Impact of Malolactic
Fermentation Strain on Wine
Composition
Lucy Joseph
U.C. Davis Department of Viticulture
and Enology
Outline
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Introduction to malolactic fermentation (MLF)
Lactic acid bacteria metabolism
Commercial inoculum
Wine matrix effects
– Interaction with oak
• Timing of inoculation
Malolactic Fermentation
• Any wine containing malic acid could be
considered unstable.
• Certain indigenous bacteria can metabolize malic
acid as a very poor carbon and energy source in
the fermenter and in the bottle.
• Conversion of malic to lactic acid by a controlled
malolactic fermentation prior to bottling
eliminates the instability.
• It can be a problem to start, very slow activity,
long time to finish and can start and stop.
MALOLACTIC BACTERIA
• What are they?
– “bacteria”: single-celled non-nucleated
microorganisms
– “lactic acid bacteria”: produce lactic acid from
sugar
• Acid loving
• Nutritional fastidious
• Carry out many food fermentations
Malolactic bacteria are lactic acid bacteria
which can convert malic acid to lactic acid:
CO2
Malic Acid
Lactic Acid + Carbon Dioxide
stronger acid
weaker acid
2 carboxyl groups
1 carboxyl group
Oenococcus oeni
• Only two species
• Oenoccocus oeni (formerly
Leuconostoc oenos) are only
found in wine
• Oenococcus kitaharae was
‘discovered’ in 2006 in the spoiled
remains of a sake mash (shochu,
high pH) and lacks the gene for
malolactic fermentation
What happens during a
malolactic fermentation
• Deacidification
– Each gram per liter of malic converted to lactic
creates a loss of 7.46 mM/L of titratable H+ ions, or
1.12 grams/L as tartaric measured by titration (TA)
• pH changes
• Micronutrients are sequestered
• Secondary metabolites can contribute to the
flavor profile
THE COURSE OF THE MLF
Bacterial growth is finished several days
after the conversion of malate to lactate.
Full bacteria growth in wine is only 107
cells/mL, and the malate has usually
disappeared at 106 cells/mL. (Diacetyl may
be being formed at that time.)
Bacteria
Oenococcus
oeni
Positives
Negatives
Increase in volatile acidity
Reduction of total acidity
(high pH and residual sugar)
Production of biogenic amines
Increase microbial stability
and ethyl carbamate
Reduction of ketone and aldehyde Spoilage aromas (mousy, sweat,
equivalents (reduces SO2 use)
sulfur)
Loss of varietal aromas and
Reduction of grassy, vegetative notes
fruity esters
Enhanced mouthfeel
Increase of diacetyl and other aroma
and flavor compounds
Out-competes other bacteria
Enhanced color stability
(co-pigmentation)
Lactobacillus
plantarum
Excess diacetyl production
Reduction in total acidity
Loss of color (high pH)
Sensitive to low pH and high
alcohol
No acetic acid production
Sluggish fermentation
Production of spoilage aromas
Relevant Metabolic Activity in
Oenococcus oeni
ML Metabolism
Buttery
Character
Diacetyl
Acetaldehyde Conversion
J.P. Osborne et al. /FEMS Microbiology Letters 191 (2000)
Mousy Character
Commercial Malolactic Strains
Oenococcus oeni
Lactobacillus plantarum
Commercial Strains - Inoculation
• Direct inoculation
Bacteria is pre-adapted and can be added
directly to the fermentation
• One step strains
Bacteria need to be rehydrated and grown for
24 hours prior to addition
• Traditional
Requires growth and build up of inoculum
prior to addition
Commercial Strains-Selection
Criteria
 pH tolerance
 Alcohol tolerance
 SO2 tolerance
 Temperature range
 Competitive ability
 Stuck MLF
 Biogenic amine production
Commercial Strains-Sensory
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Diacetyl production
Color stability
Mouthfeel
Varietal enhancement
Avoidance of defects, i.e. vegetative, sulfur
Interaction with oak
Wine Matrix Effects
Cultivar & Strain Influence
Compounds found in MLF wines by
GC-MS
Purge and trap system of Montrachet wines
•4-Methyl-3-pentenoic acid
•Methyl acetate (Sweet, solvent-like)
•Ethyl hexanoate (Fruity, rum-like)
•Hexyl acetate (Fruity)
Freon 114 extraction of Epernay 2 wines
•1,12-Tridecadiene*
•Hexadecanoic acid (mild waxy)*
•1,2-Benzene dicarboxylic acid (mild ester)*
•Farnesol (floral)*
*Spectral fit < 900
Am. J. Enol. Vitic., Vol. 43, No. 3, 1992
R. M. AVEDOVECH, M. R. McDANIEL, B. T. WATSON, and W. E. SANDINE
Other Reported Flavor Enhancers
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1-hexanol - fruity
ethyl acetate - fruity
ethyl lactate - buttery
diethyl succinate - brandy
butyrolactone – aroma enhancer
glycoaldehyde – aroma complexity,
browning
MLF in Oak
Bacteria can breakdown glycosides in solution to
release the aglycone
Breakdown of Glycosides
Glycosidic activities for a selection of Oenococcus oeni strains on four substrates: p-nitrophenyl-β-Dglucoside, p-nitrophenyl-α-L-arabinofuranoside, p-nitrophenyl-α-L-rhamnopyranoside, p-nitrophenylβ-D-xyloside
Tannat Wines- Different MLF-Before Aging
Control (gray filled square), MLF with DSM 7008 (black filled square)and D-11 (open square)
J. Agric. Food Chem. 2009, 57, 6271–6278. E. Boido, K Medina, L. Farina, F. Carrau, G. Versini, E. Dellacassa
Timing of Inoculation
Inoculation
Advantage
Disadvantage
Prefermentation
MLF completion
Reduced nutrients for AF
Production of yeast inhibitors such
as acetic acid
EarlyFermentation
Simple-shorter production time
Increased acetic acid production
Incompatibility of yeast and
bacteria
MidFermentation
Postfermentation
Tends to avoid MLF failure
Allows optimization of management
throughout the fermentation
Better domination of the MLF by inoculated
strain
More traditional, allowing AF to complete
before MLF completion
MLF occurs after AF is complete allowing
better control of temperature and SO2 levels
MLF can be done in barrels
Less color loss
Incompatibility issues
Some compatibility issues
Nutrients are depleted
Inhibitors may be high i.e. alcohol
Summary
• Reduction in acid
• Production of desirable compounds (diacetyl)
• Production of other flavor compounds during
growth (1-hexanol, ethyl acetate, ethyl lactate, diethyl
succinate, butyrolactone, glycoaldehyde, glyoxal, 2,3butanediol, caprylic acid, hydroxycinnamic acid)
• Release of aglycones from glycosides in the
wine