Enzymes: Catalyst for the conversion of Unfermentable Sugars to Fermentable Sugars

Barley or any other raw materials (corn, wheat, rice, etc.) are the sources of sugar that ultimately gets converted into the alcohol. These raw materials contain starch in them. The starch is a long chain of glucose having the alpha-1,4 and the alpha-1,6 glycosidic bond between the glucose molecules. Starch in itself is naturally unfermentable sugar and hence, can not directly be converted to alcohol by yeast. So, starch should first be converted to fermentable sugar like glucose, maltose, etc.

The conversion of unfermentable sugar is done by the enzymes that the barley kernel possesses. Else, external sources like synthetic enzymes are to be used for the conversion process to proceed smoothly. Among many enzymes that actively work for the conversion process, the following are the most important ones.

1. Amyloglucosidase (AMG)

The commercial name provided is Novozyme. AMG (Amyloglucosidase or glucoamylase) is prepared from a selective strain of fungus Aspergillus niger. The AMG can be used instead of conventional alpha-amylase. This enzyme can breakdown starch completely to glucose. Typical AMGs can withstand temperatures of 80 degrees Celsius for up to 5 minutes and denatures after that period but can withstand 60 degrees for more than an hour.

The main action of AMG is to hydrolyze alpha-1,4 and branching alpha-1,6 linkage in the starch. typically, glucose cleavage occurs from the non-reducing end. The optimum pH for AMG is around 4.0 and the temperature is about 75 degrees Celsius.

2.      Limit dextrinase (Also known as pullulanase)

The primary function of limit dextrinase is to de-branch the alpha-1,6 glycosidic bond of the starch making it more suitable for fermentation. The optimum temperature for limit dextrinase to work actively of malt extract is 60 -62.5 degrees Celsius as opposed to 50 degrees for purified limit dextrinase. Hence, malt extract limit dextrinase is more preferred to purified one. The activity of limit dextrinase increased as the pH of wort (mashed malt) decreased from 5.8 to 5.4.

3.      Alpha-amylase

Alpha-amylase is the enzyme responsible for breaking large, complex, insoluble starch molecules into smaller, soluble molecules. It is stable in hot, watery mashes and will convert starch to soluble sugars in a temperature range from 145 to 158 °F (63 to 70 °C). It requires calcium as a co-factor. The optimum pH is 5.3 to 5.7.

Naturally, barley contains this enzyme itself and does not necessarily require external addition when producing malt whiskey or grain whiskey. But, when barley is not used as the enzyme source, synthetic enzymes are used extensively. For instance, in the production of vodka, rum, brandy, tequila, etc., when barley is not used, the external enzyme source is essential.

These can be produced from bacteria (bacterial amylase) and from Fungi (Fungal amylase).

3.1 Bacterial amylase

Bacterial amylase is mostly generated from the genus Bacillus. Mostly bacterial amylase is thermostable can work at around 100 to 110 degrees Celsius.

3.2 Fungal amylase

Fungal amylase is mostly produced from Aspergillus species. The enzyme has higher acidic tolerance (pH < 3) which avoids bacterial contamination in the wort. Fungal amylase is more preferred over other microbial sources due to their more accepted GRAS (Generally Recognized As Safe) status.

In standard brewing and distilling practices alpha-amylase, beta-amylase, and limit dextrinase are mainly responsible for the production of the fermentable sugars found in brewers’ worts and distillers washes. Alpha-amylase will attack un-gelatinized starch in a limited manner by pitting during malting. During mashing, the diastase complex of enzymes will attack gelatinized/solubilized starch quickly to produce the glucose, maltose, maltotriose, maltotetraose, and dextrins found in the wort.

3.      Beta amylase

Beta amylase is the other mash enzyme capable of degrading starch. Through its action, it is the enzyme largely responsible for creating large amounts of fermentable sugar. It breaks starch down systematically to produce maltose. Beta amylase is active between 131 and 149 °F (55 and 65 °C). But like all enzymes, its activity reaches a peak, declines, and then drops precipitously as temperature increases. The rate is also dependent on the amount of enzyme present. It takes time for all of the enzymes to be destroyed, but what is still intact works very quickly. So as the mash temperature approaches 149 °F (65 °C), beta-amylase is operating at its fastest rate but it is also being denatured.

Diastatic Power (DP) is the total activity of malt enzymes that hydrolyze starch to fermentable sugars. The starch-degrading enzymes contributing to this process are alpha-amylase, beta-amylase, limit dextrinase, and alpha-glucosidase. The driving force for DP appears to be beta-amylase, which usually correlates better with DP than the other starch-degrading enzymes and has the highest activity of all starch-degrading enzymes in malt. Behind malt extract, DP is usually considered the second most important malt quality measurement. For complete conversion of starch to sugars, high levels of barley malt DP are especially important when adding substantial amounts of unmalted adjunct to the mash tun during brewing. See adjuncts. Mashing converts malt starch into fermentable sugars; however, beta-amylase and other starch-degrading enzymes are inactivated as the temperature during mashing increases and DP disappears. There is a dilemma in this phenomenon in that as temperatures rise, starch becomes gelatinized and is a better substrate for starch-degrading enzymes.

The combined action of each enzyme at different phases during the fermentation process helps for the complete conversion of sugars into fermentable forms like glucose, maltose, etc. These simpler sugars are then consumed by yeasts to produce ethanol or alcohol as the primary products.