Download - Bio preservation of Meat and Meat products
BIO-PRESERVATION OF MEAT AND MEAT PRODUCTS
G.SUNDARESANMVM 15026
Credit seminar
Department of Livestock Products Technology (Meat Science), Madras Veterinary College, Chennai – 600 007.
IntroductionMeat is highly desirable, nutritious and rich in protein,
highly perishable.
Ancient preservation- physical treatment and application of preservatives.
(e.g) Heat treatment and application of salt.
(Talukder et al. 2014)
Major concerns of meat preservation
Food Quality
Making food desirable to eat-
Good taste
Color
Texture (Nath et al., 2006)
Food Safety
Infectious agents Toxic chemicals Foreign objects
Drying, Curing, Smoking, Canning, Chilling, Freezing, Direct microbial control
The modern physical and chemical methods of food
preservation are more effective than the traditional methods,
with drawbacks.
Chemical preservatives - Toxicity
Change organoleptic and nutritional properties
It made research interest on the natural and effective preservatives
Existing Preservative Techniques
Bio preservation – an alternative
Fermentation, bacteriocins, bacteriophages and bacteriophage-encoded
enzymes fall in this concept.
(Elsser et al.,2013)
Meat preservation
Extending the shelf life of the food by
continuous fight against the spoilage organism.
Meat Bio-preservation
Extending the shelf life and food safety by natural
or controlled microbiota or their antimicrobial
components.
Definitions
Principles of bio preservation
Hindering the growth activity of the microorganisms.
e.g. pH
Killing the microorganism.
e.g. Bacteriocins
Anti-microbial metabolites
Organic acidsLactic acid Acetic acidPropionic acid
Bacteriocins Class I Class II Class III
Molecular compounds Hydrogen peroxide Carbon dioxide Diacetyl Peptides
Mechanism of action of anti-microbial metabolites
Organic acids:
By lowering pH.
OxidasesHydrogen and Lactoperoxidase
• H2O2 producing reactions lower the
oxygen.
• Oxidize sulfhydryl groups of cell
proteins and membrane lipids.
+
Diacetyl• Interfering with the utilization
of amino acids
VARIOUS METHODS OF BIO PRESERVATION
1. Fermentation
2. Bacteriocin application
3. Bacteriophages
4. Endolysins
Fermentation
Fermentation is a metabolic process that converts sugar to acids, gases, or alcohol. It occurs in yeast and bacteria.
Definition:
This process based on the growth of micro-organism in foods
it may be natural or added.
Mainly comprised of lactic acid bacteria.
Will produce the organic acid and other compounds.
This compounds exhibits the preservative action.
Fermentation of meat by protective cultures
Selection of Cultures
• Generally Recognised As Safe (GRAS)
• Ability to produce antimicrobials in meats
• Limited sensory changes
• Limited acid production
• Weak protease activity
• Limited gas production
Starter culture for meat fermentation
Meat starter cultures are preparations that contain active or
dormant microorganisms that develop the desired metabolic
activity in the meat (Hammes, Bantleon, and Min 1990).
General procedure for the preparation of fermented meat products
Fermentation in meat and meat products
Preparation of meat emulsion
Shift it in casing at -2.2
to 1.1° C
Addition of starter culture
Fermentation-lactic acid
Cooking or drying
Upstream bioprocessing Inoculum development media development Enrichment
Downstream bioprocessing Separation of biomass Semi concentration Purification Laboratory and field trial
fermented meat products
Effects of culture on the meat & meat products
Fermented sausagesAddition of curing agent and sugar and
fermentation condition preventing the growth of pathogens in fermented sausages (Lucke, 1998 b).
Raw ham and Ready to eatInjection of Psychotropic lactic acid bacteria even under the
aerobic condition observed approximately 2 log reduction of Listeria monocytogenes during the Vacuum packaged conditions.
Semi processed raw meatStrains of lactic acid bacteria that improves the self-life and freshness
of refrigerated semi-processed meat such as bacon and sausages.
Salted-Semi processed raw meatPediococcus starter grow and form the acids. Thus restricting the growth
of any clostridium botulinum and extend the shelf-life.
Probiotics
"live micro-organisms which, when administered in adequate amounts, confer a health benefit on the host". (WHO 2001) e.g. Lactobacillus plantarum
Definition
Probiotics are poor survivors in meat – Challenging environment .
Micro-encapsulation may be an option for formulation of fermented meat products with viable health-promoting bacteria (Muthukumarasamy and Holley 2006).
Safety issues of fermented meat and meat products
I. Biogenic amines CadaverinePutrescineTyramine
Toxicological symptom
Other symptom
1. Pseudo allergic reaction2. Interaction with drugs
1. Gastric ulcers2. Blood pressure problems3. Nervous symptoms
II. Antibiotic resistance
1. Gene translocation
Suzzi and Gardini, 2003
Bacteriocins
Ribosomally synthesized extracellularly released bioactive peptides or peptide complexes, having bactericidal or bacteriostatic activity.
