vidutiniškai Sm KF – 3,84 %, Sm TF – 6,30 %, Sh KF – 6,30 %, TF – 7,99 %), tačiau padidino marinuotos mėsos VR (atitinkamai, nuo 0,08 % P. pentosaceus KF Sm iki 4,30 % P. acidilactici TF Sh) ir sumažino VN (vidutiniškai Sm KF – 13,34 %, Sm TF – 7,94 %, Sh KF – 10,55 %, ShTF – 9,10 %). KF apdoroti Sm ir Sh daugeliu atvejų didino mėsos švelnumą, lyginant su TF apdorotais (nuo 2,37 % P. acidilactici Sh iki 45,89 % P. pentosaceus Sh) ir sumažino TR kiekį (Sm – 0,90 %, Sh – 4,56 %, lyginant su kontroliniais mėginiais).

Fermentacijos technologija, naudotos PRB bei augalo rūšis turėjo reikšmingos įtakos mėsos drėgmės nuostoliams, VR, VN, švelnumui ir TR kiekiui (p ≤ 0,05) ir veiksnių sąveika nustatyta reikšminga (p ≤ 0,0001).

Augaliniai bioproduktai patikimai mažino (L*) ir (a*) mėsos spalvų koordinates (atitinkamai, vidutiniškai Sm 29,07 % ir Sh 31,26 %, ir Sm 35,49 % ir Sh 34,23 %) (3.2.1.3 lentelė).

Marinavimas Sm bioproduktais sumažino jautienos gelsvumą vidutiniškai 17,78 %, o Sh – padidino vidutiniškai 51,64 %. Mėsos (L*), (a*) ir (b*) spalvų koordinatės reikšmingai kito priklausomai nuo augalinių bioproduktų apdorojimui taikytos fermentacijos technologijos (p ≤ 0,04; p ≤

229

0,0001; p ≤ 0,0001), naudotų PRB (p ≤ 0,0001) ir augalo rūšies (p ≤ 0,0001).

3.2.1.3 lentelė. Jautienos nugarinės, apdorotos Sm ir Sh augaliniais bioproduktais spalvų koordinatės

P. L. sakei P. acidilactici 7 P. pentosaceus 8 TF KF TF KF TF KF

S. montana L* 29,42

±0,28c 25,82 ±0,20a

27,56 ±0,25b

26,97 ±0,22a

25,49 ±0,21a

26,59 ±0,29a

a* 14,65±0,16d 9,33±0,08b 9,02±0,08b 8,62±0,07a 7,90±0,09a 9,16±0,08b b* 3,51±0,06c 1,78±0,03b 1,38±0,02a 0,63±0,01a 1,40±0,02a 2,40±0,19b TR 1,65±0,01a 2,44±0,04d 2,01±0,03b 2,45±0,04d 2,46±0,05d 2,25±0,06c

S. hortensis L* 25,82

±0,29b 25,34 ±0,30b

24,91 ±0,20a

27,29 ±0,26c

27,41 ±0,28c

26,09 ±0,30b

a* 11,64 ±0,08c

7,03 ±0,06b

9,16 ±0,08b

11,97 ±0,13c

11,93 ±0,11c

11,37 ±0,12c

b* 3,83±0,05c 1,42±0,02a 2,87±0,03b 4,28±0,05d 4,20±0,05d 4,23±0,04d TR 4,14±0,05e 1,68±0,01c 0,98±0,01a 2,27±0,03d 1,70±0,03c 2,00±0,02d

Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, KF – kietafazė fermentacija, TF – tradicinė fermentacija, TR – tarpraumeniniai riebalai, L* − šviesumas, a* – rausvumas, b* – gelsvumas, P – parametrai, p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Alternatyvioje terpėje pagausintų P. acidilactici KTU05-7, P.

pentosaceus KTU05-8 ir L. sakei KTU05-6 įtaka marinuotos kiaulienos kokybės rodikliams. Patikimi skirtumai (p ≤ 0,05) nustatyti tarp PRB marinuotų ir nemarinuotų kiaulienos skerdenų dalių (sprandinės, mentės, kumpio, INR ir nugarinės) mėsos technologinių rodiklių (3.2.1.4 lentelė).

Apdorojimas alternatyviu marinatu sumažino kiaulienos pH (nuo 2,81 % iki 21,17 %, atitinkamai, P. pentosaceus marinuotos mentės ir L. sakei marinuoto INR) ir VR (nuo 2,52 % iki 27,45 %, atitinkamai, P. acidilactici marinuoto kumpio ir P. pentosaceus marinuoto INR).

Mažiausi drėgmės nuostoliai nustatyti L. sakei marinuotuose mėginiuose (vidutiniškai 46,63 %), didžiausi – P. acidilactici (vidutiniškai 75,30 %). Didesni VN (vidutiniškai, marinuotų P. acidilactici – 24,47 %, P. pentosaceus – 23,90 % ir L. sakei – 20,83 %) ir švelnumas (nuo 2,09 % L. sakei marinuotos sprandinės iki 7,33 % P. acidilactici marinuoto kumpio) nustatyti marinuotos mėsos mėginių, lyginant su nemarinuotais.

230

3.2.1.4 lentelė. Skirtingų kiaulienos skerdenos dalių, marinuotų alternatyvioje terpėje pagausintomis PRB, technologiniai rodikliai.

Mėg.

TR, %

pH Drėgmės nuostoliai,

%

Vandens rišlumas,

%

Virimo nuostoliai,

%

Švelnum., kg/cm2

Sprandinė

Pa 5,46±0,03b 6,20±0,1b 3,20±0,03d 56,7±0,10a 44,2±0,23d 0,84±0,01a Pp 5,21±0,01a 6,35±0,1b 2,25±0,02c 56,9±0,10a 44,6±0,47d 0,85±0,02a Ls 5,42±0,04b 6,03±0,1a 2,54±0,02c 58,9±0,12b 40,4±0,21b 0,86±0,02b K. 5,57±0,03c 6,70±0,3c 1,52±0,01a 67,8±0,18c 38,3±0,31a 0,88±0,01b

Mentė Pa 3,90±0,01b 5,88±0,2a 2,85±0,03c 56,1±0,10b 44,3±0,47d 0,85±0,03a Pp 3,85±0,01a 5,89±0,1a 2,60±0,03c 53,8±0,11a 44,3±0,43d 0,86±0,04a Ls 3,72±0,02a 5,84±0,1a 2,55±0,02c 60,2±0,16c 43,2±0,38c 0,88±0,02b K. 5,91±0,04d 6,06±0,1b 1,84±0,01a 64,3±0,17d 39,4±0,34a 0,90±0,01b

Kumpis Pa 4,66±0,04a 5,24±0,1b 3,10±0,03c 63,6±0,16c 43,8±0,40c 1,02±0,05a Pp 4,90±0,02b 5,33±0,0b 2,86±0,03c 53,1±0,10a 44,3±0,43c 1,04±0,03a Ls 4,89±0,03b 5,09±0,1a 1,95±0,01a 56,7±0,14a 44,6±0,51d 1,06±0,04b K. 5,29±0,01c 6,42±0,2c 1,74±0,01a 65,2±0,15c 35,1±0,23a 1,10±0,02c

M. longissimus dorsi Pa 2,53±0,01a 5,30±0,1b 2,62±0,03c 66,2±0,15c 41,2±0,31d 0,84±0,08a Pp 2,44±0,02a 5,52±0,1b 2,45±0,02b 54,0±0,11a 39,1±0,30c 0,86±0,04a Ls 2,76±0,01b 5,14±0,1a 2,76±0,03c 53,7±0,10a 40,7±0,36d 0,91±0,04c K. 3,24±0,01d 6,52±0,2c 2,01±0,02a 74,5±0,18d 29,4±0,20a 0,89±0,01b

Nugarinė Pa 3,50±0,02a 5,30±0,1c 3,14±0,03d 55,1±0,10b 42,8±0,41d 1,02±0,03a Pp 3,41±0,01a 5,41±0,1c 3,07±0,03d 55,5±0,11b 43,4±0,44d 1,06±0,06b Ls 3,61±0,02b 5,06±0,0a 2,76±0,03c 52,6±0,11a 40,8±0,38c 1,08±0,05b K. 4,88±0,02c 6,35±0,1d 1,55±0,01a 58,9±0,12c 32,9±0,21a 1,11±0,04c Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, SM – sausosios medžiagos, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

PRB marinatai patikimai mažino TR kiekį (nuo 1,97 % P. acidilactici marinuotoje sprandinėje iki 34,86 % P. pentosaceus marinuotoje mentėje). Marinuotos kiaulienos technologinių savybių pokyčiams reikšmingos įtakos turėjo marinato gamybai naudotos PRB (p ≤ 0,0001) ir kiaulienos skerdenos dalis (p ≤ 0,0001) ir analizuotų veiksnių sąveika nustatyta reikšminga (p ≤ 0,0001).

Kiaulienos marinavimas PRB didino mėsos šviesumą (L*) ir rausvumą (a*) (atitinkamai, vidutiniškai 20,56 % ir 12,54 %), tačiau mažino gelsvumą (b*) (vidutiniškai 15,20 %). Spalvų pokytis mariuojant mėsą reikšmingai kito priklausomai nuo PRB (p ≤ 0,0001), kiaulienos skerdenos dalies (p ≤

231

0,0001) ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001) (3.2.1.5 lentelė).

3.2.1.5 lentelė. Skirtingų kiaulienos skerdenos dalių, marinuotų alternatyvioje terpėje pagausintomis PRB, spalvų koordinatės

Mėg L* a* b* L* a* b* Sprandinė Mentė Pa 52,5±0,22b 6,14±0,10b 15,97±0,16a 52,2±0,24b 13,73±0,14b 6,86±0,1a Pp 52,5±0,21b 5,79±0,05a 16,67±0,19b 54,1±0,25c 14,01±0,13b 8,24±0,1b Ls 53,0±0,22c 5,56±0,08a 15,68±0,18a 56,0±0,26d 13,63±0,12a 7,13±0,1a K. 48,3±0,19a 5,23±0,08a 16,91±0,18b 50,4±0,22a 13,44±0,14a 9,04±0,1c Kumpis M. longissimus dorsi Pa 63,1±0,28b 10,2±0,12c 10,4±0,13a 59,4±0,25a 4,72±0,9b 11,13±0,4a Pp 64,9±0,30c 9,56±0,08b 12,15±0,15c 62,9±0,28c 5,94±0,6c 10,75±0,6a Ls 66,0±0,26d 8,97±0,09a 12,20±0,15c 61,2±0,28b 4,66±0,5d 11,06±0,2a K. 62,47±0,3a 8,85±0,10a 13,97±0,16d 57,8±0,25a 3,54±0,5a 11,25±0,2b

Nugarinė Pa Pp Ls K. L* 55,6±0,20c 55,6±0,21c 55,0±0,28c 45,6±0,19a a* 7,00±0,10b 6,87±0,09a 6,69±0,08a 6,22±0,11a b* 11,09±0,15b 10,58±0,14a 10,43±0,13a 12,30±0,15c Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, L* − šviesumas, a* − rausvumas, b* − gelsvumas, p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Lyginant marinatų įtaką BA formavimuisi (3.2.1.2 a pav.), nustatyta, kad

marinatų gamybai naudotos PRB (p ≤ 0,0001) turėjo reikšmingos įtakos suminiam BA kiekiui (vidutiniškai, marinuojant atitinkamais marinatais, padidėjo P. acidilactici – 11,78 %, L. sakei – 36,03 % ir P. pentosaceus – 58,10 %).

Suminiam BA kiekiui mėsoje reikšmingos įtakos turėjo skerdenos dalis (p ≤ 0,0001): sprandinės, INR ir nugarinės marinuotuose mėginiuose suminis BA kiekis padidėjo nuo 8,85 % (P. acidilactici marinuotoje nugarinėje) iki 146,80 % (L. sakei marinuotame INR), mentės ir kumpio mėginiuose daugeliu atvejų sumažėjo: nuo 4,82 % (P. acidilactici marinuotoje mentėje) iki 38,33 % (P. acidilactici marinuotame kumpyje).

Jusliškai priimtinesni įvardinti marinuoti mėsos mėginiai (vidutiniškai nuo 2,16 % L. sakei iki 6,46 % P. pentosaceus marinuotoje kiaulienoje), lyginant su kontroliniais, o priimtiniausi – sprandinės mėginiai (priimtinesni vidutiniškai 19,26 %) (3.2.1.2 b pav.). Bendram mėginių priimtinumui reikšmingos įtakos turėjo PRB (p ≤ 0,0001), kiaulienos skerdenos dalis (p ≤ 0,0001) ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001).

232

a)

b) 3.2.1.2 pav. Alternatyviu marinatu apdorotų skirtingų kiaulienos

skerdenos dalių a) suminis BA kiekis ir b) bendrasis priimtinumas. Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, BA – biogeniniai aminai, Mld – M. longissimus dorsi, S – sprandinė, M – mentė, K – kumpis, N – nugarinė, p – skirtumo tarp

rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05

Alternatyvioje terpėje pagausintų P. acidilactici KTU05-7, P.

pentosaceus KTU05-8 ir L. sakei KTU05-6 įtaka marinuotos jautienos kokybės rodikliams. Skirtingų jautienos skerdenos dalių technologinėms savybėms reikšmingos įtakos turėjo PRB (p ≤ 0,0001), jautienos skerdenos dalis (p ≤ 0,0001) ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001) (3.2.1.6 lentelė).

Marinuotos jautienos nustatytas mažesnis pH (vidutiniškai nuo 11,33 % P. acidilactici iki 15,33% P. pentosaceus), didesni drėgmės nuostoliai (vidutiniškai nuo 14,45 % P. pentosaceus iki 16,36 % P. acidilactici) ir mažesnis TR kiekis (vidutiniškai nuo 3,99 % P. acidilactici iki 11,14 % P. pentosaceus), lyginant su nemarinuotais mėginiais. P. acidilactici, P. pentosaceus ir L. sakei marinatai mažino mėsos VR (atitinkamai, vidutiniškai 0,99 %, 0,30 % ir 1,21 %) ir didino VN (atitinkamai, vidutiniškai, 2,55 %, 1,52 % ir 0,81 %). Marinavimas padidino kumpio mėginių švelnumą (10,83 %) ir INR (1,95 %), tačiau priešingos tendencijos nustatytos marinuotų sprandinės (4,08 %), mentės (99,60 %) ir nugarinės (13,39 %) mėginių atitinkamų rodiklių.

Jautienos marinavimas daugeliu atvejų didino mėsos šviesumą (L*) (nuo 0,96 % iki 40,63 %, atitinkamai, P. acidilactici marinuotos mentės ir L. sakei marinuoto kumpio) ir gelsvumą (b*) (nuo 0,26 % iki 266,67 %, atitinkamai, P. pentosaceus marinuoto kumpio ir P. acidilactici marinuoto INR) bei mažino rausvumą (a*) (atitinkamai, vidutiniškai 16,58 % P. acidilactici, 33,35 % P. pentosaceus ir 24,32 % L. sakei), lyginant su kontroliniais mėginiais (3.2.1.7 lentelė).

0

50

100

150

S M K Mld N

mg/kg

Pa PpLs K

0

2

4

6

S M K Mld N

Bal

ai(m

in 0

; m

ax 6

)

Pa LsPp K

233

3.2.1.6 lentelė. Skirtingų jautienos skerdenos dalių, marinuotų alternatyvioje terpėje pagausintomis PRB, technologiniai rodikliai.

Mėg.

