the formation of off-characters in wine - ucanr

UC DAVIS VITICULTURE AND ENOLOGY

WF101: CURRENT ISSUES IN FERMENTATION MANAGEMENT

JULY 27TH, 2018

ANITA OBERHOLSTER

The formation of off-characters in wine


Off-characters in wine

• Off-flavors

• Off-colors

• Hazes or cloudiness

• Sediments or precipitates


Origin of Off-Characters

•Vineyard fruit and flora

•Winery flora

•Oxidation/reduction reactions

•Wine stabilization


Non-microbial off-characters coming from the vineyard•Eucalyptus

•Gives a eucalypt, fresh or minty character

to wine•Can be either pos or neg perceived

•1,8-cineole (Eucalyptol) is the aroma compound

•Distance from Eucalyptus trees very important

•High amount in grape skins, even more in stems and leaves

•Highest levels in Eucalypus leaves when included in ferments

•Rejection threshold is 27.5 ppb (ug/L)

•Aroma threshold – 1.1 ppb (ug/L)


Non-microbial off-characters coming from the vineyard•Brown marmorated stink bug

•Gives a green musty aroma

•Compound is trans-2-decenal•Detection threshold – 0.5 ppb (ug/L)

•Rejection threshold is 13.0 ppb (ug/L)

•3 bugs/cluster will have a negative impact

Mohekar et al. Am J Enol Vitic. 2016


• Best correlation with these 7 volatile phenols (GC-MS) to predict smoke exposure of grapes

Smoke compounds: volatile phenols

Guaiacol 4-methylguaiacol

Hayasaka et al, (2013) JAFC 61, 25‐33; Parker et al. (2012) JAFC 60, 2629‐2637; Pollnitz et al. (2004) JAFC 52, 3244‐3252; Wilkinson et al (2011) AJGWR 17, 522‐528.


Smoke taint compounds: volatiles and non-volatiles

Volatile phenols

Non-volatile phenols


Phenol-glycosides found in smoke affected grapes

Hayasaka et al. (2013) JAFC 61, 25‐33; Parker et al. (2012) JAFC 60, 2629‐2637;Noestheden et al, (2017) JAFC 65, 8418‐8425; Noestheden, (2018) JAFC 66, 695‐703; Pollnitz et al. (2004) JAFC 52, 3244‐3252; Wilkinson et al (2011) AJGWR 17,

• Phenolic glycosides by LC-MS/MS


Sensory threshold levels of VP’s

Parker et al. (2012) JAFC 62, 2629‐2637


Current recommendation

• No 100% fix for smoke taint

• Unpredictable due to precursors• Evolves during wine aging

• Actions that can minimize impact• Less skin contact – change wine style

• Cool fermentations to limit extraction

• Fruity yeast

• Wood contact to add complexity

• Reduce smoke-taint compounds• Fining and reverse osmosis treatment


What does the numbers mean• We do not know

• Due to varied amount of smoke-exposure, varieties, analysis methods and sensory evaluations

• Need to build a library of smoke-related compound composition, matrix and smoke taint evaluation


What does the numbers mean• Best guess

• Even a zero amount of free VP’s does not guarantee no smoke taint due to precursors

• Free in smoke-free grapes: nd-3.4 μg/kg

• Bound in smoke-free grapes: 4.1-10 μg/kg

• Free in smoked grapes: nd-30 μg/kg

• Bound in smoked grapes: 35.8 -2233 μg/kg


(Non)-microbial off-characters: winery

•Oxidative aromas•Reduction of varietal character; sherry, apple, green, vegetal/grassy, nutty/almond, pungent

•Mostly due to formation of acetaldehyde•Sensory threshold 100-125 mg/L

•>125 mg/L bruised apple, nutty and sherry aromas



•Acetaldehyde formation•Under oxidative conditions yeast can oxidize EtOH to CH3CHO (acetaldehyde)

•Note under these conditions acetic acid and ethyl acetate may also be produced

•Chemical oxidation of EtOH with wine aging

•Often via Acetobacter or film yeast

•Laccase activity – enzymatic oxidation


Yeast aromas – good or bad?