Rapidly digested by proteases in human digestive tract.
This bacteriocins are first discovered in 1925 by Andre Gratia
Alternative to antibiotics and chemical preservatives. (Joerger et al., 2000).
Bacteriocins vs antibiotics
s.no Characteristic Antibiotics Bacteriocins 1 Synthesis Secondary
metabolite Ribosomal
2 Application Clinical Food
3 Activity Varying spectrum Narrow spectrum
4 Host cell immunity No Yes
5 Mode of actionCell membrane or intracellular targets
Mostly pore formation, but in a few cases possibly cell wall biosynthesis
6Toxicity side effect
Yes None known
ADVANTAGES OF BACTERIOCINS
BACTERIOCINS
Reduce the use of
chemical preservatives
Better preservation
Decrease the risk of disease
transmission
Extra protection
It satisfy the consumer demand
FACTORS PROMOTING USE
OF BACTERIOCINS ASBIOPRESERVATIVES
Not alteringNutritionalquality
Effective under wide pH &
temperature range
Activity is not lostPresence of other
preservatives Effective in low concentrations
Broad spectrum of activity from food grade LAB
Preference over traditional preservative
Safe (FDA) use of nisin for > 40 years
in several countries
Bio synthesis of Bacteriocins
Nisin A
Regulatory Gene
Cyclize &Dehydrydrate
Pre peptide
ABC Transpoter
Cellular space Proteases
Active form of Bacteriocin
Extra cellular space
Lipoprotein & Transporter
complex
Class IClass IIClass III
Classification of Bacteriocins
Class I Bacteriocins or Lantibiotics
• Small peptides (<5kD) • More than 50 amino acids• Made up of unusual amino acids (lanthionine, and
methyl-lanthionine)
Lantibiotics
Class I a Class I b
Linear Cationic and hydrophobic peptides
Globular No net charge
Cleveland et al., 2001
Nisin Mersacidin
Mechanism of action of Class I Bacteriocins
Class II bacteriocins or non Lantibiotics
1. Class IIa or pediocin-like bacteriocins2. Class IIb or two-peptides bacteriocins3. Class IIc4. Class IId
Cationic Hydrophobic Heat-stable Membrane active peptides
Klenhammer, 1993
(e.g) Enterocin NKR-5-3C (Ent53C).
Ent53C showed very strong microbial activity (in Nano molar range) against Listeria spp.
Class IIa Bacteriocins (Pediocin like bacteriocins) - anti-listerial bacteriocins
Mechanism of action
Enterocin
Class IIb bacteriocins (Two-peptide bactetiocins)
Killing activity of this bacteriocin is based on the synergistic activity of both the peptide.
1. Type E (enhanced)
1. Type S (synergistic)
Action mechanism
(e.g.) Acidocin L
Class-IIc - Circular bacteriocins
Unique structural feature of a head-to-tail cyclization of their backbones.
structural stability, higher thermal stress resistance, and superior stability against proteolytic digestion, compared to their linear counterparts.
However, the biosynthetic mechanisms currently remains unknown.
(e.g.) Lactococcin A
class IId – Non pediocin like bacteriocins - Leaderless bacteriocins
Diversity of primary structure. (e.g.) Lacticin Q
Action mechanism
The highly cationic lacticin Q molecules rapidly bind with negatively charged phospholipid bilayer membrane.
After which, the lacticin Q molecular mass translocate itself from the outer to inner membrane as the pore closes
Class III Bacteriocins or Bacteriolysins
Large > 15kDa
Heat-labile antimicrobial proteins
Have a domain-type structure
Different domains have different
functions for translocation,
receptor binding, and bactericidal
activity
Anti-microbial action
Production of Bacteriocins
Methods of bacteriocin application in meat and meat products
I. Inoculation of meat with Lactic acid producing bacteria
- starter or protective culture
- Bacteriocins
II. Purified or semi purified bacteriocins
III. Use of previously fermented meat with bacteriocin producing strains
(Thomas et al., 2000)
Bacteriocin producing strains
Purified bacteriocins
VI. Edible cellulosic films - New
Application in meat products
Several bacteriocins from the bacteriocin producing strains used as a food preservatives.
Still very limited.
Bacteriocins like Pediocin has very strong anti-listerial activity, But they are not currently approved.