SM, %

pH Drėgmės nuostoliai,

%

Vandens rišlumas,

%

Virimo nuostoliai,

%

Švelnumas, kg/cm2

Sprandinė Pa 18,7±0,13a 5,03±0,01a 2,14±0,03b 61,4±0,16a 50,53±0,3c 3,82±0,01b Pp 19,5±0,11b 5,26±0,01b 1,86±0,02a 61,4±0,11a 49,03±0,3c 4,64±0,04c Ls 19,7±0,14b 5,11±0,03a 2,07±0,01b 62,9±0,14b 44,63±0,4a 2,68±0,02a K. 20,8±0,12c 5,96±0,02c 2,55±0,03c 62,9±0,15b 46,78±0,4b 3,57±0,03b

Mentė Pa 19,3±0,15b 5,20±0,02b 1,84±0,01a 62,8±0,12c 47,53±0,4b 4,43±0,03b Pp 18,8±0,11a 4,94±0,01a 2,12±0,03b 62,1±0,11b 46,67±0,4a 5,39±0,02c Ls 18,5±0,12a 5,05±0,03a 1,92±0,02a 61,4±0,14a 49,36±0,3d 5,21±0,01c K. 19,8±0,13c 5,72±0,05c 2,63±0,01c 62,2±0,12b 48,39±0,2c 2,51±0,02a

Kumpis Pa 22,0±0,11b 5,98±0,04c 2,17±0,03a 61,8±0,12a 49,71±0,3b 5,91±0,01b Pp 21,3±0,12b 5,38±0,03a 2,30±0,01b 62,3±0,13b 48,22±0,4a 4,05±0,02a Ls 20,2±0,14a 5,75±0,02b 1,94±0,01a 63,0±0,15c 51,71±0,3b 5,85±0,04c K. 24,3±0,13c 6,57±0,04a 2,41±0,02c 61,9±0,11a 53,56±0,3c 5,91±0,06b

M. longissimus dorsi Pa 20,5±0,09a 5,23±0,03b 1,76±0,01a 63,1±0,16c 45,79±0,3c 3,87±0,02c Pp 21,1±0,10b 5,04±0,02a 1,96±0,02a 62,4±0,16b 45,05±0,3b 2,59±0,01a Ls 20,1±0,12a 5,02±0,01a 1,88±0,04a 62,3±0,14b 43,33±0,2a 2,60±0,01a K. 21,3±0,11b 5,75±0,04c 2,04±0,03b 61,9±0,11a 42,74±0,4a 3,08±0,02b

Nugarinė Pa 18,8±0,11a 5,91±0,02b 1,98±0,02a 81,2±0.24b 45,56±0,3b 3,31±0,01a Pp 19,4±0,12b 5,44±0,04a 1,86±0,01a 80,3±0.19b 47,41±0,3c 5,45±0,03d Ls 18,6±0,09a 6,02±0,02b 2,14±0,01b 81,5±0.18c 46,48±0,2b 4,37±0,03c K. 22,5±0,13c 6,87±0,04c 2,25±0,05c 77,5±0.20a 42,75±0,2a 3,86±0,01b Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, SM – sausosios medžiagos, , p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Spalvų koordinačių pokyčiui reikšmingos įtakos turėjo marinatų gamybai naudotos PRB (p ≤ 0,0001), jautienos skerdenos dalis (p ≤ 0,0001) ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001). Analizuoti veiksniai (marinato gamybai naudotos PRB ir skerdenos dalis) turėjo reikšmingos įtakos BA formavimuisi marinuotoje jautienoje (p ≤ 0,0001) (3.2.1.7 lentelė). Lyginant skirtingų marinatų įtaką suminiam BA kiekiui nustatyta, kad PRB yra reikšmingas faktorius: mažausias suminis BA kiekis nustatytas P. acidilactici marinuotoje jautienoje (58,04 %), lyginant su P. pentosaceus (64,73 % didesnis, lyginant su kontroliniais mėginiais) ir L. sakei (66,12 % didesnis, lyginant su kontroliniais mėginiais).

234

3.2.1.7 lentelė. Skirtingų jautienos skerdenos dalių, marinuotų alternatyvioje terpėje pagausintomis PRB, saugos ir kokybės rodikliai

Mėg.

Tarpraum. riebalai,

% BP

Spalvų koordinatės BA,

mg/kg L* a* b* Sprandinė

Pa 3,37±0,01a 5,4±0,05c 42,06±0,20c 10,46±0,20b 4,45±0,03b 88,83 Pp 3,70±0,04b 5,6±0,06d 41,02±0,20b 7,26±0,19a 2,84±0,02a 99,05 Ls 3,82±0,03b 5,1±0,05c 36,86±0,16a 8,31±0,20a 4,71±0,05b 105,37 K. 3,37±0,03a 4,5±0,03a 38,16±0,17a 18,90±0,29d 4,85±0,05c 61,57

Mentė Pa 1,39±0,02b 4,9±0,04a 45,04±0,20b 13,99±0,21a 8,18±0,05c 96,63 Pp 1,23±0,02a 4,8±0,04a 48,56±0,18c 16,15±0,19b 7,52±0,03b 68,09 Ls 1,37±0,01b 5,3±0,06b 43,42±0,19a 13,91±0,31a 7,36±0,10b 88,73 K. 1,59±0,01c 4,9±0,04a 44,61±0,20b 19,48±0,32c 6,47±0,09a 101,53

Kumpis Pa 1,60±0,03b 5,2±0,04a 31,05±0,15a 12,84±0,19a 4,96±0,04b 38,38 Pp 1,50±0,02a 4,9±0,04a 34,32±0,20b 13,17±0,18b 3,85±0,03a 53,79 Ls 1,77±0,01c 5,5±0,05b 43,06±0,20d 12,57±0,26a 6,08±0,09c 148,73 K. 1,48±0,04a 5,1±0,05a 30,69±0,15a 15,79±0,31c 3,84±0,05a 62,23

M. longissimus dorsi Pa 1,32±0,01c 5,5±0,06a 40,24±0,20c 15,19±0,26b 8,25±0,05d 98,39 Pp 0,95±0,03a 5,4±0,06a 38,23±0,19b 11,69±0,15a 7,04±0,08c 162,52 Ls 0,83±0,02a 5,8±0,07b 36,92±0,16a 15,87±0,26c 5,90±0,08b 105,37 K. 1,13±0,02b 5,5±0,07a 38,03±0,16b 15,16±0,26b 2,25±0,03a 65,85

Nugarinė Pa 1.51±0,02a 5,1±0,05b 30,79±0,15a 15,31±0,26c 4,81±0,03c 93,80 Pp 1.60±0,04a 4,8±0,04a 41,02±0,23d 7,26±0,12a 2,84±0,02a 102,51 Ls 1.82±0,01b 5,5±0,05c 33,62±0,16b 11,12±0,28b 4,17±0,06b 114,58 K. 2.23±0,03c 5,1±0,05b 29,89±0,14a 14,12±0,26b 2,93±0,01a 86,17 Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, L* − šviesumas, a* − rausvumas, b* − gelsvumas, BA – biogeniniai aminai, BP – bendrasis priimtinumas, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Marinavimas turėjo teigiamos įtakos mėginių bendram priimtinumui

(3.2.1.7 lentelė) ir daugeliu atvejų jį didino. Bendras priimtinumas patikimai priklausė nuo marinato gamybai naudotų PRB (p ≤ 0,0001), jautienos skerdenos dalies (p ≤ 0,0001) ir šių faktorių sąveika buvo reikšminga (p ≤ 0,0001).

Alternatyvioje terpėje pagausintų P. acidilactici KTU05-7 įtaka

tradiciškai ir ekologiškai pagamintos marinuotos vištienos kokybės rodikliams. Vištienos skerdenėlės dalis (p ≤ 0,0001) ir vištienos gamybos

235

būdas (p ≤ 0,0001) turėjo reikšmingos įtakos marinuotų gaminių technologiniams rodikliams (3.2.1.3 pav.).

a) b)

c) d) 3.2.1.3 pav. Alternatyvioje terpėje pagausintomis PRB apdorotos kiaulienos

a) pH ir drėgmės nuostoliai, b) vandens rišlumas ir virimo nuostoliai, c) švelnumas ir tarpraumeninių riebalų kiekis, d) suminis biogeninių aminų

kiekis. Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, L* − šviesumas, a* − rausvumas, b* − gelsvumas,K – kontrolė, TR – tarpraumeninių riebalų kiekis, VN –

virimo nuostoliai, VR – vandens rišlumas, DN – drėgmės nuostoliai, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05

Marinavimas P. acidilactici sumažino ekologiškai ir tradiciškai

pagamintos vištienos pH (atitinkamai, vidutiniškai 11,75 % ir 11,76 %) ir padidino drėgmės nuostolius (nuo 2,20 % iki 30,23 %, atitinkamai, tradiciškai pagamintose blauzdelėse, šlaunelėse ir ekologiškai pagamintose šlaunelėse). Lyginant su nemarinuotais mėginiais, marinuotų mėginių nustatyti didesni VR ir VN (nuo 0,20 % iki 0,34 % ir nuo 1,96 % iki 3,09 %, atitinkamai, ekologiškai ir tradiciškai pagamintos vištienos). P. acidilactici marinatas didino vištienos švelnumą (nuo 8,08 % iki 26,12 %, atitinkamai,

4,5

5

5,5

6

6,5

00,20,40,60,8

11,2

KPaKPa KPaKPa KPaKPa

Ekol.Trad.Ekol.Trad.Ekol.Trad.

Krūtinėlė Blauzdelė Šlaunelė

pHDN, %

Drėgmės nuostoliai pH

61,56262,56363,5

05

101520

KPaKPa KPaKPa KPaKPa

Ekol.Trad.Ekol.Trad.Ekol.Trad.

Krūtinėlė Blauzdelė Šlaunelė

VR, %VN, %

Virimo nuostoliai Vandens rišlumas

00,511,522,53

00,20,40,60,8

1

KPaKPa KPaKPa KPaKPa

Ekol.Trad.Ekol.Trad.Ekol.Trad.

Krūtinėlė Blauzdelė Šlaunelė

Šveln, kg/cm2

TR, %

TR Švelnumas

020406080

100

K Pa KPa K Pa KPa K Pa KPa

Ekol. Trad. Ekol. Trad. Ekol.Trad.

Krūtinėlė Blauzdelė Šlaunelė

mg/kg

L* a* b*

236

ekologiškai ir tradiciškai pagamintų blauzdelių) ir TR kiekį (vidutiniškai 7,26 % ekologiškai ir 6,30 % tradiciškai pagamintos vištienos).

Analizuoti veiksniai (vištienos gamybos būdas (p ≤ 0,0001) ir skerdenėlės dalis (p ≤ 0,0001) turėjo reikšmingos įtakos marinuotos, tradiciškai ir ekologiškai pagamintos vištienos šviesumo (L*) (atitinkamai, 3,27 % ir 2,72 %) bei rausvumo (a*) didėjimui ir gelsvumo (b*) koordinačių mažėjimui (atitinkamai, 8,99 % ir 3,35 % bei 12,55 % ir 6,75 %).

Didesnis suminis BA kiekis nustatytas tradiciškai pagamintoje vištienoje (vidutiniškai, 157,36 % krūtinėlėje, 160,0 % blauzdelėje ir 110,31 % šlaunelėje), lyginant su ekologiškai pagaminta, bei priklausė nuo vištienos gamybos būdo (p ≤ 0,0001) ir skerdenėlės dalies (p ≤ 0,0001) (3.2.1.3 pav.). Marinavimas P. acidilactici sumažino BA kiekį ekologiškai pagamintoje vištienoje (nuo 31,63 % blauzdelėje iki 42,66 % šlaunelėje), o tradiciškai pagamintoje padidino (nuo 110,25 % šlaunelėje iki 218,94 % blauzdelėje).

3.2.2. Policiklinių aromatinių angliavandenilių mažinimas šaltai

rūkytose dešrose, taikant kietafazę fermentaciją antimikrobinėmis savybėmis pasižyminčiomis PRB

PRB antimikrobinis aktyvumas. Didžiausias PRB antimikrobinis

aktyvumas nustatytas prieš Y. pseudotuberculosis (vidutiniškai 17,33 mm), o mažiausias – prieš B. cereus (vidutiniškai 8,07 mm) (3.2.2.1 lentelė). P. acidilactici, P. pentosaceus ir L. sakei antimikrobinis aktyvumas patikimai priklausė nuo PRB ir patogeninių mikroorganizmų rūšies (prieš P. aeruginosa p ≤ 0,0001, S. aureus p ≤ 0,002, L. monocytogenes p ≤ 0,006, E. coli p ≤ 0,003, S. typhimurium p ≤ 0,0001, Y. enterolitica p ≤ 0,001, Y. pseudotuberculosis p ≤ 0,0001).

3.2.2.1 lentelė. PRB antimikrobinis aktyvumas prieš pasirinktus patogenus

Mikroorganizmas Inhibicinė zona/mm* P. acidilactici 7 P. pentosaceus 9 L. sakei

P. aeruginosa 11,0±0,5b 15,0±0,6d 13,0±0,4c S. aureus 12,0±0,6c 13,0±0,5c 10,0±0,6b L. monocytogenes 11,0±0,3b 9,6±0,5a 11,0±0,3b E. coli 11,0±0,3b 11,0±0,6b 8,6± 0,7a S. typhimurium 13,0±0,5b 12,5±0,3b 9,5±0,4a Y. enterolitica 12,5±0,3c 13,0±0,5c 10,9±0,3b Y. pseudotuberculosis 15,0±0,4d 18,0±0,4e 19,0±0,5e B. cereus 8,0±0,5a 8.0±0,5a 8,2±0,4a * Šulinėlių diametras buvo 6 mm; a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

237

PRB įtaka šaltai rūkytų dešrų saugos rodikliams. Dešrų paviršiaus apdorojimas alternatyvioje terpėje pagausintomis PRB sumažino suminį PAA kiekį vidinėje ir išorinėje dešrų dalyse (vidutiniškai, atitinkamai, prieš rūkymą 25,50 % ir 28,29 %, po rūkymo – 17,69 % ir 20,63 %), lyginant su neapdorotais mėginiais (3.2.2.2 lentelė). Naudotos PRB turėjo reikšmingos įtakos benz[a]antraceno (BaA) (p = 0,0001), benzo[a]pireno (BaP) (p = 0,001), benzo-[b]fluoranteno (BbF) (p = 0,0001) ir chriseno (Chr) (p = 0,0001) susidarymui šaltai rūkytose dešrose. Nustatyti patikimi BaA (p = 0,0001), BaP (p = 0,0001), BbF (p = 0,035) ir Chr (p = 0,0001) kiekybiniai pokyčiai skirtingose dešrų dalyse (išorinėje ir vidinėje). Analizuoti veiksniai (PRB rūšis p ≤ 0,0001, produkto dalis (išorinė ar vidinė) p ≤ 0,0001, apdorojimo būdas (prieš rūkymą ar po jo) p ≤ 0,0001 ir jų tarpusavo sąveika (p ≤ 0,0001) buvo reikšminga BA formavimuisi šaltai rūkytose dešrose (3.2.2.2 lentelė). Lyginant dešrų apdorojimo procesų ir PRB įtaką BA susidarymui, nustatytos skirtingos tendencijos: apdorojimas prieš rūkymą L. sakei ir P. acidilactici sumažino (atitinkamai, 12,61 % ir 45,32 %), o P. pentosaceus – padidino (3,25 %), tuo tarpu apdorojimas PRB po rūkymo visais atvejais padidino (atitinkamai, 37,67 %, 48,55 % ir 40,99 %) suminį BA kiekį išorinėje dešrų dalyje, lyginant su vidine. Šaltai rūkytų dešrų apdorojimas prieš rūkymą ir po jo didino suminį BA kiekį (atitinkamai, vidutiniškai 45,41 % ir 42,61 %).

3.2.2.2 lentelė. PAA ir suminis BA kiekis šaltai rūkytose dešrose.

