August 28, 2023 knowledge Nate Ferguson

Decoding Ester Production in Yeast Fermentation

Esters are complicated - but YOU can figure them out!

The goal of this article is to provide you with the resources and information so that you can make enriched decisions when trying to influence ester production in yeast fermentation.

This is a long article and will be broken down into a few sections

General knowledge & summary
Acetate esters
Ethyl or Fatty Acid (FA) Esters
How to control ester profile
Edge cases and oddities

If you don't want to read the whole thing - that's okay. Go to the section that is for you!

Happy learning!

1. General Knowledge & Summary

Esters are a type of chemical compound naturally produced during fermentation that are generally considered to be positive in the finished beer. Esters are produced in yeast cells by reacting alcohol (fusel alcohol or ethanol) with an acid (acetyl-CoA or acyl-CoA). Esters produced by yeast can be placed into two groups; acetate esters (fusel alcohol and acetyl-CoA/acetic acid) or ethyl esters (ethanol and acyl-CoA). Controlling esters becomes tricky as many different elements will alter the concentrations of the precursors needed to produce these compounds; I will link to those knowledge sources throughout.

There are other sources of esters from your ingredients - mainly bound esters in hops and fruits - but those are not what we are referring to today; see BIOTRANSFORMATION RESOURCES.

Esters are commonly misunderstood

Brewers will often make changes to their process in an attempt to increase esters, only to see them reduce or worse, produce off-flavours.

Ester production is multivariable and some strains act very differently.

This document is meant as a guide, not as a bible. Some strains will deviate from what we discuss below simply due to their own genetics.

Why do yeast cells produce esters?

It is commonly believed that yeast do so for 2 reasons:

Esters attract insects that in the wild are crucial for yeast to move from food source to food source
Esters allow for the breakdown of toxic fusel alcohols and aldehydes

How are Esters formed?

In most yeast strains, esters are produced within the cytoplasm of the cell where they passively diffuse (leave) across the membrane and into the beer. This is true for both of the two groups of esters; Acetate Esters (Fusel Alcohol Esters) & Ethyl Esters (Fatty Acid and ethanol Esters). In both cases, adding more oxygen will enhance both, however, how we control the other elements will influence what flavour the yeast produces.

Of these two, acetate esters are the most common in most standard fermentations, however, some strains are able to produce more or less ethyl esters.

(Reminder, This is a long article) - As promised, here is the summary.

Summary:

The following practices consistently enhance esters:

Increased wort aeration
- Increased wort aeration not leading to increased esters is normally a sign of another issue
- BEST PRACTICES - OXYGENATION
Proper mineral content (Zinc and Magnesium) will enhance esters formation (or remove their inhibition)

The following will increase esters, but... will also likely increase off-flavours, which can be dealt with but requires more attention and care.

Increased FAN
Lower pitch rate
Lower viability yeast culture (not recommended)
High Gravity Fermentation
Repitching yeast (increase with repitch - assumes proper nutrition)

The following variably increase ester content

Fermentation temperature (high or low temp OR swings in fermentation temperature)
Simple sugars (dextrose or glucose additions)

The following consistently decrease esters

Increased CO2 content during fermentation (tall vessels or pressurized fermentation)
Low O2 content (also cause more fusels and aldehydes)
Higher pitch rate

If you want to get to the nerdy stuff, keep scrolling!

2. Acetate Esters

Common acetate Esters:

Isoamyl acetate: Banana (Hefeweizen)
Isobutyl acetate: General fruity (English)
2-phenyl ethyl Acetate: Floral (Lagers)
Ethyl acetate: Pear/Solvent (Belgians and others)

Acetate esters are formed through the esterification of a Fusel or higher alcohol and Acetyl-CoA OR Acetic Acid; see “TROUBLESHOOTING - FUSEL ALCOHOLS” to learn more about Fusel Alcohols. Whether or not Acetyl-CoA or Acetic Acid is used will depend on many things mainly the genetics of the strain. Whether Acetyl-CoA or Acetic Acid - both methods will produce the same ester, however, the enzymes and chemistry are different.

These esters are our classic fermentation esters.

To make more of anything, we need more of the ingredients or precursors. For Acetate Esters that is Acetyl-CoA and Higher Alcohols. If we can increase both within the cell, we will yield more esters.

How to increase Acetyl-CoA:

Acetyl-CoA is an essential element of not just yeast cell metabolism but for humans too(that’s you….I hope) where in both, it is primarily bound to pyruvate targeting carbon to the TCA cycle for aerobic respiration. For this reason, we generally see the molecule become more active during aerobic conditions. Acetyl-CoA is produced through a process known as Beta-Oxidation where fatty acids (oils) are broken down. If the cell has access to more fatty acids from either the wort or it has created them itself, then it will have more acetyl-CoA to react with any fusel alcohols present. The process of beta-oxidation and esterification has many enzymes that are catalyzed with Magnesium and Zinc which when supplied at appropriate levels, ensures metabolism occurs smoothly and esters are generated.

How to increase Fusel Alcohols

I do not recommend you try and increase Fusel Alcohols!

There are likely more than enough fusel alcohols below the flavour threshold (you can’t taste them) present in your beer which esters to be produced from. Focus on increasing Acetyl-CoA!

