General Trends Vintage 2012
• Ideal Growing Season
• Record Harvest – High Quality Fruit
• Low YAN Compared to Previous Harvests
• More AF Issues than MLF
YAN and Fermentation
• YAN values dropped from ~205 to ~145
• Nitrogen/Nutrient Supplementation– DAP alone– Complex Nutrients– Organic Nutrients– Yeast Rehydration Survival Factors
• No Pattern to Stuck and Sluggish Ferments
Yeast Nutrition
AssimilableNitrogen
Vitamins
Minerals
The most important and indispensable factors for a good fermentation and aromatic quality of the wines
Yeast Rehydration
Survival Factors
Where to Look ?
• Poor Fermentation Causes – No Shortage!– Low Population of Viable Yeast– Fermentation Temperature Spikes – Heat, Cold– Microbial Competition – Early, Late– Toxins – Microbial, Al, Pb, Pesticide Residues– Yeast Genetic Mechanisms – Prions - Epigenetics– Nutrition
• C / N Balance - Brix / YAN• Macro nutrient shortage• Micro nutrient shortage• Trace nutrient shortage
Yeast Nutrition
Yeast Growth Factors
Nitrogen YAN = Ammonia + alpha amino acids Vitamins
8 B complex vitamins - B1 – thiamine B2 – riboflavin B3 – niacin B5 – pantothenic acid B6 – pyridoxine B7 – biotin B9 – folic acid B12 - cobalamin Minerals
8 Major Minerals - Ca, Co, Cu, Fe, Mn, Mo, Ni, Zn
All factors are important but…
Supplement additions are usually based on YAN alone
So…
Balancing Nitogen does not always balance total nutrition
K Mg
Macro nutrients >100 ppm
Micro nutrients <1 ppm Trace Nutrients – Se, B, Na, Inositol, ???
! !
Unhappy Yeast Consequences
• Stuck or Sluggish Fermentations
• Elevated C8 + C10 Fatty Acids
• High SO2
• H2S – Other Aroma Consequences
Factors Influencing MLF
• Ethanol• pH• FSO2• TSO2• Temperature• Malate or < ~1 g/L• Polyphenolsundetermined• Nutrient Status may be proportional to initial YAN• AF stuck or sluggish• Microbial Status < 103 – 104 cells/mL• Fatty Acids C8 + C10 concentration
Many of the values depend upon the
specific Oenococcus strain being used
C8+C10 FA World Survey
Countries C8 (mg/L) C10 (mg/L)
England 61.5±27.4 14.5±6.0
Portugal 34.8±18.6 8.7±5.6
Switzerland 24.8±18.7 7.8±1.8
Italy 24.4±14.2 7.5±3.5
Germany 23.9±8.5 6.4±2.7
France 21.9±12.9 4.5±1.0
China 18.0±8.7 1.1±0.2
USA 12.2±7.0 2.8±1.4
Spain 11.4±2.3 2.5±0.5
Chile 8.1±5.8 1.9±0.8
South Africa 3.9±1.2 1.9±0.5
Average levels of fatty acids measured post- AF according to the country of origin. 282 wines, 3 vintages, red and white, North and South Hemisphere.
C8+C10 FA vs MLF
0
10
20
30
40
50
C8 C10
C8 a
nd C
10 co
ncen
tratio
ns (m
g/L)
Figure 2: Of the 282 wines studied: 156 presented no problems with MFL (group I in green) and showed average levels of C8 (9.0 ± 5.0) and C10 (2.1 ± 1.1) relatively low. The 126 wines in which MLF was problematic (group II in pink) showed significantly higher average levels of C8 (34.4 ± 7.4) and C10 (9.3 ± 2.5).
C8+C10 FA vs MLF
0
0,5
1
1,5
2
2,5
0 10 20 30 40 50
Days after inoculation
L-m
alic
aci
d (g
/L)
MLF monitored in a wine with C8 =5 mg/L and C10=2 mg/L* (♦), in the same wine with C8 =20 mg/L and C10=4 mg/L (●) and in the same wine with C8 =50 mg/L and C10=20 mg/L (■). Trials were made in triplicate. Malolactic starter is inoculated at the initial population of 106 cell/mL at day 0.