PAA L. sakei P. acidilactici 7 P. pentosaceus 9 Kontrolė

PR PoR PR PoR PR PoR NA AV PR

Vidinė dalis BaA, ng/g

0,068±0,005b

0,054±0,003a

0,061±0,004a

0,063±0,010b

0,065±0,007b

0,062±0,005b

0,072±0,006c

0,069±0,005c

Chr, ng/g

0,149±0,007b

0,168±0,009c

0,204±0,006e

0,144±0,011b

0,168±0,005c

0,104±0,010a

0,197±0,013d

0,169±0,014c

BbF, ng/g

0,021±0,009a

0,033±0,005b

0,030±0,004b

0,039±0,002c

0,031±0,001b

0,051±0,004d

0,033±0,003b

0,021±0,002a

BaP, ng/g

0,039±0,004b

0,030±0,002a

0,036±0,007b

0,034±0,003a

0,034±0,005a

0,042±0,003b

0,081±0,007d

0,079±0,004d

∑PAA 0,227a 0,285c 0,331c 0,280c 0,298d 0,259b 0,383f 0,328d ∑BA 49,50b 41,63a 93,26e 53,76d 57,77c 46,43b 40,93a 40,39a

Išorinė dalis BaA, ng/g

0,072± 0,003b

0,065±0,004a

0,069±0,005a

0,071±0,011b

0,069±0,004a

0,072±0,003b

0,086±0,009c

0,074±0,003b

Chr, ng/g

0,167±0,011b

0,184±0,015d

0,212±0,014e

0,163±0,014b

0,184±0,003d

0,124±0,012a

0,212±0,011e

0,182±0,013d

BbF, ng/g

0,040±0,007a

0,051±0,007b

0,041±0,005a

0,062±0,008c

0,054±0,007b

0,073±0,005d

0,037±0,005a

0,048±0,003b

238

3.2.2.2 lentelės tęsinys PAA L. sakei P. acidilactici 7 P. pentosaceus 9 Kontrolė

PR PoR PR PoR PR PoR NA AV PR

BaP, ng/g

0,062±0,005b

0,054±0,006a

0,051±0,009a

0,072±0,007b

0,068±0,002b

0,059±0,004a

0,106±0,010d

0,098±0,007d

∑PAA 0,341b 0,354b 0,373c 0,368c 0,375c 0,328a 0,441e 0,402d ∑BA 43,26a 57,31d 50.99b 79,86f 59,65c 65,46e 40,23a 40,00a PAA – policikliniai aromatiniai angliavandeniliai, BaA – benz[a]anthracenas, BbF – benzo-[b]fluorantenas, BaP – benzo[a]pirenas, Chr – chrisenas, PR – š/r dešros apdorotos PRB prieš rūkymą, PoR – š/r dešros apdorotos PRB po rūkymo, BA – biogeniniai aminai, AV – apdorota vandeniu, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės eilutėse pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

Apdorojimui naudotos PRB turėjo reikšmingos įtakos šaltai rūkytų dešrų bendram priimtinumui (p ≤ 0,0001) (3.2.2.1 pav.).

3.2.2.1 pav. Bendras priimtinumas ir emocijos, užfiksuotosas FaceReader programine įranga. Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, PR – šaltai rūkytos dešros apdorotos PRB prieš rūkymą; PoR – šaltai rūkytos dešros apdorotos PRB po rūkymo, K – kontrolė, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05

Paviršiaus apdorojimas po rūkymo mažino bendrą priimtinumą

(vidutiniškai, L. sakei – 12,50 %, P. acidilactici – 2,08 % ir P. pentosaceus – 6,25 %), o apdorojimas P. pentosaceus prieš rūkymą – didino 4,17 %, lyginant su neapdorotais mėginiais. Įvertinus vertintojų veido emocines raiškas, kaip emocinį atsaką į dešrų juslinį priimtinumą (FaceReader programine įranga), nustatyta, kad pastarųjų sąsajos su bendru priimtinumu, įvertintu hedoninėje intensyvumo skalėje, yra statistiškai reikšmingos (p = 0.0001) (bendras priimtinumas ir emocinė raiška „laimingas“, bendras priimtinumas ir emocinė raiška „liūdnas“, bendras priimtinumas ir emocinė raiška „piktas“).

0123456

00,10,20,30,40,50,6

PR PoR PR PoR PR PoR

Pa Ls Pp K

Taš

kai

Em

ocij

a

Laimingas Liūdnas Piktas Taškai

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3.2.3. Augalinių bioproduktų įtaka maltos mėsos produktų kokybei

Augalinių bioproduktų įtaka MMP mikrobiologiniam stabilumui. Fermentacijos metodas ir fermentacijai naudotos PRB turėjo reikšmingos įtakos (p ≤ 0,05) nepageidaujamų mikroorganizmų kiekiui (3.2.3.1 lentelė).

3.2.3.1 lentelė. MMP su 3 % Sm priedu mikrobiologiniai rodikliai. MMP mėginiai

Laikymo laikas, val. 0 24 72 120

Bendras mezofilinių bakterijų skaičius, log10 KSV/g Kontrolė 5,21±0,32a 6,09±0,62b 6,25±0,72c 6,77±0,43d Ls TF 4,81±0,82a 5,14±0,64b 5,33±0,54b 6,01±0,32c Ls KF 4,63±0,62a 5,13±0,85c 5,27±0,22c 5,69±0,28d Pa TF 4,65±0,41a 5,21±0,27c 5,42±0,27c 5,59±0,27d Pa KF 3,62±0,21a 4,05±0,37c 4,18±0,25c 4,43±0,29d Pp TF 4,91±0,61a 5,45±0,30c 5,69±0,31c 6,04±0,37d Pp KF 4,59±0,42a 5,29±0,54c 5,38±0,45c 5,64±0,34d

Koliforminių bakterijų skaičius, log10 KSV/g Kontrolė 4,02±0,35a 4,30±0,42b 5,03±0,24d 6,83±0,58e Ls TF 3,81±0,41a 4,00±0,33a 4,67±0,58c 4,95±0,37c Ls KF 3,61±0,49a 3,79±0,91a 4,44±0,42c 4,60±0,42c Pa TF 3,23±0,62a 3,39±0,87a 3,95±0,31b 4,20±0,30c Pa KF 2,94±0,65a 3,19±0,59a 3,61±0,72c 3,82±0,44c Pp TF 3,71±0,59a 3,90±0,58a 4,62±0,70c 4,82±0,24c Pp KF 3,59±0,29a 3,77±0,33a 4,31±0,85b 4,67±0,26c

Mielių ir pelėsinių grybų skaičius, log10 KSV/g

Kontrolė 2,92±0,44a 3,30±0,50a 3,92±0,45c 3,98±0,23c Ls TF 1,81±0,38a 2,14±0,24a 2,31±0,32b 2,36±0,14b Ls KF 1,61±0,18a 1,82±0,89a 2,02±0,29b 2,07±0,35b Pa TF 1,23±0,33a

1,49±0,56a 2,15±0,30c 2,19±0,25c Pa KF 1,54±0,24a

1,64±0,83a 1,84±0,24b 1,89±0,60b Pp TF 2,11±0,49a 2,18±0,50a 2,43±0,27b 2,46±0,24b Pp KF 1,88±0,25a 2,10±0,33b 2,12±0,21b 2,13±0,52b Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, Kontrolė – MMP be Sm, Sm – S. montana, KF – kietafazė, TF – tradicinė fermentacija, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės eilutėse pažymėti skirtingomis raidėmis, tarpusavyje patikimai skiriasi (p < 0,05)

Sm bioproduktai, priklausomai nuo PRB naudotos jų gamybai, patikimai

mažino mezofilinių (p ≤ 0,0001) ir koliforminių (p ≤ 0,0001) bakterijų bei mielių ir pelėsinių grybų (p ≤ 0,003) kiekį MMP.

Didžiausiu antimikrobiniu aktyvumu pasižymėjo P. acidilactici fermentuoti Sm bioproduktai (vidutiniškai mezofilinių bakterijų skaičių MMP sumažino 23,45 %, koliforminių 28,22 %, mielių ir pelėsinių grybų 50,75 %), lyginant su kontroliniais MMP. KF fermentuoti Sm bioproduktai mažino mezofilinių ir koliforminių bakterijų kiekį (atitinkamai, 22,01 % ir

240

8,13 %) MMP, tačiau didino mielių ir pelėsinių grybų kiekį 1,79 %, lyginant su Sm TF bioproduktais.

Sm bioproduktų įtaka AJ kiekiui MMP. Analizuoti veiksniai (fermentacijai naudotos PRB ir fermentacijos sąlygos) turėjo reikšmingos įtakos AJ kiekiui MMP, praturtintuose Sm (p ≤ 0,005). Sm bioproduktai padidino suminį AJ kiekį nuo 15,46 iki 23,66 kartų (atitinkamai, MMP, pagamintuose su L. sakei TF ir P. pentosaceus TF apdorotais Sm), lyginant su kontroliniais MMP. KF apdoroti Sm bioproduktai didino AJ kiekį MMP (0,84 %), lyginant su TF. Sm bioproduktai padidino ρ-kimeno, γ-terpineno ir karvakrolio kiekį MMP (atitinkamai, vidutiniškai 36,6; 7,1 ir 5,9 karto), lyginant su kontroliniais MMP.

Sm ir Sh bioproduktų įtaka MMP technologiniams ir saugos rodikliams. MMP, praturtintų Sm ir Sh bioproduktais, bendram priimtinumui reikšmingos įtakos turėjo augalų fermentacijai naudotos PRB (p ≤ 0,0001), augalo rūšis (p ≤ 0,0001), bioprodukto kiekis (p ≤ 0,0001) ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001). MMP pagaminti su 3 % Sm ir Sh įvertinti priimtiniausiais, o MMP su 7 % bioproduktų priedu įvertinti nepriimtinais ir toliau analizuojami nebuvo. Priimtiniausiais įvertinti MMP su P. pentosaceus fermentuotais TF ir KF Sm ir Sh bioproduktais (atitinkamai, 5,5 ir 6,0 ir 5,8 ir 5,8 taškai).

Fermentacijos technologija turėjo reikšmingos įtakos (p ≤ 0,05) MMP kokybės rodikliams (3.2.3.2 lentelė). KF apdoroti Sm bioproduktai didino MMP pH, o Sh – mažino (atitinkamai, vidutiniškai 0,22 % ir 0,50 %, lyginant su TF fermentuotais bioproduktais ir, atitinkamai, 0,16 % ir 0,80 %, lyginant su kontrolinias mėginiais). MMP drėgmės nuostoliams reikšmingos įtakos turėjo Sm ir Sh fermentacijos technologija (p ≤ 0,0001 ir p ≤ 0,004) ir mažesni drėgmės nuostoliai nustatyti MMP, pagamintų su KF fermentuotais Sm ir Sh bioproduktais (atitinkamai, vidutiniškai 50,52 % ir 14,81 %), lyginant su MMP, pagamintais su TF apdorotais bioproduktais.

Augaliniai bioproduktai daugeliu atvejų didino MMP VR, lyginant su kontroliniais mėginiais (nuo 2,78 % MMP su TF apdorotais P. acidilactici Sh iki 20,47 % MMP su KF apdorotais P. acidilactici Sm). Didesni VR nustatyti MMP su KF apdorotais Sm ir Sh (atitinkamai, vidutiniškai, 8,43 % ir 8,89 %), lyginant su MMP, pagamintais su TF apdorotais bioproduktais. Sm ir Sh didino MMP VN (vidutiniškai 1,16 (Sm TF) ir 1,09 (Sm KF) bei 1,11 (Sh TF) ir 0,92 (Sm KF) kartų) ir švelnumą (atitinkamai, vidutiniškai 39,29 % ir 36,90 %), lyginant su kontroliniais MMP. Augaliniais bioproduktais praturtintų MMP VR ir VN patikimai priklausė nuo augalinių bioproduktų fermentacijos technologijos (p ≤ 0,0001) bei jų fermentacijai naudotų PRB (atitinkamai, p ≤ 0,001 ir p ≤ 0,0001) ir faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001).

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3.2.3.2 lentelė. MMP, pagamintų su Sm ir Sh bioproduktais, technologiniai rodikliai.

Augaliniai bioproduktai patikimai mažino MMP (L*) ir (b*) spalvų

koordinates (atitinkamai, vidutiniškai Sm 16,89 % ir Sh 20,06 % bei Sm 14,44 % ir Sh 9,49 %) (3.2.3.3 lentelė). Augalinių bioproduktų priedas didino MMP rausvumą (a*) nuo 22,38 % iki 68,86 % (atitinkamai, MMP pagamintų su KF ir TF P. acidilactici apdorotais Sh MMP, lyginant su kontroliniais mėginiais). TF apdoroti Sm ir Sh didino MMP rausvumą (atitinkamai 11,0 % ir 20,63 %), lyginant su MMP, pagamintais su KF augaliniais bioproduktais.

Analizuoti veiksnai: bioproduktų fermentacijos technologija (p ≤ 0,0001), fermentacijai naudotos PRB (p ≤ 0,0001) turėjo reikšmingos įtakos BA formavimuisi MMP ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001) (3.2.3.3 lentelė). Didesnis suminis BA kiekis nustatytas MMP, pagamintuose su Sh bioproduktais (nuo 125,59 % su L. sakei TF iki 223,35 % su P. acidilactici TF apdorotais augaliniais produktais). Mažesnis suminis BA kiekis nustatytas MMP, pagamintuose su Sm bioproduktais (vidutiniškai, 0,89 % TF ir 34,87 % KF), lyginant su kontroliniais MMP. Lyginant augalinių bioproduktų fermentacijos procesų ir naudotų PRB įtaką BA kiekiui MMP, nustatytos skirtingos tendencijos: KF apdoroti Sm

MMP SM,% pH Drėgmės nuostoliai,%

Vandens rišlumas,%

Virimo nuostoliai,%

S. montana (3 %)

Ls TF 29,67±0,03c 6,16±0,01c 2,23±0,26b 65,97±0,23b 26,36±0,18b

KF 30,60±0,03d 6,08±0,02b 1,20±0,17a 67,75±0,41b 25,32±0,11a

Pa TF 29,56±0,04c 6,08±0,01b 3,06±0,41c 63,45±0,46a 26,57±0,09b KF 26,94±0,02a 6,04±0,03a 1,21±0,19a 72,44±0,79e 25,25±0,17a

Pp TF 27,84±0,02b 6,02±0,04a 2,16±0,24b 64,01±0,32a 32,28±0,26d KF 30,18±0,03c 6,10±0,02b 1,19±0,11a 69,41±0,20c 29,32±0,19c

S. hortensis (3 %)

Ls TF 28,67±0,03b 6,01±0,03a 2,19±0,26b 59,10±0,61a 22,60±0,30a KF 30,55±0,03c 6,05±0,04b 1,86±0,18a 65,99±0,66b 25,88±0,21b

Pa TF 29,84±0,04c 6,00±0,03a 2,11±0,20b 61,80±0,61a 30,42±0,33c KF 31,40±0,04d 6,03±0,02b 1,74±0,19a 65,23±0,68b 19,98±0,23a

Pp TF 29,20±0,03b 6,01±0,02a 2,03±0,20b 65,11±0,67b 28,45±0,25b KF 26,59±0,02a 6,03±0,04b 1,79±0,15a 71,28±0,70d 21,25±0,18a

K 37,06±0,04e 6,07±0,03d 1,96±0,15c 60,13±0,45b 24,46±0,22c Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, SM – sausosios medžiagos, Sm – Satureja montana, Sh – Satureja hortensis, KF – kietafazė fermentacija, TF – tradicinė fermentacija, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

242

mažino, o Sh didino suminį BA kiekį MMP (atitinkamai, vidutiniškai 29,97 % ir 0,19 %), o P. acidilactici fermentuoti bioproduktai efektyviausiai mažino suminį BA kiekį MMP (atitinkamai, 53,94 % ir 13,38 %).

3.2.3.3 lentelė. MMP, pagamintų su Sm ir Sh bioproduktais, technologiniai rodikliai ir suminis biogeninių aminų kiekis.

MMP be glitimo (manų kruopų pakeitimo MMP receptūroje žirnių

skaidulomis) kokybės rodikliai. Patikimi skirtumai (p ≤ 0,05) nustatyti tarp MMP, pagamintų su manų kruopomis ir žirnių skaidulomis kokybės rodiklių (3.2.3.1 pav.).

Žirnių skaidulos didino MMP drėgmės nuostolius (4,3 %), pH (0,91 %), ir švelnumą (1,68 kartus) bei mažino VN (2,90 %) ir VR (8,9 %), lyginant su MMP, pagamintais su manų kruopomis.

Manų kruopų priedas didino MMP šviesumo (L*) (7,2 %), rausvumo (a*) (5,1 %) ir gelsvumo (b*) (0,5 %) koordinates, lyginant su MMP su žirnių skaidulomis.

Didesnis suminis BA kiekis nustatytas MMP su žirnių skaidulomis (23,9 %), lyginant su MMP su manų kruopų priedu.

