Sep 21, 2015: Ester Production in Fermentation
Think of the last saison you tasted: did you notice any banana or clove flavors? These aren’t there because the beer was fermented over bananas and cloves but because of compounds produced by yeast during fermentation. The spicy clove notes come from phenols, which are spicy flavored compounds containing a phenyl ring and a hydroxyl (OH) group. The fruity flavors, such as banana in the case of saisons, come from compounds called esters. Esters are naturally occurring compounds that show up in fruits, flowers, and fermentation vessels, along with a myriad of other places. In low concentrations esters impart fruity character to the beer they are in, but at too high concentrations they can taste like solvents, especially ethyl acetate. Ester compounds are defined by an ester group that connects two organic molecules. The connecting ester group is made up of a carbon bonded to two oxygen atoms, one by a double bond. A visualization of esters can be seen in Figure 1.
Esters are widely considered some of the most important flavor compounds in beer, and there are a wide variety of these esters. Most have fruity flavors like apple, pear, or banana. Each ester’s flavor is different and most have different flavor thresholds. Ethyl acetate is the most abundant ester that occurs in beer. It also has the highest flavor threshold at 33 ppm (most have a flavor threshold 10x less than this). A table of esters and their associated flavors can be seen below.
|Compound||Flavor Notes||Flavor Threshold||Structure|
|Ethyl acetate||Nail polish remover, fruity, solvent-y||33 ppm|
|Methyl butyrate||Apples, pineapples||43 ppb|
|Iso-butyl acetate||Bananas, floral, fruity||64 ppb|
|Ethyl butyrate||Tropical fruits, pineapple, juicy fruit||400 ppb|
|Iso-amyl acetate||Sweet, bananas, pear drops, circus peanuts||1.6 ppm|
|Ethyl hexanoate||Apples, pears, anise||225 ppb|
|Ethyl octanoate||Apples||200 ppb|
|2-Phenylethyl acetate||Roses, honey||200 ppb|
Esters are produced in two ways in beer, both of which start with an alcohol and a carboxylic acid and end with an ester. The first way is by spontaneous chemical reactions called condensation reactions. The other method is by enzymatic processes carried out by yeast during fermentation. The condensation that results from reacting a carboxylic acid with an alcohol produces an ester and water (see Figure 2). This process is slow and would not produce enough esters to affect the flavor of the beer. Cellular processes within the yeast are what create most of the esters that are tasted in beer. The reactions start a fatty acid reacting with Adenosine Tri-Phosphate (ATP) and a coenzyme ASH (CoASH), to form acyl CoASH. Acyl CoASH molecules then interact with ester forming enzymes to combine with alcohols (mostly ethanol) and produce esters.
After esters have been made by yeast, they start to be broken down by enzymes called esterases. This process happens at a slower rate than the creation of esters, thus the concentration of esters does not decrease because of it. Particularly, of note is that acetate esters are broken down at a much slower rate than other esters. This rate difference, in combination with ethanol being the most common alcohol reacted, explains why ethyl acetate is the most abundant ester found in beers.
The formation of esters has a huge impact on beer flavor. Not only do esters result in fruity flavor notes, but their formation also reduces the amount of off-flavored compounds such as certain fatty acids. Many short chain fatty acids like butyric acid, octanoic acid, etc. impose rancid, sweaty, or animal-like flavors and aromas to the beer. The fruity notes resulting from the formation of esters make for a rather agreeable tradeoff.
Many factors influence ester production during fermentation, including yeast strain and pitching rate, oxygen and free amino nitrogen levels in wort, temperature, and what organic acids and alcohols are present. The first factor is yeast strain. There are many different kinds of yeasts in several different genera (more than 500 species). Only a limited number of these yeast, though, are used for fermentation due to other fermentation byproducts they may create. The most common strains of yeast used are Saccharomyces cerevisiae and Saccharomyces pastorianus (lager yeast). There are a few other species of yeast that are also used, but those two make up the bulk of brewing yeast. Ale yeasts usually produce more esters than lager yeasts meaning there are more fruity and floral flavors in ales than there are in lagers. There are however different amounts and ratios of esters produced by each different strain of yeast within a species. These different ratios are the result of the yeasts making different kinds of organic acids and alcohols to react and create esters. The yeast will make esters out of whatever acids and alcohols are available. However some yeast produce different acids preferentially. All this means that the final flavor of the beer is strongly dictated by the strain of yeast selected for fermentation.
