Key points on MLF

Question 1

11 factors contributing to MLF

Answer 1

1. pH (most important)
2. S02
3. Nutrient Composition
4. Oxygen
5. CO2
6. Alcohol
7. Temperature
8. Organic Acids
9. Phenolics or phenolic acids
10. Presence of other LAB
11. Bacteriophage

Question 2

MLF inhibited by (7)

Answer 2

1. Use of SO2 (should be used after)
2. Early racking (it likes solids)
3. Downward pH adjustment
4. Low temp
5. Filtration/Fining
6. Addition of Fumaric acid (only outside EU)
7. Bacteriocin (lysozyme) addition (egg whites and are the best additive to prevent MLF)

Question 3

How to inhibit MLF (4)

Answer 3

1. Add SO2
   - 0.8 mg/L molecular pH adjusted
2. Temperature
   - Below 18°C and as low as 13°C
3. High ethanol does not always inhibit because often coupled with high pH
4. Acidity
   - Adding tartaric acid (better than malic acid) will help but may not stop it
   - Wines pH 3.3 and below need to be stimulated while wines greater than 3.3 will be difficult to stop

Question 4

Inhibition 2 (3)

Answer 4

1. Elimination of viable bacteria
2. Sterile filtration and sterile bottling
3. Chemical inhibitors

Question 5

Lysozyme

Answer 5

Egg whites that are used as an antimicrobial agent. When we use this we need to avoid adding close to bottling it is a protein and is capable of combining tannins

Question 6

Importance of Lysozyme

Answer 6

It is included among wine additives able to control fermentation processes and avoid spoilage during wine making. Because it is an animal product its not vegan.

Question 7

Amount of Lysozyme added

Answer 7

The amount of lysozyme added normally ranges between 25 and 50 g/hL.

Question 8

Organic acids that can be added (3)

Answer 8

1. Fumarate inhibitory at low concentrations
2. Fatty acids (C8 and C10 released by yeasts)
   can also be inhibitory
3. Malate stimulates growth prior to malolactic fermentation

Question 9

Oxygen (3)

Answer 9

1. Stimulatory to growth
2. Affects spectrum of end products
3. Can produce more energy (and acetic acid) in presence of oxygen (TCA cycle, via citrate a/o by pentose phosphate pathway –> Eterof. LAB)

Question 10

Carbon Dioxide (2)

Answer 10

1. Stimulatory to malolatic fermentation
2. Mechanism unknown

Question 11

Bacteriophage

Answer 11

1. Bacterial “viruses” that can be spread from one bacterium to another and that cause cell death
2. Not known if this is a problem in wine production or not; it is a problem in other lactic acid bacteria fermentations

Question 12

Presence of Other Lactic Acid Bacteria positive

Answer 12

1. Mixed cultures may yield “better” complexity
2. Can be stimulatory
   - Increase in pH (Difficult to start MLF below 3.3-3.4 pH)

Question 13

Presence of Other Lactic Acid Bacteria Negative

Answer 13

Can be inhibitory
- Bacteriocin production (Lysozyme)
- Competition for nutrients

Question 14

Nutrient Composition (4)

Answer 14

1. Lactic acid bacteria are fastidious: numerous growth requirements
2. Aging on yeast lees increases micronutrient content via autolysis
3. Extended skin contact enhances lactic acid bacteria
4. Higher solids or a less clarification degree enhance lactic acid bacteria

Remember Red wines have more nutirents because they offer more solids

Question 15

Yeast and Bacteria interaction (Metabolization)

Answer 15

Yeast and bacteria are competing for nutrients
( Yeast are mainly interested in sugars
and are able to metabolize amino acids)

Question 16

Yeast and Bacteria interaction (bacteria growth)

Answer 16

Yeast can both inhibit and stimulate bacterial growth

Question 17

What inhibits bacterial growth

Answer 17

Formation of ethanol, bisulfite and fatty acid by yeast

Question 18

What else stimulates bacterial growth

Answer 18

SO2 binding and the products of yeast autolysis after “sur lies” aging

Question 19

Death Phase

Answer 19

Bacteria can stimulate death phase in yeast

Question 20

Choice of strain of Bacteria and yeast (5)

Answer 20

1. compatibility
2. Production of desirable characteristics
3. Ability to complete MLF
4. Vigor
5. Availability of freeze dried inoculum

Question 21

Inoculated MLF (4)

Answer 21

1. Better control over both timing and organisms present
2. Difficult to maintain inocula
3. Starter culture must be “pure”
4. Percent inoculation: 1-50% depending upon vigor of culture

Question 22

Inoculum prep (4)

Answer 22

1. Start culture from a medium supporting good growth of organism
2. Inoculate “diluted” juice (with water) with starter and addition of nutrients
3. Use #2 to inoculate full strength wine or juice with addition of nutrients
4. Use #3 to inoculate the rest of wine

Question 23

Spontaneous MLF (3)

Answer 23

1. Uncontrolled timing of process
2. Risk of unwanted species/strains
3. Off-characters can be produced if MLF when undesired

Question 24

Timing of MLF (4 options)

Answer 24

1. Prior to yeast fermentation
2. Simultaneous with yeast fermentation
3. Mid-way through yeast fermentation
4. After yeast fermentation

Question 25

Pre Ferm Inoculum (2)

Answer 25

1. Decreases yeast nutrients
   - Stuck/sluggish fermentation
   - Production of off-characters
2. May lead to production of inhibitory compounds (acetic acid) due to presence of oxygen

Question 26

Simultaneous yeast inoculation (3)

Answer 26

1. See increase in acetic acid
2. See a decrease in viability of both yeast and bacteria
3. Yeast “rebound” better than bacteria

Question 27

Mid Ferm inoculation (4)

Answer 27

1. Nutrients left for bacteria
2. Ethanol low and not inhibitory
3. Yeast-produced SO2 may be inhibitory
4. May lead to arrest of yeast fermentation

Question 28

Post Ferm Inoculation (4)

Answer 28

1. Nutrients have been depleted
2. Ethanol concentration high
3. Concentration of other yeast inhibitory compounds also high
4. Better temperature control