MMP Švelnumas, kg/cm2

Spalvų koordinatės BA, mg/kg L* a* b*

S. montana (3 %) Ls TF 0,27±0,02b 61,01±1,02c 6,22±0,74d 13,62±0,54c 13,65b

KF 0,29±0.02c 58,80±2,31a 5,29±0,69b 12,54±0,26a 11,44a

Pa TF 0,32±0.03c 59,61±1,89b 5,39±0,88b 13,59±0,27b 44,31f KF 0,20±0.02a 58,76±1,26a 5,09±0,53a 13,17±0,19b 20,41d Pp TF 0,20±0.01a 59,36±0,97b 6,21±0,42d 14,57±0,16c 43,01d KF 0,25±0.02b 58,50±1,07a 5,67±0,29c 13,23±0,22b 34,50e

S. hortensis (3 %) Ls TF 0,25±0.01a 60,27±1,23b 5,60±0,61c 14,11±0,17b 76,61c KF 0,27±0.02a 56,78±1,51a 5,16±0,54b 13,80±0,13a 91,19c Pa TF 0,31±0.03b 59,47±1,46b 5,94±0,60c 14,19±0,10b 109,81d KF 0,26±0.02a 56,84±1,32a 5,03±0,48a 13,58±0,14a 95,12d Pp TF 0,24±0.02a 59,34±1,41b 6,94±0,72d 14,39±0,16b 87,58c KF 0,26±0.03a 55,71±1,33a 5,13±0,55b 13,81±0,14a 83,13c K 0,42±0.03d 70,61±2,17e 4,11±0,43a 15,17±0.17c 33,96e Pastaba: Ls – L. sakei, Pa – P. acidilactici, Pp – P. pentosaceus, L* − šviesumas, a* - rausvumas, b* - gelsvumas, BA – biogeniniai aminai, Sm – Satureja montana, Sh – Satureja hortensis, KF – kietafazė fermentacija, TF – tradicinė fermentacija, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05, p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės stulpeliuose pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p < 0,05)

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3.2.3.1 pav. MMP, pagamintų su žirnių skaidulomis ir manų kruopomis,

kokybės rodikliai ir suminis BA kiekis. Paaiškinimai: SM- sausosios medžiagos, DN – drėgmės nuostoliai, VR – vandens rišlumas, VN – virimo nuostoliai, K – kontrolė, L* − šviesumas, a* - rausvumas, b* - gelsvumas, BA – biogeniniai aminai, p – skirtumo tarp

rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05. 3.2.4. Augalinių bioproduktų įtaka varškės sūrių kokybės ir saugos

rodikliams Sm ir Rc bioproduktai stabdė mielių augimą VS paviršiuje ir prailgino

vartoti tinkamumo terminą (atitinkamai, 5 ir 3 paromis), lyginant su kontroliniais mėginiais (3.2.4.1 lentelė) bei turėjo reikšmingos įtakos VS saugos ir kokybės rodikliams, kurie reikšmingai priklausė nuo augalinių bioproduktų fermentacijos metodo (p ≤ 0,0001), fermentacijai naudotų PRB (p ≤ 0,0001), augalo rūšies (p ≤ 0,0001) ir šių faktorių tarpusavio sąveika buvo reikšminga (p ≤ 0,0001).

3.2.4.1 lentelė. Sm ir Rc džiovintų augalų ir jų bioproduktų įtaka mielių augimui VS laikymo metu

Laikymo trukmė, dienomis

L. sakei ir augalų bioproduktai Džiovinti augalai Kontrolė Sm Rc Sm Rc

4 - - - - - 5 - - - + + 7 - - + + + 9 - + + + +

11 + + + + ++ Sm – Satureja montana, Rc – Rhaponticus carthamoides, VS – varškės sūris, (-) nėra mielių kolonijų, (+) pirmieji mielių pėdsakai; (++) didelis kiekis mielių kolonijų; p – skirtumo tarp rezultatų reikšmių patikimumas p – patikimas, kai ≤ 0,05

AJ kiekis VS, praturtintuose 3 % Sm ir Rc bioproduktų, patikimai

priklausė nuo augalo rūšies (p ≤ 0,05). Sm ir Rc bioproduktai padidino suminį AJ kiekį (atitinkamai, vidutiniškai 7,03 ir 8,96 kartus), lyginant su

0

20

40

60

SM, % pH DN, % VR, % VN, % K,kg/cm2

L* a* b* BA

MMP su žirnių skaidulomis

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kontroliniais VS mėginiais, o VS su TF fermentuotais bioproduktais pasižymėjo didesniu AJ kiekiu (atitinkamai, vidutiniškai, 68,54 % ir 13,10 %), lyginant su VS, pagamintais su KF apdorotais bioproduktais. Sm bioproduktai padidino karvakrolio ir timolio (atitinkamai, vidutiniškai 21,88 ir 12.68 kartus), o Rc – timolio ir ρ-kimeno kiekį (atitinkamai, vidutiniškai 25,24 ir 18,86 kartus). Karvakrolio VS, pagamintuose su Rc, nenustatyta.

Analizuoti veiksniai turėjo reikšmingos įtakos suminiam BA kiekiui VS, praturtintuose augaliniais bioproduktais (augalų fermentacijos būdas p ≤ 0,0001, fermentacijai naudotos PRB p ≤ 0,0001, augalo rūšis p ≤ 0,0001 ir jų sąveika p ≤ 0,0001 buvo reikšminga) (3.2.4.2 lentelė).

3.2.4.2 lentelė. VS su Sm ir Rc bioproduktais saugos ir kokybės rodikliai.

Mėg.

Su fermentuotais augalų bioproduktais K

Su nef. Sm ir

Rc L. sakei P. acidilactici 7 P. pentosaceus 8

TF KF TF KF TF KF VS pagaminti su Sm

pH 24

4,68 ±0.05c

4,38 ±0.02a

4,54 ±0,04b

4,41 ±0,03a

4,76 ±0,05d

4,51 ±0,04b

4,79 ±0,06d

4,69 ±0,05c

pH 48

4,21 ±0.03a

4,13 ±0.04a

4,38 ±0,03b

4,08 ±0,02a

4,52 ±0,04c

4,22 ±0,03a

4,53 ±0,05c

4,38 ±0,04b

BTR 0,20 ±0.01b

0,10 ±0.01a

0,20 ±0,01b

0,10 ±0,03a

0,10 ±0,01a

0,10 ±0,02a

0,10 ±0,01a

0,10 ±0,01a

L(+) 5,61 ±0.07c

7,44 ±0.09f

5,36 ±0,07b

6,74 ±0,01d

5,14 ±0,07b

5,95 ±0,08c

4,68 ±0,05a

5,24 ±0,06b

D(-) 1,39 ±0.03a

1,17 ±0.02a

2,35 ±0,04c

1,94 ±0,03b

3,28 ±0,04d

2,47 ±0,03c

1,78 ±0,02b

2,14 ±0,03b

BA 145,37f 28,07b 130,3f 67,31c 135,42f 111,61e 4,77a 30,18b VS pagaminti su Rc

pH 24

4,38 ±0,03b

4,21 ±0,02a

4,42 ±0,04b

4,15 ±0,02a

4,44 ±0,04b

4,40 ±0,04b

4,54 ±0,05c

4,39 ±0,03b

pH 48

4,18 ±0,02b

4,09 ±0,02a

4,32 ±0,03c

4,11 ±0,02a

4,34 ±0,03c

4,11 ±0,02a

4,39 ±0,03c

4,25 ±0,03b

BTR

0,19 ±0,02b

0,11 ±0,01a

0,21 ±0,02b

0,12 ±0,01a

0,10 ±0,01a

0,10 ±0,01a

0,11 ±0,01a

0,12 ±0,01a

L(+) 2,18 ±0,06a

5,44 ±0,07c

2,94 ±0,09b

5,44 ±0,07c

5,94 ±0,03d

4,94 ±0,03c

3,17 ±0,03b

2,00 ±0,04a

D(-) 3,47 ±0,03c

1,30 ±0,02a

2,76 ±0,04b

2,45 ±0,03b

3,45 ±0,04c

2,68 ±0,04b

2,34 ±0,03b

2,79 ±0,04b

BA 171,27f 3,69a 100,28d 66,1c 109,15d 95,46c 0,34a 106,16d Pastaba: BA – biogeniniai aminai, Sm – S. montana, Rc – R.carthamoides, KF – kietafazė fermentacija, TF – tradicinė fermentacija, VS – varškės sūris, K – kontrolė, p – skirtumo tarp rezultatų reikšmių patikimumas, p – patikimas, kai ≤ 0,05, a, b, c, d, e,f – atskirų duomenų vidurkiai, lentelės eilutėse pažymėti skirtingomis raidėmis, tarpusavyje statistiškai patikimai skiriasi (p <0,05)

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Lyginant augalinių bioproduktų apdorojimo technologijos ir PRB įtaką BA formavimuisi VS nustatyta, kad KF apdoroti Sm ir Rc bioproduktai mažino suminį BA kiekį (atitinkamai, vidutiniškai 48,87 % ir 48,17 %), o mažiausias suminis BA kiekis nustatytas VS, pagamintuose su P. acidilactici fermentuotais bioproduktais (atitinkamai, 80,69 % ir 97,85 % mažesnis), lyginant su VS, pagamintais su TF apdorotais bioproduktais. Nefermentuoti Sm ir Rc augalai didino suminį BA kiekį VS (atitinkamai, vidutiniškai 6,3 ir 312,2 karto), lyginant su kontroliniais VS mėginiais. Mažesnės pH vertės nustatytos VS pagamintų su KF apdorotais Sm ir Rc bioproduktais (atitinkamai, vidutiniškai 4,87 % ir 3,87 %) (lyginant su TF) bei vidutiniškai 5,56 % (VS su Sm) ir 4,54 % (VS s Rc) mažesnės (lyginant su kontroliniais mėginiais). BTR patikimai priklausė nuo augalinių bioproduktų fermentacijos metodo (p ≤ 0,0001) ir naudotų fermentacijai PRB (p ≤ 0,0001). Mažesnis D(-) ir didesnis L(+) pieno rūgšties izomerų kiekis nustatytas VS, pagamintuose su KF apdorotais Sm ir Rc bioproduktais (atitinkamai, 19,32 % ir 32,02 % bei 24,71 % ir 72,58 %), lyginant su VS, pagamintais su TF apdorotais bioproduktais. Sm ir Rc fermentacijos technologija, naudotos PRB ir augalo rūšis turėjo reikšmingos įtakos VS bendram priimtinumui (p ≤ 0,0001) ir daugeliu atvejų VS su bioproduktais nustatyti priimtinesni (vidutiniškai, 15,35 % (Sm) ir 17,54 % (Rc), lyginant su kontroliniais VS mėginiais). Lyginant VS su TF ir KF fermentuotais bioproduktais, nustatyta, kad priimtinesni VS gaunami su KF fermentuotais Sm ir Sh (atitinkamai, 13,88 % ir 12,81 %), lyginant su TF. Priimtiniausi įvardinti VS, pagaminti su KF P. acidilactici Sm ir VS su KF L. sakei Rc (atitinkamai, 4,8 taško). Įvertinus vertintojų veido emocines raiškas, kaip emocinį atsaką į VS juslinį priimtinumą (FaceReader programine įranga), nustatyta, kad pastarųjų sąsajos su bendru priimtinumu, įvertintu hedoninėje intensyvumo skalėje, yra statistiškai reikšmingos (p ≤ 0,05) (bendras priimtinumas ir emocinė raiška „laimingas“, bendras priimtinumas ir emocinė raiška „liūdnas“).

4. REZULTATŲ APTARIMAS

PRB - augalų bioproduktai – natūrali priemonė gyvūninės kilmės

maisto produktų praturtinimui ir tvarumo padidinimui. Gyvūninės kilmės maisto produktų kokybės gerinimas, tai daugelį veiksnių apjungiantis procesas: pirminę žaliavų gamybą, maisto mokslą ir gamybos technologijas bei žmonių mitybos poreikius [202]. Fermentacija PRB didina biologiškai aktyvių junginių prieinamumą, produktų mitybinę vertę, gerina juslines bei antimikrobines savybes [283]. Bakteriocinus produkuojančių PRB taikymas

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maisto pramonėje ypač aktualus, siekiant užtikrinti produktų tvarumą [266]. Pramonėje naudojamos PRB geba prisitaikyti prie skirtingų substratų, daugintis ir produkuoti metabolitus kietoje bei skystoje terpėje (KF ar TF), tačiau yra jautrios substratų cheminei sudėčiai [266, 347]. Savitą cheminę sudėtį turinčių augalų ir jų bioproduktų (topinambų, linų sėmenų ir sojų miltų su sumažintu riebalų kiekiu, žirnių skaidulų, Sm, Sh, Rc) apdorojimui, daugeliu atvejų, KF taikymas yra efektyvesnis nei TF (gaunamas didesnis PRB KSV/g substrate, kuris kinta nuo 6,21 log10 KSV/g P. acidilactici TF topinambuose iki 9,98 log10 KSV/g P. pentosaceus KF sojų miltuose).

Vaistinių – prieskoninių augalų naudojimas maisto produktų gamybai yra taikomas ne tik dėl išskirtinių juslinių savybių suteikimo, bet ir dėl minėtų augalų antimikrobinio aktyvumo. Satureja ir Rhaponticum rūšių augalai pasižymi antimikrobiniu, antioksidaciniu ir antigrybiniu aktyvumu [39, 84, 194, 293; 345; 371, 375], o dauguma PRB yra saugios ir pasižymi antimikrobiniu aktyvumu bei teigiama įtaka maisto produktų tekstūros ir fizikinėms cheminėms savybėms [73, 297]. Dėl šios priežasties PRB – augalų bioproduktų sukūrimas ir pritaikymas gyvūninės kilmės maisto produktų vertės bei tvarumo didinimui yra labai perspektyvus. Sm, Sh ir Rc fermentuoti P. acidilactici, L. sakei ir P. pentosaceus pasižymi antimikrobiniu aktyvumu prieš sporas formuojančias aerobines mezofilines bakterijas, Enterobakterijas, mieles ir pelėsinius grybus, ir ši jų savybė priklauso nuo fermentacijos metodo (p ≤ 0,0001), fermentacijai naudotų PRB (p ≤ 0,0001) ir analizuotų veiksnių įtaka bioproduktų antimikrobiniam aktyvumui yra reikšminga (p ≤ 0,0001).

Sm, Sh ir Rc augalai naudojami kaip biologiškai aktyvūs ir saugūs maisto priedai, mažinantys nepageidaujamos mikrofloros augimą, veikiant jų sudėtyje esantiems eteriniams aliejams (EA). EA komponentai (karvakrolis, timolis, linalolis) pasižymi stipriu antimikrobiniu ir antioksidaciniu aktyvumu [44, 87, 124, 129, 130, 250, 359]. Nustatyta, kad šioje disertacijoje tirtų augalinių ekstraktų antimikrobinis aktyvumas padidėjo, atlikus fermentaciją PRB, ir priklausė nuo patogeninių mikroorganizmų rūšies (bioproduktų fermentacijai naudota PRB ir augalo rūšis turėjo reikšmingos įtakos, atitinkamai, E.coli p ≤ 0,010, ir p ≤ 0,002, B. subtilis p ≤ 0,0001 ir p ≤ 0,0001, P. fluorescens biovar. III p ≤ 0,023 ir p ≤ 0,001 bei P. fluorescens biovar. V p ≤ 0,001 ir p ≤ 0,001 slopinimui.

Fermentacijos PRB metu sumažėja substrato pH, o tai yra svarbus veiksnys galutinio produkto juslinių savybių formavimuisi, cheminio bei biologinio tvarumo padidinimui [294, 314, 332]. PRB mažina substrato pH, ir didina BTR, nes didėja pieno rūgšties kiekis. Fermentacijos metu heterofermentinės PRB gamina įvairias organines rūgštis, mažina terpės pH ir didina BTR [293, 332], ir priklausomai nuo PRB rūšies bei substrato

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cheminės sudėties savitumo išskiria L(+) ir D(-) pieno rūgšties izomerų mišinį [114, 373].