• Grape-derived – provide varietal distinction √

• Yeast and fermentation-derived• Sulfur compounds • Esters √ mostly• Higher alcohols high conc• Carbonyls high conc• Volatile acids • Volatile phenols



•Reductive aromas•Match stick, burnt rubber, rotten egg aromas

•Usually due to sulfur compounds

• Produced by yeast• Degradation of sulfur-containing amino acids

• Degradation of sulfur-containing pesticides

• Release and/or metabolism of grape-derived sulfur-containing precursors


Sulfur compounds

Swiegers et al., (2005) Austr. J. Grape Wine Res. 11: 139-173

Compound Conc in wine (g/L)

Aroma threshold (g/L)

Aroma Descriptor

Hydrogen sulfide Trace80 10-80 Rotten egg

Methanethiol(methyl mercaptan)

5.1, 2.1 0.3 Cooked cabbage,putrefaction, onion, rubber

Ethanethiol(ethyl mercaptan)

1.9-18.7 1.1 Onion, rubber, natural gas

Dimethyl Sulfide 1.4-61.9 25 Asparagus, corn, molasses


Hydrogen sulfide• Amount produced varies with:

•Amount of sulfur compounds available (inorganic sulfur in vineyard, use of pesticides and fungicides)

•Yeast strain

•Fermentation conditions• Rate of fermentation

• Use of SO2

• Presence of metal ions

•Nutrient status of environment• Vitamin deficiency

• Level of total nitrogen


Timing of Formation of H2SBr

ix

Time

H2S


Thiols (mercaptans)

• Mercaptans such as ethanethiol can be formed by reaction of H2S with EtOH or CH3CHO

• Formation of sulfides such as DMS (dimethylsulfide, asparagus, corn, molasses) not clear

• Yeast or SO2 can reduce disulfides to thiols (mercaptans) such as ethane- and methanethiol – can be removed by copper

methanethiol ethanethiol


Microbial off-characters from the vineyard and winery

•Vineyard flora

•Winery flora

•Fermentation microbes•Saccharomyces yeast

•Non-Saccharomyces yeast

•Spoilage microbes•Acetic acid bacteria

•Lactic acid bacteria

•Brettanomyces


Microbial off-characters from the vineyard and winery

•Low fermentation temp and low/late sulfite additions favor non-Saccharomyces yeasts and bacteria

•Warm fermentation temp favors bacteria

•O2 exposure favors non-Sacch and bacteria

Oenococcus oeniSaccharomyces cerevisiae Brettanomyces bruxellensis


• Produced mainly by yeast (through lipid and acetyl-CoA metabolism)•Variable amounts, mixed strains higher levels

of esters compared to fermentations with Saccharomyces cerevisiae

•Also variety depended

•Some esters produced by yeast from specific grape precursors

Esters


Swiegers et al., 2005 Austr. J. Grape Wine Res. 11: 139-173

Compound Wine (mg/L)

Threshold(mg/L)

Aroma descriptor

Ethyl acetate 22.5-63.5 7.5* Fruity, VA, nail polish

Isoamyl acetate 0.1-3.4 0.03* Banana, pear

Isobutyl acetate 0.01-1.6 1.6*** Banana, fruity

2-Phenylethyl acetate

0-18.5 0.25* Flowery, rose, fruity,

Hexyl acetate 0-4.8 0.07** Sweet, perfumeEthyl butanoate 0.01-1.8 0.02* Floral, fruity

Ethyl hexanoate 0.03-3.4 0.05* Green apple

Ethyl octanoate 0.05-3.8 0.02* Sweet soap

Ethyl decanoate 0-2.1 0.2**** Floral, soap

*10% ethanol, **wine, ***beer, ****synthetic wine


Ethyl Acetate

• Most prominent ester is ethyl acetate• Acetic acid + ethanol = ethyl acetate

• Small quantity produced by yeast during fermentation• Depended on strain, temp of fermentation, amino

nitrogen content of juice and SO2 levels

• Larger amounts produced by aerobic acetic acid bacteria during barrel aging

• Some possibly produced by lactic acid bacteria


Ethyl Acetate

• Aroma: nail polish, solvent, glue• Aroma threshold 7.5 – 12 mg/L

• Low levels, fruity aromas

• Wine normal 22.5-63.5 mg/L, spoiled 150 mg/L


Higher alcoholsCompound Wine

(mg/L)Threshold (mg/L)