If you do want to try - see our article on “TROUBLESHOOTING - FUSEL ALCOHOLS” and do the opposite.

If we increase Acetyl-CoA and Fusels, we should get more acetate esters. These esters generally taste fruity to floral and form the backbone of what most people think of when we hear about a yeast strain being estery. Different strains will have different concentrations of each ester depending on the amount of and what type of fusel alcohol they produce (which is directly related to their Amino Acid needs as a cell).

How to increase Acetate Esters:

More Oxygen = More Acetate Esters
Ensuring proper Magnesium and zinc content of the wort
Glucose/Dextrose: The mechanism is unknown but anecdotally shown to do so; I do this too

3. Ethyl or Fatty Acid (FA) Esters

Common Ethyl Esters:

Ethyl Hexanoate: Aniseed or apple
Ethyl Octanoate: Sour Apple
Ethyl Hexanoate: Pineapple

Ethyl Esters are a different class of esters that are not commonly talked about, but, can play a large role in fermentation flavour. Similar to Acetate esters, if we have more precursors then we will have more final products, which in this case are esters.

The difference is what reacts

Where Acetate esters have Fusel alcohols and Acetyl-CoA, Ethyl Esters have Ethanol (instead of fusel alcohols) and Acyl-CoA which is the bonding of Acetyl-CoA to a Fatty Acid. The different fatty acids found in the cell are what contribute to the different esters we see produced this way.

So - what are fatty acids?

Fatty acids are mainly produced in the cytoplasm of the cells and are mainly used within the membrane of the cell. The cell can consume fatty acids from their environment, however, studies have not shown these fatty acids to not be directly active when it comes to ester production; likely indirectly affect it but that is out of scope for this post. Fatty acids are mainly synthesized from acetyl-CoA (it's kind of a big deal) along with a few other molecules that are used to produce the needed fatty acids. Each strain will produce different fatty acids based on its genetics which will alter the availability of fatty acids will be available for ethyl ester production. When it comes to altering ethyl esters, the biggest thing we can do is to make FA synthesis occur as smoothly and easily as possible.

How do we increase Fatty Acid Synthesis?

Simple.

Oxygen. Zinc. Magnesium. Other trace nutrients and generally Healthy cells

How to increase Ethyl Esters:

Ample Acetyl-CoA access (healthy cells and lots of oxygen).
Proper mineral concentrations (Zinc, magnesium, manganese, Copper, and Iron are important in decreasing priority).
Free phosphate appears to be important to Ethyl esters but the mechanisms are unknown - acid rests might increase ethyl esters.
Keeping trub in the wort might increase Ethyl Esters due to increased available Fatty Acids - only anecdotal evidence.

4. How to Control Ester profile:

When dialling in an Ester profile I always recommend the following approach:

FIRST, ensure you are providing sufficient oxygen:

Acetate Esters come from Acetyl CoA and Higher alcohols being fused together. If insufficient Oxygen is present, insufficient amounts of Acetyl CoA will be produced resulting in a beer that has esters but also tastes hot or burning due to the fusel alcohols not being degraded.
Ensuring proper aeration is achieved removes this issue.

Ensure proper trace nutrient levels:

Adding yeast nutrients will help ensure cells have available energy to produce Acetyl-CoA.
if any nutrients are insufficient, amino acids or other essential molecules required for ester production will not be produced reducing esters.

Increase your fermentation temperature to increase Esters:

If you do this and fusel alcohols increase, this likely means you do not have enough oxygen
- This might be due to the oxygen becoming less soluble at higher temperatures
- See our BEST PRACTICES - OXYGENATION for more information

Assess the amount of FAN present in your wort

FAN is required for fusel alcohols and therefore Acetate Esters. Higher FAN levels will typically generate more fusels which if you have proper wort aeration will be converted to acetate esters
If you add more FAN and you generate a hot fermentation flavour then you do not have sufficient wort aeration

Other things that impact Ester profile

Simple sugar content has been shown to increase esters
Esters tend to increase with re-pitching

5. Edge case & Oddities:

Edge Cases

I added more oxygen and my perceivable ester concentration decreased - What is going on?

This is caused by one of two things

1. The “esters” you tasted before were not actually esters.

If this occurs, this is typically a sign that you were not initially providing enough oxygen to your cells in general
The “Esters” you were detecting before were likely aldehydes or fusels
- The cells now as sufficient energy to break down aldehydes - but not enough to produce the ester

2. More diversity of esters, but a lower concentration of each resulting in less flavour.

Each ester has a “Detection Threshold”: the concentration of the substance needed in order for you to taste the substance
This could also be the case of multiple esters now being produced with higher wort aeration
- If you add more oxygen, you may shift one or more esters below the detection threshold making the beer cleaner (I.E. instead of one compound at 10, you now have 2 compounds at 5 and the detection limit for both is 7. End result is a cleaner beer despite the total amount of esters being the same or more)

Oddities:

Hefeweizens:

Fermenting colder has been anecdotally described to increase banana ester (acetate ester)
The banana ester is isoamyl acetate which is the fusion of isoamyl alcohol and acetic acid

Things you can try - but there isn't much evidence to them yet

Acid rests increase the amount of free inorganic phosphates which have been shown to increase ethyl esters due to the increase in fatty acid synthesis.

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