Daugelis PRB išskiria į fermentuojamą substratą proteolitinius fermentus [401]. Ištyrus hidrolazių (amilazių, proteazių) kinetiką fermentuojamuose substratuose, nustatyta, kad didesnis amilolitinių fermentų aktyvumas gaunamas KF, o proteolitinių – TF apdorotuose bioproduktuose (atitinkamai, vidutiniškai nuo 0,83 iki 3,69 karto topinambuose ir sojų miltuose, ir vidutiniškai nuo 1,08 iki 4,52 karto, žirnių skaidulose ir linų sėmenyse) ir amilolitinių fermentų aktyvumas daugiau priklauso nuo fermentuojamojo substrato cheminės sudėties specifikos nei nuo jo drėgnio.

Dekarboksilaziniu aktyvumu pasižymintys mikroorganizmai nėra tinkami baltymingų žaliavų fermentavimui, nes gali inicijuoti BA formavimasi. BA kokybinė ir kiekybinė sudėtis produktuose priklauso nuo fermentuojamo substrato cheminės sudėties, pašalinės ir technologijoje naudojamos mikrofloros, aplinkos ir kitų veiksnių [47]. Nustatyta, kad augalų fermentacijai naudotos PRB (p ≤ 0,018) ir augalo rūšis (p ≤ 0,0001) turėjo reikšmingos įtakos suminiam BA kiekiui, o KF daugeliu atvejų didino BA kiekį nuo 2,1 % topinambuose iki 19,7 % sojų miltuose. Tai galima paaiškinti efektyvesniu mikroorganizmų metabolitų produkavimu, esant minimaliam drėgmės kiekiui substrate. KF dažnai naudojama pramonėje, tikslu gauti kuo didesnes metabolitų išeigas.

Fermentacija PRB turi teigiamos įtakos substrato maistinei vertei ir didina biologiškai aktyvių junginių bioprieinamumą (didina fenolinių rūgščių, BFJ, tirpių skaidulinių medžiagų ir lignanų kiekį) [186, 229, 300, 308] bei baltymų virškinamumą [16, 93]. Panašios tendencijos nustatytos išanalizavus gautus tyrimų rezultatus: PRB fermentuotuose augaluose nustatytas didesnis lignanų (vidutiniškai, TF 27,9 %, KF – 41,0 %), BFJ ir LRSGA (atitinkamai, vidutiniškai TF – 371,76 %, KF – 396,0 % ir TF – 202,92 %, KF – 218,47 %), tačiau mažesnis ARs ir β-gliukanų kiekis (vidutiniškai, TF – 15,13 %, KF – 17,02 % ir TF – 58,80 %, KF – 59,93 %). Choung ir kt. [58] ir [313] publikavo, kad KF didina BFJ kiekį substrate ir jo antioksidacinį aktyvumą, [89, 90] bei [173] publikavo, kad fermentacijos metu didėja substrato LRSGA. Mokslininkai Oyewole ir kt. [274] nustatė, kad fermentuojant didėja lignanų kiekis ir priklauso nuo glikozidinių kompleksų, kurių sudėtyje yra lignanų, išardymo augalinėje matricoje veikiant amilolitiniams fermentams.

Alternatyvioje terpėje pagausintų pieno rūgšties bakterijų taikymas

kiaulienos, jautienos ir vištienos kokybės rodiklių pagerinimui. Mėsos fermentacijos metu vykstantys fiziniai, biocheminiai ir mikrobiologiniai pokyčiai suteikia galutiniam produktui išskirtines juslines, maistines ir

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technologines savybes. PRB, produkuojančios antimikrobinius junginius (bakteriocinus, organines rūgštis ir kt.), gali būti naudojamos kaip natūralūs konservantai, padedantys ilgesnį laiką išlaikyti mėsos gaminių spalvą bei pagerinantys tekstūrą, juslines savybes bei teigiamai veikiantys žmonių sveikatą [98, 137, 276]. Marinavimas, tai mėsos apdorojimo būdas, pagerinantis svarbiausias mėsos technologines savybes: VR, švelnumą, juslines savybes, sumažina drėgmės ir virimo nuostolius, stabilizuoja spalvą [149, 163]. Kiaulienos, jautienos ir vištienos apdorojimas PRB pagrindu pagamintais marinatais daugeliu atvejų pagerino technologines gaminių savybes: padidino tradiciškai ir ekologiškai pagamintos vištienos VR (atitinkamai, 0,2 % ir 0,3 %) ir švelnumą (nuo 8,1 % iki 26,1 %), kiaulienos ir jautienos švelnumą (atitinkamai, vidutiniškai 6,4 % ir 18,9 % (P. acidilactici marinatas), 4,6 % ir 27,7 % (P. pentosaceus marinatas), 2,2 % ir 15,9 % (L. sakei marinatas).

Mėsos spalva ir pH yra svarbūs kokybės rodikliai, o jų kitimas technologinių procesų metu yra griežtai kontroliuojamas [228]. Marinavimas natūraliais, PRB pagrindu pagamintais marinatais, didina kiaulienos, jautienos ir vištienos (L*) koordinates (atitinkamai, 20,56 %, 8,73 % ir 3,0 %) ir kiaulienos rausvumą (a*) (12,54 %), tačiau sumažina jautienos ir vištienos rausvumą (a*) (atitinkamai, vidutiniškai, 24,75 % ir 6,17 %). Taip pat, sumažėja kiaulienos ir vištienos gelsvumo (b*) koordinatės (atitinkamai, 15,20 % ir 9,65 %). Jautienos gelsvumo (b*) koordinatės padidėja (55,79 %). Marinavimo metu spalva kinta dėl mioglobino sąveikos su azoto monoksidu bei veikiant pH (mėsa šviesėja) [222]. Nekontroliuojant marinavimo proceso, gali atsirasti nepageidaujami mėsos pokyčiai: pakisti spalva, įvykti riebalų oksidacija, padidėti fermentų aktyvumas [25].

PRB pasižymi dideliu aminopeptidazinių ir dekarboksilazinių fermentų aktyvumu, kuriems veikiant susidaro nepageidaujami junginiai – BA. Histaminas (HIS), tiraminas (TIR), rečiau feniletilaminas (FEN) yra pagrindiniai BA, siejami su neigiamu poveikiu vartotojų sveikatai: histamininiu apsinuodijimu, maisto netoleravimu, migrena, hipertenzija, smegenų hemoragija ir širdies darbo sutrikimais [363]. Putrescinas (PUT) (7,13–131,29 mg/kg) ir kadaverinas (KAD) (4,77–65,71 mg/kg) buvo pagrindiniai BA daugelyje kiaulienos, jautienos ir vištienos mėginių, o HIS kiekis kito nuo 1,37–44,85 mg/kg. Beutling ir kt. [35] nustatė, kad neigiamą poveikį vartotojų sveikatai gali sukelti su maistu gaunamas 10–40 g HIS, 5–10 g TIR, 25 g triptamino ir 5 g PHE kiekis, o toksiška dozė 10–80 g HIS bei 25–250 mg TIR. Marinatų gamybai naudotos PRB turėjo reikšmingos įtakos (p ≤ 0,0001) suminiam BA kiekiui kiaulienoje ir jautienoje (atitinkamai, vidutiniškai suminis BA kiekis padidėjo marinuojant P.

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acidilactici marinatu – 11,78 % ir 58,04 %, L. sakei – 36,03 % ir 64,71 % ir P. pentosaceus – 58,10 % ir 66,12 %). Marinavimas P. acidilactici sumažino BA kiekį ekologiškai pagamintoje vištienoje vidutiniškai 39,48 %, o tradiciškai pagamintoje padidino 158,05 %. PRB skatina akumuliacinėmis savybėmis pasižyminčių BA susidarymą, todėl jų kiekį PRB apdorotuose maisto produktuose būtina kontroliuoti [35, 214).

Augalinių bioproduktų įtaka maltos mėsos produktų ir kiaulienos

bei jautienos nugarinės kokybės ir saugos rodikliams. Vienas iš natūralių būdų padidinti maisto produktų saugą, tvarumą ir funkcionalumą, tai naudoti antimikrobinėmis savybėmis pasižyminčių PRB ir aromatinių augalų kompleksinius bioproduktus. Priimtiniausi MMP ir kiaulienos bei jautienos nugarinės mėginiai buvo apdoroti, atitinkamai, 3 % ir 2 % bioproduktų. Didelis aromatinių augalų kiekis yra nepriimtinas vartotojams dėl stipraus, šildančio, aštraus kvapo ir skonio [135], MMP su 5 % ir 7 % bioproduktų priedo buvo įvardinti kaip nepriimtini.

3 % Sm bioprodukto užtikrino MMP biologinę saugą: po 5 parų laikymo +4 ºC temperatūroje MMP su Sm priedu padidėjo nepageidaujamų mikroorganizmų kiekis, tačiau koliforminių bakterijų ir pelėsinių grybų kiekis neviršijo Europos Komisijos Parlamento reglamento Nr. 852/2004 nustatytų normų. Sm bioprodukto antimikrobiniam aktyvumui prieš mezofilines ir koliformines bakterijas, mieles ir pelėsinius grybus reikšmingos įtakos turėjo bioprodukto fermentacijos būdas (TF ir KF) ir naudotos PRB (p ≤ 0,05). Didžiausiu antimikrobiniu aktyvumu pasižymėjo P. acidilactici fermentuoti Sm bioproduktai (vidutiniškai, mezofilinių bakterijų skaičių MMP sumažino 23,45 %, koliforminių 28,22 %, mielių ir pelėsinių grybų 50,75 %), lyginant su kontroliniais MMP. KF fermentuoti Sm bioproduktai sumažino mezofilinių ir koliforminių bakterijų kiekį (atitinkamai, 22,01 % ir 8,13 %) MMP, tačiau padidino mielių ir pelėsinių grybų kiekį 1,79 %, lyginant MMP, pagamintais su Sm TF bioproduktais. Didesnis antimikrobinis aktyvumas nustatytas ne atskirų bioproduktų komponentų (PRB ir Sm), o jų derinio (sinergistinio poveikio). Sm bioproduktų antimikrobinis aktyvumas gali priklausyti nuo fermentacijos metu susidarančios pieno rūgšties [21], PRB produkuojamų bakteriocinų [59] ir Satureja rūšies augalams būdingų antimikrobinių savybių [333].

Sm yra intensyviu aromatu pasižymintis prieskoninis augalas, dažniausiai naudojamas mėsos ir žuvies produktų gamyboje. Jo EA sudėtyje esantys AJ (karvakrolis, ρ-kimenas, timolis) pasižymi antimikrobiniu ir antoksidaciniu aktyvumu [280, 167, 345]. Suminis AJ kiekis MMP su Sm bioproduktais padidėjo nuo 15,46 iki 23,66 kartų, Sm bioproduktai padidino ρ-kimeno, γ-terpineno ir karvakrolio kiekį MMP (atitinkamai, vidutiniškai 36,6; 7,1 ir

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5,9 karto), lyginant su kontroliniais MMP. Mėsos oksidacijos metu formuojasi nepageidaujamo kvapo, toksiški junginiai, mėsos spalva šviesėja, trumpėja jos tinkamumo vartoti terminas, didėja drėgmės nuostoliai, prastėja produkto kokybė [66, 99]. Antioksidacinėmis savybėmis pasižyminčių augalų naudojimas gali sumažinti mėsos oksidaciją ir padidinti švelnumą [66]. MMP kokybės rodikliams reikšmingos įtakos turėjo Sm ir Sh fermentacijos būdas (p ≤ 0,05), o kiaulienos ir jautienos nugarinės – augalų fermentacijos technologija (p ≤ 0,0001), naudotos PRB (p ≤ 0,0001) ir augalo rūšis (p ≤ 0,0001). Sm ir Sh bioproduktai daugeliu atvejų didino MMP, kiaulienos ir jautienos VR (atitinkamai, vidutiniškai, 11,71 % ir 7,69 %, 0,69 % ir 1,45 %, 1,19 % ir 2,50 %) ir švelnumą (išskyrus kiaulieną, mažino atitinkamai, 115,87 % ir 39,05 %) (atitinkamai, vidutiniškai, 39,29 % ir 36,90 % bei 30,40 % ir 31,52 %). Mokslininkai Lantto ir kt. [212] publikavo, kad augalai, savo sudėtyje turintys skaidulinių medžiagų, naudojami mėsos produktų tekstūros gerinimui, o mokslininkas Ke (2006) įrodė, kad fermentacija PRB lėtina riebalų oksidaciją ir padidina mėsos VR bei švelnumą.

Aromatiniuose augaluose esantys antioksidantai stabilizuoja mėsos spalvą ir prailgina jos tinkamumo vartoti laiką [181]. Augaliniai bioproduktai turėjo reikšmingos įtakos MMP, pagamintų iš kiaulienos ir jautienos nugarinės, spalvų pokyčiams: daugeliu atvejų mažino šviesumą (L*) (atitinkamai, Sm 16,89 % ir Sh 20,06 %, Sm 17,24 % ir Sh 5,48 %, ir Sm 29,07 % ir Sh 31,26 %) bei ir gelsvumą (b*) (atitinkamai, vidutiniškai Sm 14,44 % ir Sh 9,49 %, Sm % 24,18 ir Sh 16,63 %, ir Sm 17,78 %, o Sh didino vidutiniškai 51,64 %). MMP ir kiaulienos nugarinės rausvumo (a*) koordinates Sm ir Sh bioproduktai didino (atitinkamai, vidutiniškai 37,35 % ir 37,06 %, 117,24 % ir 58,37 %), o jautienos mažino (atitinkamai, vidutiniškai 35,49 % ir 34,23 %). Mėsos spalva yra labai svarbus kokybės rodiklis, todėl būtina slopinti pigmentų oksidaciją. Priimtiniausia yra rausvos spalvos mėsa, kuriai įtakos turi mėsoje vyraujančios mioglobino formos (šviesiai rausva – oksimioglobinas, raudona – mioglobinas, pilka – ruda – metmioglobinas) [245]. Faustman ir kt. [102] nustatė, kad riebalų oksidacija spartina mioglobino oksidaciją ir skatina mėsos spalvos intensyvumo mažėjimą, todėl būtų perspektyvu naudoti riebalų oksidaciją slopinančius augalus ir bakteriocinus produkuojančias PRB šių procesų lėtinimui [181, 184].

Nepaisant to, kad augaliniai bioproduktai daugeliu atvejų gerino mėsos ir jos produktų technologines savybes, jie gali skatinti BA formavimąsi. Fermentacijos metu vykstantis baltymų skilimas padidina mėsos produktų funkcionalumą, tačiau tuo pačiu metu sudaromos palankios sąlygos dekarboksilaziniu aktyvumu pasižymintiems mikroorganizmams inicijuoti

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BA formavimasi [76]. Mėsoje BA būtina kontroliuoti dėl jų neigiamo poveikio žmonių sveikatai, taip pat šie junginiai įvardijami kaip mėsos šviežumo indikatoriai [24]. Sm bioproduktai mažino suminį BA kiekį MMP (0,89 % (TF) ir 34,87 % (KF)), o Sh didino (168,94 % (TF) ir 164,47 % (KF)), lyginant su kontroliniais mėginiais, o jų kiekis priklausė nuo bioproduktų fermentacijos technologijos (p ≤ 0,0001), fermentacijai naudotų PRB (p ≤ 0,0001) ir šių faktorių tarpusavio sąveikos (p ≤ 0,0001). PUT, HIS, SPR ir SPD buvo pagrindiniai BA kontroliniuose mėginiuose, KAD ir TIR – MMP, pagamintuose su Sm, o PUT, KAD ir HIS – MMP, pagamintuose su Sh. Ruiz-Capillaz ir Jiménez-Colmenero [327] nustatė, kad TIR, KAD, PUT ir HIS yra pagrindiniai BA apdorotoje mėsoje, o jų koncentracija didėja apdorojimo ir laikymo metu. Hernández-Jover ir kt. [143] ir [327] nustatė, kad SPD ir SPR yra randami tik šviežioje mėsoje, o jų kiekis apdorojimo ir laikymo metu mažėja.