C8+C10 FA California 2011
• 225 random fermentation samples from across California
• Less than 5% above 15 mg/L C8+C10 FA
• All others below levels of concern
• Two problem sluggish MLF tested ~22 mg/L C8+C10 FA
• The problem MLF above were ~30 FA including C6
• Medium Chain FA assay at ETS Labs
C8+C10 FA vs MLF
Country Wine without MLF issues Wines with MLF issues
Number of wines analyzed
s C8 (mg/L) C10 (mg/L) Number of
wines analyzed C8 (mg/L) C10 (mg/L) France 48 12.9±2.1 2.7±0.6 44 37.3±9.7 10.6±5.7 Italy 22 11.2±1.2 3.4±0.4 22 38.4±8.4 12.6±6.6 South Africa 17 3.5±1.6 1.7±0.5 15 11.1±5.4 2.8±0.3 Australia 16 2.9±1.5 1.0±0.4 12 12.3±3.2 3.6±2.2
? MLF issues are not solely due to C8 +C10 Fatty Acid levels
? Native MLF by endogenous strains have varying tolerance for C8 +C10 Fatty Acid levels
? Undiscovered factors may reduce tolerance of C8 +C10 Fatty Acid levels
YAN Effect on FA
0
10
20
30
40
50
60
75 125 175 225 275
Available nitrogen (mg/L)
C8 (m
g/L)
at
the
end
of t
he A
F
Phytosanitary Treatments and Effect on MLF
• Bordeaux 2010 vintage was very dry at end of season
• Washout by rain was minimal
• Led to persistence of abnormally high residues in must– Folpet residue
• Two phytosanitary compounds were >100 ug/L– Boscalid– Dimethomorph
Folpet Effect on C8+C10 FA
C8+C10 = f(Folpet µg/L)
0
10
20
30
40
50
60
0 20 40 60 80 100 120 140 160 Folpet (µg/L)
C8+C10 (mg/L)
Folpet is a protective leaf fungicide
Indirect effect on MLF through elevated FA
>25 mg/L C8+C10 FA can inhibit MLF
Boscalid vs MLF
Parameters Values Temperature 20°C pH 3,7 ASV (% vol.) 12,4 Total SO2 (mg/L) 22 L-malic acid 1,8 Fatty acids (octanoic acid + decanoic acid in mg/L) 8 Brettanomyces (cells/mL using quantitative PCR analysis) <1/10 mL Lactic bacteria (cells/mL using epifluorescence) 2.106 cell/mL Boscalid (µg/L) 262
Chemical Name: 3-pyridinecarboxamide, 2-chloro-N-(4’chloro[1,1’biphenyl]-2-yl) Common Name: Boscalid (BAS 510) Trade Names: Emerald, Endura, and Pristine Chemical Class: Carboxamide aka anilide
Dimethomorph vs MLF
Tank Dimethomorph (µ g/L)
Number of days before MLF completion (from bacteria inoculation carried out 24h afterwards
each time)
Octanoic and decanoic acids (mg/L) at the end of
AF I Not detected 12 7 II Not detected 14 14 III Not detected 11 12 IV 92 54 8 V Not detected 10 5 VI 191,5 42 6 VII Not detected 18 12
Dimethomorph is a systemic fungicide which protects plants from molds, as well as killing molds on plants and preventing their spread. It is a cinnamic acid derivative and a member of the morpholine chemical family.
Dealing with Difficult MLF
PREVENTIVE
Correct Nitrogen Deficiencies in Must/Juice
Rehydrate Yeast with a Nutrient Containing Survival Factors
PROACTIVE
Utilize Co-Inocculation Strategy – Early (24 hr after yeast) or Late (~5 brix left)
After Determination of Elevated Levels of C8+C10 FA – Test Stuck/Sluggish for FA - Use FA Resistant Oenococcus Strain for MLF
REMEDIAL
Detoxify Wine by Yeast Hull FA Adsorption – MLF Restart Protocol
Co-Inocculation vs Sequential
Type of inocculation
Step Objective
Early co-inocculation
24-48 h after AF start-up
Time gain and fault preventionOptimized fermentation management
Late co-inocculation
At 3-5 brix remaining
Monopolization of the ecosystem and fault prevention. Secures a traditional vinification (AF then MLF)
Sequential inocculation
After AF,at pressing
MLF after post AF macerationMLF in barrels
Spring inocculation and/or restart
Later…Remedial
Restarting stuck MLFSpring MLF
Choosing the type of inoculation to implement:
Co-Inocculation – For You?
Pre-fermentative maceration
Alcoholicfermentation
MalolacticFermentation
Sequential inoculation
Late co-inoculation
Early co-inoculation
• Lower Stress on Oenococcus at Inocculation
• No latency between AF and MLF – control indigenous microflora
• Enococcus, a facultative heterofermenter, does not produce Acetic Acid from 6 carbon sugars-Glucose/Fructose
• Biogenic Amines – Strain Specific – Genetic Screening
Thank You!
Peter Salamone, PhDTechnical Manager, NA
Laffort USA
1460 Cader Lane, Suite CPetaluma, CA 94054
(707 [email protected]