The amount of yeast added to a fermenter also dictates ester production. This is due to the fact that yeast produce esters mostly during their growth and reproduction phase. After the yeast grow a sizeable colony, they start converting sugar into alcohol and CO2 and ester production slows significantly. Pitching large amounts of yeast will ensure a healthy population of yeast later on and helps start a strong fermentation, but the growth and reproduction stage is significantly shortened and far less esters are produced. The obvious answer is to pitch less yeast. Under-pitching will lead to much higher rates of ester production but yeast health can suffer and the fermentation may be slow or stop completely. These are problems that are worse than not having enough esters. The trick is to find a happy medium. Make sure there is enough yeast pitched to have a strong healthy fermentations and a small enough amount of yeast to produce more esters.
One of the easiest conditions to control during fermentation that will change ester production is temperature. Higher temperatures result in more ester production. Saisons and other beers with very fruity characteristics (usually ales) are fermented between 70°F and 75°F (21° - 24°C). Lower fermentation temperatures in the range of 64°F to 70°F (18 - 21°C) result in the production of less of these fruity or floral esters and produce more spicy phenolic compounds like vanillin, 4-vinyl guaiacol, and eugenol which impart vanilla, smoky, or clove-like flavors to the beer. Lager fermentations usually occur under even lower temperatures, 50°F to 55°F (10° - 13°C), resulting in the production of very few ester compounds.
Wort composition plays a large role in the formation of esters. This makes sense because if the yeast live in the wort then they probably pull most of their nutrients from the wort. The most influential substance in the wort is oxygen. If there is too much oxygen the yeast will use it to make unsaturated fatty acids and sterols for generating cell walls. This process uses up a lot of acyl CoASH meaning the yeast can’t use the acyl CoASH to make esters. Low oxygen concentrations help keep levels of acyl CoASH high resulting in more ester production. However, if oxygen levels are too low,the yeast will not be able to grow and divide. This can result in slow or incomplete fermentations and even massive cell death called autolysis.
Other wort constituents that make a difference in ester production are sugar content and free amino nitrogen. Sugars, in addition to being turned into yeast, are also used to make acyl CoASH. More acyl CoASH means more esters, so increasing sugar content in wort should increase ester production by yeast. Nitrogen is also used in the production of acyl CoASH as well as the enzymes used to produce esters. Increasing the concentration of nitrogen in wort should also increase ester production.
In summary, the ratios and amounts of esters are some of the most influential factors in finished beer flavors, and therefore important to control. Yeast strain selection plays a large role in what types of esters are produced. The amount of yeast pitched must be high enough to conduct a healthy and strong fermentation, but a small enough amount to allow for the production of more esters. Oxygen content in the wort has similar effects and needs. Too much oxygen will result in less esters being created by the yeast, but too little oxygen will result in poor cell growth and increase the risk of autolysis and a stuck fermentation. Controlling temperature will help supervise how much ester production occurs. The relationship is direct proportional, with warmer temperatures resulting in more esters being produced. Sugar and nitrogen content also affect the formation of esters. With more of these nutrients available in solution, the yeast are able to make more esters. So by selecting the correct yeast and controlling the pitch rate, temperature, and a few basic wort constituent concentrations, ester profiles in beer can be controlled, giving brewers better control over the flavor of their product.
How does one tell what esters are present and what tastes good? By tasting the beer, of course! Analytical Flavor Systems’ Gastrograph Review application allows for detailed flavor analysis to be done on any beer produced. This data can be used to visualize how different factors affect the flavor of a beer. The system stores data on every product reviewed and allows for the comparison of flavors based on differences in manufacturing processes. Using this powerful tool, brewers everywhere can ensure that their beer has the best ratio and amount of flavorful ester compounds. Quality and consistency in beer flavor are key in the success of any brewer - all the way down to esters!
- Peddie H.A.B., “Ester Formation in Brewery Fermentations” Journal of the Institute of Brewing, https://beersensoryscience.wordpress.com/2011/02/04/esters/ Sept-Oct, 1990, Vol. 96, pp. 327-331 http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1990.tb01039.x/pdf
- Engam S., “Organoleptic Threshold Values of some Alcohols and Esters in Beer” Journal of the Institute of Brewing, 1972, Vol. 78, pp. 33-36 http://onlinelibrary.wiley.com/doi/10.1002/j.2050-0416.1972.tb03425.x/pdf