Žirnių skaidulos MMP be glitimo gamybai. Produktų be glitimo

kūrimas gali užtikrinti celiakija sergančių ir glitimo baltymų netoleruojančių vartotojų poreikius įvairiems ir natūraliems maisto produktams [253, 256, 404]. Glitimas yra svarbus maisto produktų tekstūrai bei juslinėms savybėms, todėl jo pašalinimas ar pakeitimas kitomis žaliavomis yra sunkus uždavinys gamintojams, kurių tikslas eliminuoti iš produktų alergijas sukeliančius baltymus, tačiau išlaikyti naujai sukurtų produktų maistinę vertę bei priimtinumą [341, 404]. Alternatyva glitimo turinčioms žaliavoms gali būti ryžių ir kukurūzų krakmolas, ankštinių kultūrų miltai ir skaidulinės medžiagos [252]. Kuriant MMP be glitimo, gaminių receptūra buvo koreguojama manų kruopas keičiant žirnių skaidulomis. Skaidulinės medžiagos gali būti naudojamos mėsos produktų gamyboje kaip rišamoji ir užpildanti medžiaga, pakeičianti dalį riebalų, didinanti bendrą priimtinumą ir juslines savybes, funkcionaliąją vertę, pH, VR bei emulsinių sistemų stabilumą [372]. Patikimi skirtumai (p ≤ 0,05) nustatyti tarp MMP, pagamintų su manų kruopomis ir žirnių skaidulomis kokybės rodiklių. Žirnių skaidulos turėjo teigiamos įtakos MMP technologiniams rodikliams: didino VR (8,90 %) ir švelnumą (1,68 kartus) bei mažino VN (2,90 %). Elleuch ir kt. [95] ir Pinero ir kt. [301] nustatė, kad skaidulinių medžiagų įterpimas į mėsos produktus didina VR ir sausųjų medžiagų kiekį juose.

Labai svarbu užtikrinti, kad naujai įterpiamos žaliavos neturėtų neigiamo poveikio mėsos gaminių spalvai [388]. Manų kruopos yra šviesios spalvos, neutralaus skonio ir kvapo [37], o žirnių skaidulų spalva kinta nuo baltos iki žalios (priklausomai nuo rūšies ir apdorojimo būdo). MMP, pagaminti su žirnių skaidulomis, buvo tamsesni (6,76 %), rausvesni (a*) (4,87 %) ir gelsvesni (b*) (0,53 %), lyginant su MMP, pagamintais su manų kruopomis.

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Žalia ankštinių augalų spalva gali pabloginti mėsos produktų spalvą ir sumažinti mėsos produktų bendrą priimtinumą [396]. Didesnis suminis BA kiekis nustatytas MMP su žirnių skaidulomis (23,90 %), lyginant su MMP, pagamintais su manų kruopų priedu. BA kiekis bei kompozicija priklauso nuo mėsos fermentacijos bei laikymo sąlygų [103], fermentuoti žirnių skaidulų bioproduktai padidino BA kiekį MMP.

PRB įtaka šaltai rūkytų dešrų saugos rodikliams. Mėsa ir jos

produktai yra vieni iš pagrindinių PAA šaltinių [91]. Pastaruoju metu didėja susidomėjimas mikroorganizmų gebėjimu absorbuoti nepageidaujamus cheminius junginius iš maisto matricų ir taip sumažinti jų kiekį iki vartotojams nekenksmingo. [2, 3, 4] nustatė, kad PRB mažina PAA formavimąsi rūkytuose mėsos produktuose. Tirtuose, PRB neapdorotuose, rūkytų dešrų mėginiuose PAA kiekis nesiekė 2,0 mg/kg (2,0 mg/kg yra toksiškas kiekis) [219]. Šaltai rūkytų dešrų paviršiaus apdorojimas PRB reikšmingai sumažino PAA kiekį: BaA (p = 0,0001), BaP (p = 0,001), BbF (p = 0,0001) ir Chr (p = 0,0001). Dešrų paviršiaus apdorojimui naudotų PRB substratų pH kito nuo 4,21 iki 4,42. Zhao ir kt. [411] nustatė, kad didžiausiu BaP surišimo pajėgumą PRB pasiekia, esant terpės pH 4,0–5,0. Didesnis PAA kiekis nustatytas išoriniame šaltai rūkytų dešrų sluoksnyje, nei vidiniame, tačiau apdorojimas PRB sumažino suminį PAA kiekį abiejose dešrų dalyse (vidutiniškai, atitinkamai, prieš rūkymą 25,50 % ir 28,29 %, po rūkymo – 17,69 % ir 20,63 %), lyginant su neapdorotais mėginiais. Ledesma ir kt. [217, 218] nustatė, kad didžiausi PAA kiekiai formuojasi išorinėje produkto dalyje, o [15] ir [339] nustatė, jog rūkymo metu mėsos išorėje susidarę PAA migruoja ir į vidinius gaminių sluoksnius. Zhao ir kt. [411] nustatė, kad kai kurios PRB gali degraduoti toksiškus junginius, o [112] ir [140] mano, kad PRB toksiškus junginius metabolizuoja į specifinius fermentus arba prijungia prie ląstelių sienelių komponentų.

PRB sumažino PAA kiekį šaltai rūkytose dešrose, tačiau padidino BA kiekį. Liu ir kt. [228] teigia, kad dešrų fermentacija turi įtakos BA kiekio didėjimui jose. Šaltai rūkytų dešrų apdorojimas PRB prieš rūkymą ir po jo didino suminį BA kiekį (atitinkamai, vidutiniškai 45,41 % ir 42,61 %). Dešrų paviršiaus apdorojimas PRB prieš rūkymą sumažino KAD ir SPR, o po rūkymo sumažino PUT kiekį (vidutiniškai 53 % apdrojant L. sakei ir P. acidilactici). Šaltai rūkytas dešras apdorojus P. pentosaceus PUT jose nenustatyta. Straub ir kt. [366] nustatė, kad Pediococci nepasižymi dekarboksilaziniu aktyvumu, o [365] nustatė, jog P. acidilactici KTU05-7, P. pentosaceus KTU05-9, L. sakei KTU05-6 mažino BA kiekį MMP. BA formavimasis priklauso nuo mikrofloros bei fizikinių cheminių veiksnių

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(pH, temperatūros, druskos ir cukraus kiekio bei deguonies koncentracijos). Dekarboksilaziniu aktyvumu pasižyminčių mikroorganizmų veikla taip pat didina BA kiekį produktuose [215]. Tačiau, plačiu antimikrobiniu spektru pasižyminčios bakteriocinus produkuojančios PRB gali mažinti PAA kiekį, prailginti tinkamumo vartoti terminą bei slopinti nepageidaujamos mikrofloros augimą mėsos produktuose [201]. P. acidilactici, P. pentosaceus ir L. sakei antimikrobinis aktyvumas patikimai priklausė nuo PRB ir patogeninių mikroorganizmų rūšies (prieš P. aeruginosa p ≤ 0,0001, S. aureus p ≤ 0,002, L. monocytogenes p ≤ 0,006, E. coli p ≤ 0,003, S. typhimurium p ≤ 0,0001, Y. enterolitica p ≤ 0,001, Y. pseudotuberculosis p ≤ 0,0001). PRB produkuojami junginiai pasižymi antimikrobiniu aktyvumu [9, 59, 325], o fermentacijos metu išskirtos organinės rūgštys, vandenilio peroksidas, bakteriocinai ir kt. junginiai mažina terpės pH, slopina pašalinės mikrofloros augimą, dėl šios priežasties PRB gali būti naudojamos, siekiant užtikrinti technologinio proceso kontrolę [9]. Šiame eksperimente naudotos PRB pasižymėjo antimikrobiniu aktyvumu prieš visus tirtus nepageidaujamus mikroorganizmus ir gali būti rekomenduojamos šaltai rūkytų dešrų paviršiaus apdorojimui, siekiant sumažinti mikrobinę ir cheminę taršą.

Augaliniai bioproduktai varškės sūrių tvarumo didinimui. VS yra

tradicinis rytų Europos šalių produktas, pasižymintis trumpu vartoti tinkamumo terminu [168]. Antimikrobinėmis savybėmis pasižyminčios PRB ir aromatiniai augalai (Satureja rūšies, savo EA turintys karvakrolio ir timolio) gali būti naudojami VS gamyboje kaip natūralūs konservantai, prailginantys tinkamumo vartoti terminą [52, 64, 291]. Nustatyta, kad AJ kiekis VS, praturtintuose Sm ir Rc bioproduktais, patikimai priklausė nuo augalų rūšies (p ≤ 0,05). Didesnis AJ kiekis nustatytas VS su Sm ir Rc (atitinkamai, vidutiniškai 7,03 ir 8,96 kartus), lyginant su kontroliniais VS mėginiais. Sm bioproduktai didino karvakrolio ir timolio (atitinkamai, vidutiniškai 21,88 ir 12.68 kartus), o Rc – timolio ir ρ-kimeno (atitinkamai, vidutiniškai 25,24 ir 18,86 kartus) kiekį VS mėginiuose. Karvakrolio VS, pagamintuose su Rc nebuvo nustatyta. Atskirų AJ komponentų sudėtis fermentacijos metu kito: vienų junginių kiekis sumažėjo, kitų padidėjo. Panašios AJ pokyčių tendencijos nustatytos S. montana sėklas fermentuojant PRB [176].

Fermentuoti pieno produktai yra aukštos maistinės vertės, tačiau, jie gali būti ir neigiamą poveikį vartotojų sveikatai turinčių junginių – BA šaltinis [208]. Sūriuose, ypač ilgo brandinimo, nustatomi dideli BA kiekiai, lyginant su kitomis maisto produktų grupėmis [319]. BA kiekis sūriuose priklauso nuo gamybos sąlygų, brandinimo ir laikymo trukmės, pieno pasterizavimo

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efektyvumo bei mikroorganizmų kultūrų, naudojamų rauginimui [363]. Parinkti PRB, mažinančias BA kiekį, yra svarbus uždavinys, norint užtikrinti sūrių saugą. Nustatyta, kad augalų fermentacijos būdas p ≤ 0,0001, fermentacijai naudotos PRB p ≤ 0,0001, augalo rūšis p ≤ 0,0001 turėjo reikšmingos įtakos BA formavimuisi VS ir šių faktorių sąveika buvo reikšminga (p ≤ 0,0001). Pagrindinis BA, nustatytas VS, pagamintuose su augaliniais bioproduktais buvo KAD (3,55–137,6 mg/kg), o KF apdoroti Sm ir Rc bioproduktai mažino suminį BA kiekį (atitinkamai, vidutiniškai 48,87 % ir 48,17) VS. Piene BA kiekis yra apie 1 mg/l, o sūriuose BA kiekis gali padidėti iki 1000 mg [227]. Calzada ir kt. [46] nustatė, kad bakteriocinus produkuojančios PRB gali slopinti dekarboksilaziniu aktyvumu pasižyminčių Enterobakterijų dauginimąsi pieno produktuose, o [223] nustatė, kad kai kurios Lactobacillus, Pediococcus ir Micrococcus rūšys gali sumažinti TIR ir HIS kiekį anaerobinėmis sąlygomis, veikiant monoamino oksidazėms.

Augaliniai bioproduktai pagerino varškės sūrių saugos ir kokybės rodiklius bei padidino bendrą priimtinumą: nustatytas mažesnis D(-) ir didesnis L(+) pieno rūgšties izomerų kiekis VS, su KF apdorotais Sm ir Rc bioproduktais (atitinkamai, 19,32 % ir 32,02 % bei 24,71 % ir 72,58 %), Sm ir Rc padidino VS stabilumą – bioproduktai lėtino mielių augimą ir prailgino VS vartoti tinkamumo terminą (atitinkamai, 5 ir 3 paromis), lyginant su kontroliniais mėginiais. Dėl šių priežasčių Sm ir Rc bioproduktai gali būti rekomenduojami tvaresnių ir geresnės kokybės VS gamybai.

IŠVADOS

1. Fermentacija L. sakei KTU05-6, P. acidilactici KTU05-7 ir P.

pentosaceus KTU05-8 yra tinkama technologinė priemonė Sm, Sh, Rc, linų sėmenų, topinambų, sojų miltų ir žirnių skaidulų bioproduktų gamybai, tačiau proceso sąlygos (PRB parinkimas bei substrato drėgnis) turi būti modeliuojamos, priklausomai nuo substrato cheminės sudėties ir norimų gauti bioprodukto savybių. Šie veiksniai turi reikšmingos įtakos (p ≤ 0,05) rūgštingumo rodikliams, PRB kiekiui, amilolitinių ir proteolitinių fermentų aktyvumui.

2. Fermentacijos technologija turi reikšmingos įtakos biologiškai aktyvių

junginių kiekiui ir substrato LRSGA (p ≤0,05): didina lignanų (MAT ir SEKO) bei BFJ kiekį ir substrato LRSGA, mažina ARs homologų (C15:0, C19:0, C21:0, ir C23) ir β-gliukanų koncentraciją.

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3. KF ir TF turi nevienareikšmės įtakos augalinių bioproduktų saugai, tačiau visada fermentuojamame substrate mažina nepageidaujamų mikroorganizmų kiekį, o D(-) pieno rūgšties izomerų bei BA kiekis neviršija literatūroje nurodomų vartotojų sveikatai kenksmingų kiekių.

4. PRB – augalų bioproduktų antimikrobinis aktyvumas didesnis nei

atskirų jų komponentų (augalo ar PRB) ir šiai savybei įtakos turi bioproduktų gamybai parinkta PRB rūšis (prieš E.coli p ≤ 0,010, B. subtilis p ≤ 0,0001, P. fluorescens biovar. III p ≤ 0,023 ir P. fluorescens biovar. V p ≤ 0,001) ir augalo rūšis (prieš E.coli p ≤ 0,002, prieš B. subtilis, P. fluorescens biovar. III, P. fluorescens biovar. V p ≤ 0,0001).

5. Naujai sukurtų PRB – augalų bioproduktų panaudojimas yra efektyvus

technologinis sprendimas gyvūninės kilmės maisto produktų vertei, saugai ir tvarumui padidinti:

5.1. Sm ir Sh bioproduktai yra saugūs BA aspektu ir gali būti rekomenduojami MMP gamybai, nes mažina biotaršą, didina AJ ir BFJ kiekį, stabilizuoja spalvą bei pagerina juslines savybes;

5.2. Pediococci bioproduktai yra saugūs ir didina kiaulienos VR, švelnumą, rausvumą bei bendrą priimtinumą (lyginant su nemarinuotais mėginiais).

5.3. Jautienos švelnumui padidinti galima rekomenduoti L.sakei, o bendram priimtinumui padidinti – P. acidilactici bioproduktus.

5.4. P. acidilactici marinatai yra saugūs ir didina tradiciškai ir ekologiškai pagamintos vištienos VR ir švelnumą.

5.5. PRB bioproduktai pasižymi antimikrobinėmis savybėmis ir mažina BA (KAD, SPD ir PUT) ir PAA (BaP ir Chr) kiekį šaltai rūkytose dešrose.

5.6. Sm ir Rc bioproduktai gali būti rekomenduojami VS gamyboje, siekiant padidinti jų tvarumą, AJ kiekį (timolio, karvakrolio ir p – kimeno) bei bendrą priimtinumą.