Aroma descriptor

Propanol 9-68 500** Fruity, sweet, pungent, harsh

2-methylpropanol 25.8-110 4 Fruity, wine-like

Butanol .5-8.5 150* Fusel, spirituous

Isobutanol 9-174 40* Fusel, spirituous

Isoamyl alcohol 6-490 30* Harsh, nail polish

Hexanol 0.3-12 4* Green, grass

2-Phenylethyl alcohol

4-197 10* Floral, rose

*10% ethanol, **wine


Fusel/higher alcohols

• 300 mg/L add complexity (fruity characteristics)

• 400 mg/L (strong, pungent smell and taste)

• Different yeast strains contribute variable amounts of fusel alcohols• Non-Saccharomyces yeast – higher levels of

fusel alcohols


Fusel alcohols

• Conc fusel alcohols produced:• Amount of precursor - amino acids

From Linda Bisson: The Fusel Family

Ehrlich Pathway


Fusel alcohols

• Conc fusel alcohols produced:• Amount of precursor - amino acids

• EtOH conc, fermentation temp, pH, must composition, amount of solids, skin contact time etc. influence conc of higher alcohols

From Linda Bisson: The Fusel Family


Carbonyl compounds• Acetaldehyde (bruised apple, sherry,

nutty)• Sensory threshold of 40-100 mg/L, typical

conc. in wine 10-75 mg/L

• Major intermediate in yeast fermentation

• Increase with fermentation temp

• Increase over time due to oxidation of EtOH -due to aeration


Volatile acids• Acetic acid (>90%)

•High conc. vinegar-like aroma

•Fault 0.7-1.5 g/L depending on wine style

•Production by Saccharomyces cerevisiaestrains varies widely 0.1-2 g/L

•However, commercially used strains produce less than native strains

•Odor threshold 0.1 – 0.125 g/L


Volatile acids (VA)• Acetic acid

• Excess conc. largely the result of metabolism of EtOH by aerobic acetic acid bacteria by converting EtOH to acetic acid

• LAB in presence of glucose can produce acetic acid

• During oxidation - formed acetaldehyde can be oxidized to acetic acid


Volatile Phenols• Produced from hydroxycinnamic acid

precursors in the grape must

-CO2 Reduced4-ethylphenol4-vinylphenol

4-ethyl-guaiacol4-vinyl-guaiacolferulic acid

p-coumaric acidO H

OOH

O H

O

OH O

O H

O

O H O H

O H

O


Volatile Phenols

• Trace amounts present in grapes

• Mostly produced during fermentation from precursors during fermentation• Saccharomyces cerevisiae

• 4-ethylphenol (medicinal, barnyard)• 4-ethylguaiacol (phenolic, sweet)• 4-vinyl phenol (phamaceutical)• 4-vinylguaiacol (clove-like phenolic)

Present below threshold values

Main contributor


Volatile Phenols

• Brettanomyces/Dekkera spp. • Produce high conc of 4EP, 4EG, 4EC,

regarded as spoilage organisms

• Band-aid, medicinal, pharmaceutical, barnyard-like, horsey, sweaty, leathery, mouse urine, wet dog, smoky, spicy, cheesy, rancid, metallic

• Brett – spoilage or complexity?


Brettanomyces/Dekkera spp. • Brett is not a fermentation problem but

sanitation problem in cellar/air and barrel

• Grows under difficult conditions• Tolerant of 10% EtOH

• 34% isolates grew well at pH 2

• 30% of isolates grew at 10C• 35% of isolates grew at 37 C• 10% of isolates grew at both temp

extremes

• 50% show tolerance to =>30 ppm free SO2 at pH 3.4 Lucy Joseph, culture collection, UCD


Aroma thresholds

Figure 1. Aroma thresholds of 4-ethyl guaiacol (4-EG), 4-ethyl phenol (4-EP, and 4-ethyl catechol (4-EC) in three different Australian Cabernet Sauvignon wines.www.awri.com .au


Lactic vs Brett

Lucy Joseph, culture collection, UCD

Lactic Acid Bacteria Sensory Brettanomyces SensoryVinegar, pungent, sour Vinegar, pungent, sour

Nail polish remover Chemical, solventButter, nutty, caramel Resin, woody

Geranium leaves Floral, lilac, + rancidMousy MousyBitter Metallic, bitter

Ropy, viscous, oily

Viscous, sweet

Fecal Barnyard, Sewage


Lactic acid bacteria found in wine

• Where do they come from?