NISIN is the only approved Bacteriocin by FAO/WHO 1969. (Devlieghere, et al., 2004)
Application in meat products
The most-studied bacteriocins in the
meat and meat products is -
Nisin
Enterocin AS_48
Enterocin A and B
Pediocin PA-I
(Saeed et al., 2014)
Use of Purified/ Semi purified bacteriocins
To date, the only commercially produced bacteriocins are:
Nisin produced by Lactoccocus lactis ssp. Lactis – Rogers and Whittier 1928.
Pediocin PA-1, produced by Pediococcus acidilactici (Not approved)
(Saeed et al., 2014)
The samples of raw mince meat were inoculated
with 103 CFU/g of Listeria monocytogenes and
stored at 40 c.
After 16 days, the control sample increased count
from 3log10 to 6.4log10 CFU/g.
Nisin – 2.4 log 10 CFU/g.
Raw mince meat
(Pawar et al., 2000)
Beef carcass
Beef carcass were inoculated with approximately 4 log 10
CFU/cm2 of Brochothrix thermosphatca to evaluate the
effectiveness of nisin to sanitize the carcasses.
Nisin spray treatment 1000 IU/ml reduced the population at 0 day – 1.8 log 10 CFU/cm2
at 1 day - 3.6 log 10 CFU/cm2
( Deegan et al., 2006)
Bacteriocin producing culture addition:
(Degnan et al., 1992) Demonstrated the possibility of using bacteriocin producing culture LAB and Pediococcus acidilactici (nisin and pediocin) to control the L. monocytogenes growth in the all vacuum packed Beef products
Combination with MAP (Fang and Lin, 1994) Was found to be more effective
when used in combination with modified atmosphere packaging (100% co2, 80% co2+20%o2)
Examples of microorganisms that may be controlled by bio preservation techniques
Commercial status
Subtilin, Cerein, Plantaricin, have been isolated and characterized from different bacteriocin producing strains.
But they are yet to acquire a commercial status.
Nisin (Lactobacillus lactis) and Pediocin (Pediococci acidilactici) are the only commercially.
(Nes et al., 2007)
Commercially approved bacteriocinNisin – E234 (Nisaplin)
Bacteriophages Bacterial virus that infect the bacteria and reproduce by
hijacking their host biosynthetic pathway
They are harmless to humans, animals and plants.
I. Classification II. Mechanism of action
Bacteriophage application in meat
Application in meat industry
FDA - Recently permit safe use of a bacteriophage preparation as an anti-listerial agent in Ready To Eat (RTE) meat and poultry products.
Phage (lytic) preparation with 0.1ppm concentration is sprayed directly on meat products prior to packaging at the level of 1 milliliter (ml)/500 cm2 surface area.
Bacteriophages will remain dormant unless their specific target, L. monocytogenes is encountered.
(Kathy walker, 2006).
Phage (lytic)
Lysins, or endolysins , are hydrolytic enzymes produced by bacteriophages.
It cleave the host's cell wall during the final stage of the lytic cycle.
Endolysins - Degrading the peptidoglycan of Gram positive bacteria when applied externally to the bacterial cell, thereby acting as antibacterial agents.
ENDOLYSINS
(Jhamb and Spardha, 2014)
Endolysins mechanism of action
Schematic representation of the modular structure
Mode of action of phage-encoded - endolysins.
Endolysin application
Broad killing spectrum by cleaving peptidoglycan linkage of bacterial membrane.
It exhibits the antimicrobial activity at Nano gram level .
At the same time production cost is very high, because use of genetically modified organism for the production.
MERITS AND DEMERITS OF BIO PRESERVATION
s.no MERITS DEMERITS
1 Narrow activity of spectrum
2 Limited diffusion in solid matrix
3 Sometime difficult to apply
4 Prevent the colorectal cancer Consumers are reluctant to use of meat preserved using micro-organisms
5 R
6
Bio Prn
Extended shelf-life of the meat and meat product
Decrease the of food borne pathogens transmission
Reduce use of chemical preservatives
Resistant organisms are controlled effectively (e.g) L.monocytogenes
Minimal handling and Natural way
Lack of research focus
Application deficit
Commercialization
Regulatory approval
MERITS DEMERITS
Current status
Nowadays researchers are focusing on the application of bacteriocins in foods as part of packaging films.
The potential applications of bacteriocins in the food and health care sectors are evident.
(Cagri et al., 2004)
Research focus
Commercial application and for production in large scale, both genetic and fermentative protocols need to be optimized.
E. coli has long been considered the primary prokaryotic host for cloning
(Billman Jacobe, 1996)
Alternative food-grade organisms must be Identified,
Researchers are focusing on the application of bacteriocins in foods as part of packaging films.
(Coma, 2008).
Conclusion
Extend the shelf life and food safety of meat and meat products by the use of natural or controlled microbiota and antimicrobial compounds.
This preservation can be effectively used - hurdles.
Technology can be integrated as part of preservation technology
THANK YOU