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APPENDIXES

Table 1. Moisture content (%), TTA (°N), pH, L(+) and D(-)-lactic acid isomers content (g/100 g) of SMF and SSF fermented savory plants

PP

L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF

Moisture content after 48 h fermentation, % Sm 71.85±0.6a 40.12±0.7a 69.53±1.0 38.24±0.7a 75.05±1.2b 40.1±0.5a Sh 78.32±0.5c 43.95±0.9e 71.76±1.1a 44.63±1.1b 80.14±1.3b 42.9±0.7b Rc 75.25±0.4b 42.68±0.8b 73.82±0.9b 41.31±0.8a 71.36±1.3a 42.5±0.6b

Total titratable acidity after 48h of fermentation, °N Sm 12.8±0.64a 10.3±0.27a 11.7±0.63a 11.5±0.36a 10.5±0.84a 11.4±0.5a Sh 13.5±0.39b 11.6±0.31b 13.3±0.59b 12.1±0.41b 14.3±0.69b 12.8±0.3b Rc 13.2±0.49b 11.7±0.38b 12.9±0.63b 11.4±0.32a 13.5±0.93b 11.7±0.5a

pH, after 24 h of fermentation Sm 5.67±0.04a 6.27±0.05b 5.58±0.06a 6.17±0.07b 5.53±0.05a 5.99±0.4a Sh 5.75±0.03b 6.24±0.02b 5.79±0.09b 5.99±0.06a 5.53±0.03a 6.09±0.9a Rc 5.62±0.05a 6.14±0.03a 5.77±0.06b 5.98±0.03a 5.93±0.06b 6.10±0.8b

pH, after 48 h of fermentation Sm 5.42±0.05c 5.89±0.06b 5.34±0.08c 5.93±0.07c 5.05±0.02a 5.49±0.7a Sh 4.53±0.06a 5.70±0.05a 4.41±0.05a 5.74±0.08b 5.36±0.06b 5.61±0.8b Rc 4.53±0.05a 5.87±0.03b 4.71±0.04b 5.51±0.09a 4.93±0.05c 5.42±0.6b

L(+)-lactic acid content after 48 h of fermentation, g/100g Sm 4.71±0.08a 5.92±0.09c 6.91±0.19c 6.67±0.13b 6.83±0.14e 6.13±0.1b Sh 4.94±0.08a 6.04±0.11c 6.31±0.11c 6.41±0.11b 5.42±0.09d 6.22±0.1b Rc 5.44±0.07b 2.19±0.03a 2.94±0.05a 5.44±0.06a 0.40±0.01a 5.67±0.7a

D(-)-lactic acid content after 48 h of fermentation, g/100g Sm 3.79±0.07b 5.01±0.10c 4.19±0.10a 4.02±0.12a 3.27±0.08a 4.13±0.8b Sh 3.84±0.09b 4.48±0.08b 4.21±0.09a 3.87±0.07a 3.48±0.09a 3.14±0.9a Rc 2.47±0.05a 2.94±0.03a 4.55±0.04b 6.58±0.08c 5.31±0.06b 4.57±0.7b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within rows with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana Sh – S. hortensis; Rc – R. carthamoides, P – plant product.

Table 2. Microbiological parameters of LAB, spore of aerobic mesophilic bacteria, Enterobacteria , yeast and mold of fermented savory plants

P L. sakei P. acidilactici 7 P. pentosaceus 8 NF SMF SSF SMF SSF SMF SSF

Lactic acid bacteria, log10 CFU/g Sm 8.59

±0.09b 7.86

±0.08a 7.46

±0.07a 9.20

±0.10a 9.26

±0.10a 9.32

±0.09b 3.07

±0.02a Sh 6.48

±0.07a 9.11

±0.09b 8.67

±0.08b 9.74

±0.10b 5.43

±0.06a 7.48

±0.08a 2.86

±0.02a Rc 8.46

±0.09b 8.49

±0.08a 8.43

±0.07b 9.28

±0.10a 9.20

±0.09c 9.42

±0.09b 3.76

±0.03b

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Continue of Table 2. P L. sakei P. acidilactici 7 P. pentosaceus 8 NF SMF SSF SMF SSF SMF SSF

Spore of aerobic mesophilic bacteria, log10 CFU/g

Sm 1.08±0.02a 1.42±0.03a 1.16±0.01a - - - 4.30±0.05a Sh 1.66±0.02b 1.59±0.02a 1.43±0.01a 1.3±0.1a - - 4.75±0.04a Rc 2.05±0.02c 2.12±0.02b 1.94±0.02b - - - 6.29±0.07b

Enterobacteria, log10 CFU/g Sm - - - - - - 3.43±0.04a Sh 1.2±0.01a 1.33±0.01a - - - - 3.51±0.03b Rc 1.5±0.02a - 1.9±0.02a - - - 3.48±0.04a

Yeasts and mold, log10 CFU/g Sm - - - - - - 2.65±0.03a Sh - - - - - - 3.15±0.03a Rc - - - - - - 8.29±0.09c

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; log10 CFU/g – colony-forming units per gram; Sm – S. montana Sh – S. hortensis; Rc – R. carthamoides, P – plant product.

Table 3. Moisture content (%), pH, TTA (°N), L(+) and D(-)-lactic acid isomers content (g/100 g) in fermented plant bioproducts

P L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF

Moisture content after 48 h fermentation, % DS 50.2±1.0a 38.9±1b 50.84±1.2a 21.3±0.6a 51.6±0.7a 21.5±0.9a PF 53.4±1.2a 39.1±1b 51.31±1.7a 26.3±0.6b 52.0±1.0c 29.5±1.1a Lu 79.2±1.2c 41.6±1.0c 78.66±2.2d 46.9±0.9d 77.7±2.1c 45.8±1.2c Ht 65.7±0.7 b 34.3±0.6a 57.32±1.0b 23.4±0.5a 63.7±1.5b 27.2±0.5a

Total titratable acidity after 48h of fermentation, °N DS 6.5±0.3b 5.7±0.1b 4.5±0.3a 3.3±0.7a 3.6±0.3a 2.2±0.1a PF 2.6±0.2a 2.3±0.1a 3.0±0.2a 2.6±0.6a 4.3±0.5a 4.1±0.4a Lu 13.8±0.5c 8.2±0.1c 10.0±0.3c 17.6±0.9d 4.8±0.2a 22.8±0.4e Ht 9.4±0.7b 5.0±0.1b 15.4±0.8d 5.4±0.4b 16.0±0.9d 6.0±0.5b

pH, after 24h of fermentation DS 6.11±0.2b 6.42±0.1b 6.23±0.7b 6.18±0.2b 6.46±0.5b 6.15±0.2b PF 4.78±0.3a 4.55±0.5a 4.66±0.4a 4.50±0.5a 4.70±0.5a 4.45±0.3a Lu 5.98±0.5b 6.00±0.1b 4.85±0.5a 6.04±0.7b 4.61±0.3a 5.83±0.1b Ht 4.30±0.2a 6.40±0.3b 4.5±0.1a 5.21±0.2a 4.57±0.3a 5.19±0.1a

pH, after 48h of fermentation DS 5.42±0.4b 5.59±0.5b 5.18±0.1b 5.94±0. 5b 5.40±0.5c 5.96±0.3c PF 4.65±0.3a 4.48±0.4a 4.72±0.5b 4.45±0. 4a 4.46±0.4b 4.40±0.2a Lu 4.45±0.3a 5.91±0.4b 4.39±0.2a 4.8±0.1a 4.41±0.5b 4.87±0.1a Ht 4.16±0.7a 6.18±0.3b 4.27±0.2a 6.19±0.4b 4.17±0.7a 6.1±0.5c

258

Continue of Table 3. P L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF

L(+)-lactic acid content, g/100g DS 6.74±0.1e 6.02±0.1b 6.74±0.2d 9.37±0.2d 7.85±0.2c 8.17±0.24d PF 5.53±0.4d 5.74±0.3a 6.12±0.7c 6.77±0.8c 8.07±0.9c 8.17±0.2 Lu 2.94±0.7c 5.93±0.1a 6.25±0.1c 6.25±0.1c 6.34±0.1b 3.08±0.1a Ht 0.18±0.3a 6.29±0.2c 3.79±0.1a 2.81±0.03a 4.11±0.1a 7.32±0.2c

D(-)-lactic acid content, g/100g DS 3.39±0.6b 5.09±0.1c 4.51±0.1d 4.82±0.3d 3.30±0.7b 7.05±0.2e PF 2.33±0.2a 2.17±0.2a 3.46±0.4c 2.71±0.3c 4.66±0.6c 5.02±0.7c Lu 4.73±0.9c 3.48±0.6b 4.33±0.9d 3.08±0.1c 3.17±0.1a 0.31±0.2a Ht 2.41±0.1a 3.08±0.4b 1.92±0.1a 0.18±0.2a 2.68±0.7a 4.51±0.3c Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; DS – defatted soy flour; PF – pea fiber; Lu –L. usitatissimum, Ht – H. tuberosus, P – plant product

Table 4. LAB count (log10 CFU/g) in SMF and SSF fermented defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke

P L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF

Lactic acid bacteria, log10 CFU/g DS 9.73±0.09d 8.60±0.09c 9.51±0.08d 9.89±0.10b 8.85±0.09c 9.98±0.11b PF 8.47±0.07c 9.52±0.10d 8.65±0.09c 9.39±0.09b 9.22±0.10d 9.36±0.10b Lu 7.92±0.09c 9.45±0.11d 8.72±0.09c 8.93±0.08a 9.11±0.10d 8.26±0.08a Ht 6.36±0.07a 6.65±0.08a 6.21±0.05a 8.66±0.09a 6.53±0.07a 8.80±0.09a Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; log10 CFU/g – colony-forming units per gram; DS – defatted soy flour; PF – pea fiber; Lu –L. usitatissimum, Ht – H. tuberosus, P – plant product

Table 5. Lignans (MAT and SECO) content (μg 100/g) in pea fiber

Samples NF SMF SSF Ls Pa Pp Ls Pa Pp

Matairesinol Pea fiber

72.6 ±2.1a

83.8 ±1.1b

97.2 ±1.5d

93.2 ±1.3c

88.3 ±1.2c

98.3 ±2.0d

78.3 ±1.4b

Secoisolariciresinol Pea fiber

62.0 ±2.4a

74.5 ±1.5c

121.2 ±2.4e

74.6 ±1.5c

88.1 ±1.8d

140.3 ±2.8f

87.2 ±1.7d

Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; Ls – L. sakei; Pa – P. acidilactici, Pp – P. pentosaceus; NF - nonfermented

259

Table 6. Proteolytic and amylolytic enzymes activity in defatted soy flour, pea fiber, flaxseed and Jerusalem artichoke (AU/g)

P L. sakei P. acidilactici 7 P. pentosaceus 8 SMF SSF SMF SSF SMF SSF

Amylolytic enzymes activity, AU/g DS 199.6±1.6a 715.8±3.5e 22.1±0.6a 152.2±1.3a 313.8±1.9b 183.9±1.9a PF 183.6±3.7a 195.8±14.3a 162.1±2.4d 132.2±2.6a 213.8±6.4a 193.4±3.9a Lu 269.3±1.5c 175.3±1.2a 132.2±1.8d 277.9±3.1c 235.6±2.5a 605.7±4.4e Ht 1281±5.7f 1075.0±4.7f 765.7±3.3f 390.6±3.5d 391.4±3.2b 442.3±2.9c

Proteolytic enzymes activity, AU/g DS 829.1±2.6c 723.6±4.2d 1069±4.9b 763.4±2.7d 1044±4.7b 709.4±3.5d PF 924.7±9.1d 874.3±6.2d 956.5±3.3a 861.7±3.1d 974.8±6.7a 898.6±4.9c Lu 603.6±4.7a 309.1±1.7a 927.3±4.4a 152.7±1.6a 1003±5.1b 181.8±1.8a Ht 1102±3.6f 1047±3.13f 1178±3.3c 887.3±3.8d 1095±4.2c 869.2±5.6c Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; AU/g – activity units per gram; DS – defatted soy flour; PF – pea fiber; Lu –L. usitatissimum, Ht – H. tuberosus, P – plant product

Table 7. ARs homologues content (μg/g) in pea fiber

Samples NF SMF SSF Ls Pa Pp Ls Pa Pp

C15:0

Pea fiber

200±0.6f 22±0.8d 25±0.3a 32±0.5b 22±0.4a 30±0.3b 24±0.9c C19:0

60±0.5c 55±0.3e 35±0.8b 25±0.3a 44±0.2d 51±0.6d 21±0.8c C21:0

3±0.6a 3±0.4a 42±0.6c 36±0.3b 27±0.2b 60±0.7e 18±0.4b C23:0

4±0.3a 18±0.4d 23±0.5a 21±0.6a 25±0.3a 20±0.2a 11±0.1a Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; Ls – L. sakei; Pa – P. acidilactici, Pp – P. pentosaceus; ARs – alkylresorcinols, NF – nonfermented

Table 8. Colour parameters (L*, a*, and b*) of pork loin marinated with Sm and Sh bioproducts

Meat samples

L. sakei P. acidilactici 7 P. pentosaceus 8 Control SMF SSF SMF SSF SMF SSF

S. montana L* 35.32

±0.28a 39.67 ±0.36b

37.78 ±0.39b

38.94 ±0.39b

35.40 ±0.40a

38.32 ±0.41b

47.41 ±0.47d

a* 7.42 ±0.08c

12.61 ±1.10e

10.62 ±0.09d

11.44 ±0.12d

7.93 ±0.08c

10.20 ±0.09d

4.62 ±0.06a

b* 9.21 ±0.08c

10.03 ±0.09d

8.61 ±0.07b

8.48 ±0.09b

7.34 ±0.07a

8.11 ±0.09b

9.13 ±0.10c

260

Continue of Table 8. Samples L. sakei P. acidilactici 7 P. pentosaceus 8 Control SMF SSF SMF SSF SMF SSF

S. hortensis L* 36.31

±0.39c 39.64

±0.25c 39.78

±0.30c 39.82

±0.37c 30.70 ±0.41a

29.44 ±0.18a

47.41 ±0.37e

a* 7.48 ±0.07c

9.78 ±0.09d

7.94 ±0.08c

7.19 ±0.08c

5.78 ±0.04b

5.73 ±0.05b

4.62 ±0.06a

b* 9.2 ±0.09c

10.22 ±0.12c

11.37 ±0.10d

10.80 ±0.13c

1.11 ±0.09a

2.97 ±0.04a

9.13 ±0.10c

Mean values within row with different letters are significantly different (p < 0.05). SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana; Sh – S. hortensis; L* − lightness; a* − redness; and b* − yellowness

Table 9. Parameters of pork loin marinated with Sm and Sh bioproducts

Meat samples

Moisture, %

pH DL, %

WHC, %

CL, %

SF, kg/cm2

IF, %

S. montana

Pa SMF 22.58 ±0.03a

6.0 ±0.05a

3.12 ±0.03c

65.31 ±0.61b

42.11 ±0.19c

3.16 ±0.06d

3.03 ±0.03a

Pa SSF 23.36 ±0.04b

6.27 ±0.03b

3.16 ±0.06c

67.52 ±0.60c

40.77 ±0.20b

1.77 ±0.02a

4.63 ±0.05c

Pp SMF 24.66 ±0.05b

5.98 ±0.02a

3.04 ±0.04b

68.61 ±0.45d

42.18 ±0.17c

2.05 ±0.02b

6.75 ±0.08d

Pp SSF 24.70 ±0.03b

5.97 ±0.06a

3.37 ±0.06d

66.76 ±0.51b

37.10 ±0.26a

2.45 ±0.02c

4.06 ±0.05b

Ls SMF 29.61 ±0.03c

5.86 ±0.07a

2.96 ±0.04b

63.58 ±0.61a

36.44 ±0.19a

2.14 ±0.03b

4.17 ±0.05b

Ls SSF 22.51 ±0.02a

6.59 ±0.08c

2.61 ±0.02a

67.14 ±0.23c

40.54 ±0.23b

2.03 ±0.04b

4.41 ±0.05c

S. hortensis

Pa SMF 22.25 ±0.04a

6.12 ±0.04a

3.01 ±0.03a

68.22 ±0.30d

34.65 ±0.17b

1.48 ±0.01c

4.19 ±0.04b

Pa SSF 22.23 ±0.05a

6.56 ±0.06b

2.98 ±0.03a

67.39 ±0.19c

40.99 ±0.18d

1.41 ±0.01c

3.17 ±0.03a

Pp SMF 26.08 ±0.03c

6.10 ±0.07a

3.15 ±0.04b

65.13 ±0.22a

34.21 ±0.22b

1.16 ±0.02a

4.92 ±0.06b

Pp SSF 25.20 ±0.03c

6.18 ±0.06a

3.21 ±0.05b

67.54 ±0.48c

34.50 ±0.19b

1.25 ±0.03b

4.64 ±0.03b

Ls SMF 24.71 ±0.03b

6.28 ±0.07a

2.93 ±0.03a

68.37 ±0.53d

32.56 ±0.16a

1.59 ±0.04c

4.07 ±0.04b

Ls SSF 21.62 ±0.02a

6.30 ±0.06b

3.03 ±0.03a

66.31 ±0.17b

36.54 ±0.26c

1.87 ±0.03d

5.01 ±0.06c

Control 22.65 ±0.02a

6.52 ±0.08c

3.06 ±0.02a

66.20 ±0.36b

39.45 ±0.18d

1.05 ±0.02a

5.86 ±0.08d

Mean values within column with different letters are significantly different (p < 0.05). Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; WHC – water holding capacity; IF – intramuscular fat; SMF – submerged, SSF – solid state fermentation; Sm – S. montana