• Population established in winery – difficult to remove

• Vineyards may be reservoirs for some species


Spoilage compounds produced by LacticsBacteria Compound Sensory Effect Threshold

LAB Acetic Acid Vinegar, pungent, sour 0.2 ppt

LAB Ethyl acetate Nail polish remover 7.5 ppm

Lb., Oeno. Diacetyl Butter, nutty, caramel 0.1 to 2 ppm

Lb., Pd. 2-Ethoxy-3,5-hexadiene Geranium leaves 0.1 ppb

Lb., Oeno. 2-Acetyl-tetrahydropyridine

Mousy 4 to 5 ppb

Lb., Oeno. 2-Ethyltetrahydropyridine Mousy 2 to 18 ppb

Lb., Oeno. 2-Acetyl-1-pyrroline Mousy 7 to 8 ppb

Lb., Pd. Acrolein (+anthocyanin) Bitter

Pd. b-D-Glucan Ropy, viscous, oily

Oeno. Mannitol Viscous, sweet

LAB Skatole (indole) Fecal 1.7 ppm (1.8)

LAB Biogenic Amines None (headache)

Letters in Applied Microbiology 48 (2009) 149–156 ; E.J. Bartowsky


Mousy off-flavor

• Off-flavour reminiscent of caged mice • Perceived late on the palate – non-volatile at

wine pH

• 3 known compounds causes mousy aroma

• Lactic acid bacteria (LAB) can produce all 3 compounds

• Dekkera/Brettanomyces can produce 2

ACPY

ETPY

ACTPY


Mousy off-flavor

• 2-ethyltetrahydropyridine (ETPY)• Threshold 150 g/L, up to 162 g/L can be produced by

LAB

• 2-acetyltetrahydropyridine (ACTPY)• Threshold 1.6 g/L, isolated in wine at levels of 4.8-106 g/L

• 2-acetylpyrroline (ACPY)• Threshold 0.1 g/L, detected in wine in trace – 7.8 g/L

amounts

2-acetylpyrroline

NO

2-ethyltetrahydropyridine 2-acetyltetrahydropyridine


Mousy off-flavor

• Following needed for mousy-flavorproduction• L-Lysine, L-ornithine

• Responsible for ring formations of 3 mousy heterocycles

• EtOH and acetaldehyde• Responsible for the acetyl side chain

Snowdown et al. (2006) J. Agric. Food Chem. 54: 6465-6474

2-acetylpyrroline

NO

2-ethyltetrahydropyridine 2-acetyltetrahydropyridine


Musty taints

• What off-odours are classified as musty taints?• Fungal, earthy, moldy, corky, mushroom or

straight musty

• What causes musty taints?• Haloanisoles (TCA, TBA)

• Alkylmethoxypyrazine (MDMP)

• Carbon unsaturated aliphatic compounds with carbonyl function (1-octen-3-one, 1-nonen-3-one)

• (-)-Geosmin


Musty taints: Cork taint

• Trichloroanisole (TCA) responsible for 80-85% of cork taint• Haloanisoles (TCA, TBA) ‘Musty’, ‘mouldy’,

‘wet cardboard’ character

• Generally detection threshold given as 1.4-4.6 ng/L

Sefton and Simpson, Austr. J. Grape Wine Res. 2005, 11, 226-240.


Musty taints: Cork taint• 2,4,6-trichlorophenols (TCP) produced from

naturally-occuring phenols and chlorine from sanitizers, cleaning products and town water

• Microorganisms capable of methylation are present in cork, form TCA from TCP

• Decline of TCA in corks• Suggest depletion of source of contamination

• Improved treatment of corks

• Improved testing methods

Sefton and Simpson, Austr. J. Grape Wine Res. 2005, 11, 226-240.


• Main off-flavors

• Ethyl acetate, H2S and ethanethiol, acetaldehyde, VA, volatile phenols, mousy

• Most off-flavors can be minimized or prevented by• Using clean fruit• Sufficient nutrient, oxygen and temperature

control during fermentation• Good winery sanitation and adequate SO2

use

Concluding remarks


Thank you

the formation of off-characters in wine - ucanr

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