261

Table 10. Colour parameters (L*, a* and b*) of beef loin marinated with Sm and Sh bioproducts

Meat samples

L. sakei P. acidilactici 7 P. pentosaceus 8 Control SMF SSF SMF SSF SSF SSF

S. montana L* 29.42

±0.28c 25.82 ±0.20a

27.56 ±0.25b

26.97 ±0.22a

25.49 ±0.21a

26.59 ±0.29a

38.03 ±0.36e

a* 14.65 ±0.16d

9.33 ±0.08b

9.02 ±0.08b

8.62 ±0.07a

7.90 ±0.09a

9.16 ±0.08b

15.16 ±0.15d

b* 3.51 ±0.06c

1.78 ±0.03b

1.38 ±0.02a

0.63 ±0.01a

1.40 ±0.02a

2.40 ±0.19b

2.25 ±0.25b

S. hortensis L* 25.82

±0.29b 25.34 ±0.30b

24.91 ±0.20a

27.29 ±0.26c

27.41 ±0.28c

26.09 ±0.30b

38.03 ±0.36e

a* 11.64 ±0.08c

7.03 ±0.06b

5.88 ±0.07a

11.97 ±0.13c

11.93 ±0.11c

11.37 ±0.12c

15.16 ±0.15d

b* 3.83 ±0.05c

1.42 ±0.02a

2.87 ±0.03b

4.28 ±0.05d

4.20 ±0.05d

4.23 ±0.04d

2.25 ±0.25b

Data are the mean ± SD (n = 3); SD – standard deviation; Mean values within row with different letters are significantly different (p < 0.05); SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana; Sh – S. hortensis; L* − lightness; a* − redness (or –a* of greenness); and b* − yellowness (or –b* of blueness)

Table 11. Colour parameters (L*, a* and b*) of pork meat, marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice

Sample L* a* b* Pork neck

Pa 52.45±0.22b 6.14±0.10b 15.97±0.16a Pp 52.50±0.21b 5.79±0.05a 16.67±0.19b Ls 53.04±0.22c 5.56±0.08a 15.68±0.18a Control 48.26±0.19a 5.23±0.08a 16.91±0.18b

Pork shoulder Pa 52.23±0.24b 13.73±0.14b 6.86±0.11a Pp 54.09±0.25c 14.01±0.13b 8.24±0.13b Ls 55.96±0.26d 13.63±0.12a 7.13±0.12a Control 50.37±0.22a 13.44±0.14a 9.04±0.11c

Pork ham muscle Pa 63.11±0.28b 10.92±0.12c 10.44±0.13a Pp 64.87±0.30c 9.56±0.08b 12.15±0.15c Ls 65.99±0.26d 8.97±0.09a 12.2±0.15c Control 62.47±0.27a 8.85±0.10a 13.97±0.16d

M. longissimus dorsi Pa 59.41±0.25a 4.72±0.09b 11.13±0.14a Pp 62.89±0.28c 5.94±0.06c 10.75±0.16a Ls 61.17±0.28b 4.66±0.05d 11.06±0.15a Control 57.78±0.25a 3.54±0.05a 11.25±0.15b

262

Continue of Table 11. Sample L* a* b*

Pork loin Pa 55.58±0.20c 7.00±0.10b 11.09±0.15b Pp 55.63±0.21c 6.87±0.09a 10.58±0.14a Ls 55.01±0.28c 6.69±0.08a 10.43±0.13a Control 45.63±0.19a 6.22±0.11a 12.30±0.15c Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05). Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; Sm – S. montana; Sh – S. hortensis; L* − lightness; a* − redness; and b* − yellowness

Table 12. The overall acceptability of pork meat, marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice

Overall acceptability scores (min 0 – max 6) Sample Neck Shoulder Ham Mld Pork loin

Pa 5.4±0.05c 4.9±0.04a 5.2±0.04a 5.5±0.06a 5.1±0.05b Ls 5.6±0.06d 4.8±0.04a 4.9±0.04a 5.4±0.06a 4.8±0.04a Pp 5.1±0.05c 5.3±0.06b 5.5±0.05b 5.8±0.07b 5.5±0.05c Control 4.5±0.03a 4.9±0.04a 5.1±0.05a 5.5±0.07a 5.1±0.05b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; M. longissimus dorsi – Mld

Table 13. Colour parameters (L*, a*, and b*) of organic and conventionally produced chicken meat marinated with P. acidilactici KTU05-7

Colour

Organic Conventionally Control Pa Control Pa

Breast

L* 47.3±0.14a 48.90±0.35b 51.6±0.23c 54.20±0.42d a* 14.03±0.26d 13.92±0.27c 12.87±0.34b 11.36±0.34a b* 13.98±0.11d 12.57±0.42c 11.82±0.21b 10.61±0.26a

Drumsticks

L* 46.5±0.38a 47.9±0.43b 58.3±0.31d 59.0±0.37d

a* 13.91±0.46d 13.07±0.54c 12.10±0.14b 11.07±0.42a b* 14.20±0.24d 13.86±0.27d 10.94±0.93b 9.41±0.24a

Thighs

L* 45.1±0.25a 45.90±0.44a 53.2±0.55b 55.10±0.43b

a* 14.31±0.37b 13.85±0.33b 11.01±0.27a 10.27±0.25a b* 13.54±0.42b 12.49±0.11b 11.63±0.14a 10.07±0.34a Mean values within row with different letters are significantly different (p < 0.05); Pa – P. acidilactici; L* − lightness; a* − redness; and b* − yellowness

263

Table 14. The overall acceptability of beef meat, marinated with P. acidilactici KTU05-7, P. pentosaceus KTU05-8 and L. sakei KTU05-6 strains cultivated in potato juice

Overall acceptability scores (min 0 – max 6) Samples Neck Shoulder Ham Mld Loin

Pa 5.6±0.04b 5.2±0.05b 5.4±0.05b 5.7±0.05b 5.6±0.06b Ls 5.2±0.03b 4.8±0.03a 5.3±0.04a 5.8±0.06b 5.1±0.04a Pp 4.9±0.03a 5.1±0.04b 5.4±0.04b 5.4±0.04a 5.3±0.05b Control 4.7±0.02a 4.8±0.04a 5.1±0.05a 5.4±0.03a 5.0±0.04a Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within column with different letters are significantly different (p < 0.05) Pa – P. acidilactici; Pp – P. pentosaceus; Ls – L. sakei; M. longissimus dorsi – Mld

Table 15. Facial expression intensity of cold smoked sausages sensory analysis

Facial expression

L. sakei P. acidilactici 7 P. pentosaceus 8 Control BS AS BS AS BS AS

Points 4.50

±0.01b 4.20

±0.01a 4.80

±0.03c 4.70

±0.02c 5.0

±0.05d 4.50

±0.04b 4.80

±0.05c

Happy 0.44

±0.001a 0.42

±0.05a 0.48

±0.003b 0.47

±0.002b 0.49

±0.004b 0.45

±0.004a 0.48

±0.003b

Sad 0.19

±0.001a 0.17

±0.001a 0.20

±0.002b 0.203

±0.003b 0.214

±0.001b 0.21

±0.001b 0.21

±0.002b

Angry 0.20

±0.001b 0.18

±0.001a 0.21

±0.002b 0.21

±0.002b 0.22

±0.002b 0.22

±0.002b 0.22

±0.002b Data are the mean ± SD (n = 3); SD – standard deviation. Mean values within row with different letters are significantly different (p < 0.05). LAB – lactic acid bacteria; Pa - P. acidilactici; Ls - L. sakei; Pp – P. pentosaceus; BS – cold smoked pork sausages treated with LAB before smoking; AS – cold smoked pork sausages treated with LAB after smoking

Table 16. The overall acceptability of RCMP produced with 3 %, 5 % and 7 % of biotreated Sm bioproducts

RCMP L. sakei P. acidilactici 7 P. pentosaceus 8 Control SMF SSF SMF SSF SMF SMF

Overall acceptability (scores: min 0 – max 6)

RCMP 3 % 5.0

±0.04b 6.0

±0.03c 5.5

±0.02b 6.0

±0.03b 5.5

±0.02b 6.0

±0.02c 4.0

±0.01a

RCMP 5 % 4.0

±0.02a 4.2

±0.03b 4.0

±0.03a 4.0

±0.02a 4.1

±0.03a 4.2

±0.03b 4.0

±0.02a

RCMP 7 % 1.0

±0.01a 1.0

±0.01a 0.9

±0.01a 1.0

±0.01a 0.8

±0.01a 0.9

±0.01a 4.0

±0.01c Data are the mean ± SD (n = 3); SD – standard deviation; Mean values within column with different letters are significantly different (p < 0.05); LAB – lactic acid bacteria; SMF – submerged fermentation; SSF – solid state fermentation; Sm – S. montana, RCMP - ready-to-cook minced pork meat products

264

Table 17. Microbiological parameters of RCMP produced with 3 % of Sm RCMP samples

Storage time, h 0 24 72 120

Total number of mesophilic bacteria, log10 CFU/g Control 5.21±0.32d 6.09±0.62f 6.25±0.72e 6.77±0.43d Ls SMF 4.81±0.82c 5.14±0.64c 5.33±0.54c 6.01±0.32c Ls SSF 4.63±0.62b 5.13±0.85c 5.27±0.22c 5.69±0.28b Pa SMF 4.65±0.41b 5.21±0.27d 5.42±0.27d 5.59±0.27b Pa SSF 3.62±0.21a 4.05±0.37a 4.18±0.25a 4.43±0.29a Pp SMF 4.91±0.61c 5.45±0.30e 5.69±0.31d 6.04±0.37c Pp SSF 4.59±0.42b 5.29±0.54c 5.38±0.45c 5.64±0.34b

Coliforms, log10 CFU/g Control 4.02±0.35c 4.30±0.42d 5.03±0.24d 6.83±0.58e Ls SMF 3.81±0.41b 4.00±0.33c 4.67±0.58c 4.95±0.37c Ls SSF 3.61±0.49b 3.79±0.91c 4.44±0.42b 4.60±0.42b Pa SMF 3.23±0.62a 3.39±0.87b 3.95±0.31a 4.20±0.30a Pa SSF 2.94±0.65a 3.19±0.59a 3.61±0.72a 3.82±0.44a Pp SMF 3.71±0.59b 3.90±0.58c 4.62±0.70c 4.82±0.24b Pp SSF 3.59±0.29b 3.77±0.33b 4.31±0.85b 4.67±0.26b

Yeast and mold, log10 CFU/g Control 2.92±0.44e 3.30±0.50d 3.92±0.45d 3.98±0.23d Ls SMF 1.81±0.38b 2.14±0.24b 2.31±0.32b 2.36±0.14b Ls SSF 1.61±0.18b 1.82±0.89b 2.02±0.29a 2.07±0.35a Pa SMF 1.23±0.33a

1.49±0.56a 2.15±0.30a 2.19±0.25a Pa SSF 1.54±0.24a

1.64±0.83a 1.84±0.24a 1.89±0.60a Pp SMF 2.11±0.49c 2.18±0.50b 2.43±0.27b 2.46±0.24b Pp SSF 1.88±0.25b 2.10±0.33b 2.12±0.21a 2.13±0.52a Data values are expressed as mean ± standard devation (n = 3). SD: standard deviation. Means within columns with different letters are significantly different (p ≤ 0.05). Control – RCMP without Satureja montana additives; Ls – L. sakei; Pa – P. acidilactici; Pp – P. pentosaceus; SMF – submerged fermentation and SSF – solid state fermentation.

Table 18. Quality parameters of RCMP produced with pea fiber and semolina

Meat sample Moisture, %

pH Drip loss, %

WHC, %

Cooking loss, %

Shear, kg/cm2

RCMP with pea fiber

37.06 ±0.04a

6.07 ±0.03a

1.96 ±0.15a

60.13 ±0.45b

24.46 ±0.22a

0.42 ±0.03a

RCMP with semolina

32.55 ±0.03a

6.01 ±0.04b

1.88 ±0.17b

65.97 ±0.53b

25.19 ±0.26a

0.25 ±0.01b

Data values are expressed as mean ± standard devation (n = 3); SD: standard deviation. Means within columns with different letters are significantly different (p ≤ 0.05). Remark: RCMP - ready-to-cook minced pork meat products; WHC - Water holding capacity

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Table 19. Colour parameters (L*, a* and b*) of RCMP produced with pea fiber and semolina Meat samples L* a* b* RCMP PF 65.84±1.60a 3.91±0.39b 15.09±0.24a RCMP S 70.61±2.17b 4.11±0.43b 15.17±0.17b

Data values are expressed as mean ± standard devation (n = 3); SD: standard deviation. Means within columns with different letters are significantly different (p ≤ 0.05). Remark: RCMP - ready-to-cook minced pork meat products; L* − lightness; a* − redness (or –a* of greenness); and b* − yellowness (or –b* of blueness); PF – pea fiber; S - semolina Table 20. BAs content (mg/kg) in RCMP produced with pea fiber and semolina

BAs PHE PUT CAD HIS TYR SPD SPR T

RCMP PF - 5.36

±0.06b 1.12

±0.10a 3.89

±0.13a 2.74

±0.20b 10.29 ±0.47c

18.66 ±0.53b

42.06

RCMP S - 7.81

±0.14c 0.70

±0.02b 4.71

±0.12b -

5.54 ±0.18b

15.2 ±0.62c

33.96

Data values are expressed as mean ± standard devation (n = 3); SD: standard deviation. Means within columns with different letters are significantly different (p ≤ 0.05). Remark: RCMP - ready-to-cook minced pork meat products; BAs – biogenic amines; T – total, PF – pea fiber; S - semolina

Table 21. The results of the sensory properties of UCC produced with Sm and Rc bioproducts determined in 5 point system

Plant L. sakei P. acidilactici 7 P. pentosaceus 8 UCC

UCC with NFP SMF SSF SMF SSF SMF SSF

Average of curd cheese acceptability S. montana 4.1b 4.7c 4.3b 4.8d 3.9a 4.5c 3.8a 4.1b R. carthamoides

4.3c 4.8d 4.0b 4.7d 4.3c 4.7d 3.8a 4.0b

SD: standard deviation. Means within columns with different letters are significantly different (p ≤ 0.05). Remark: SMF – traditional submerged fermentation; SSF – solid state fermentation; cfu - colony-forming units per gram, UCC – unripened curd cheese, NFP – nonfermented plant

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CURRICULUM VITAE

Name, Surname: Erika Mozūrienė Address: Department of Food Safety and Quality, Lithuanian

University of Health Sciences, Tilžės 18, LT-47181 Kaunas, Lithuania

Phone: +370 608 83450 E-mail: erika.mozuriene@lsmuni.lt Education: 2012–2016 PhD studies in Zootechics. Lithuanian University of

Health Sciences, Kaunas. 2009–2011 Master degree in Public Health. Lithuanian University

of Health Sciences, Kaunas. 2005–2009 Bachelor in Public Health. Lithuanian Veterinary

Academy, Kaunas. Professional Activity: from 2011–until now Study coordinator at the Lithuanian University of

Health Sciences.

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ACKNOWLEDGEMENTS

I would like to thank my supervisor Prof. Dr. Elena Bartkienė for her kind support and useful advices during the whole PhD process.

I would like to acknowledge for Prof. Habil. Dr. Audrius Maruška, Assoc. Prof. Dr. Ida Jākobsone, and Assoc. Prof. Dr. Vadims Bartkevics for sharing their scientific knowledge and experience.

Finally, I am thankfull for my family for being very supportive and encouraging.