Denatured Enzymes

Hey Joe . . . I heard you cooked your old alpha amylase down . . .
how are you going to mash now? I guess if you believe the threads about
mashing-off on some of the homebrew forums you’re going to keep on
mashing because enzymes are actually not heat labile. I read some of
the debate on this and see this as a classic case of error propagation.

There is absolutely no question that enzymes are heat labile and once
denatured they permanently lose their catalytic activity. This
biochemical fact is demonstrated when fruits and vegetables are
blanched and when meat is grilled. Denaturation also happens at
breweries around the world during mashing.

Some enzymes found in malt are extremely heat sensitive and never
have a chance of surviving mashing if they make it out of kilning.
These include lipoxygenase, phytase, beta-glucanse and a wide range of
proteolytic enzymes. Most mashes begin no cooler than 140 ºF (60 ºF)
and the listed enzymes have no activity in the mash because they are
almost immediately denatured during mash-in.

The two primary enzymes of interest in mashing are beta and alpha
amylase because they convert starch to fermentable sugars. In a
laboratory it can be easily demonstrated that these enzymes retain
activity for some time period when held above their denaturation
temperature. Enzymatic re-actions are typically measured by monitoring
the concentration of product over time and the change in product
concentration over time indicates enzymatic rate. Usually these
analyses are run at a fixed temperature to make the test conditions
reproducible.

The rate of enzymatic reactions is highest when the substrate
concentration is high, the product concentration is low and the
temperature is at the optimum for the enzyme (there are many other
conditions, but these are the most pertinent here). If malt and water
are mixed together at 158 ºF (70 ºC) several things begin to happen.
Starch begins to gelatinize, beta amylase begins to denature and alpha
amylase begins to cleave amylose and amylopectin in smaller molecules.
This temperature is well above the optimum temperature for beta
amylase, but that fact does not stop beta amylase activity and maltose
production is seen. Over time, the population of beta amylase enzymes
denatures and those molecules that are active keep working until they
denature. The time period depends on environmental conditions and
enzyme concentration.

The same thing happens with alpha-amylase during mash-off. The
temperature increases, the last bits of ungelatinized starch gelatinize
and alpha-amylase activity continues until the population of alpha
amylase enzymes has been completely denatured. Everything has a time
component and these reactions are not able to occur instantaneously.

So it is logical to conclude from such laboratory experiments (and the
data is out there demonstrating these phenomena) that mashing at
temperatures above the denaturation point of a particular enzyme does
not instantly stop enzymatic activity. To jump to the next step and
assume that mashing-off does not stop enzymatic is faulty logic. It is
also out of context because mash-off usually occurs long after
significant changes in the carbohydrate profile of wort continue. As I
mentioned earlier, enzymatic rates are highest when substrate
concentration is highest and this happens in the beginning of the mash.
By the time mash-off rolls around changes have slowed down
considerably. What does happen during mash-off is that the wort
viscosity is reduced, some ungelatinized starch is freed up and
alpha-amylase activity drops off, usually after the last bits of starch
are converted.

Response by Ashton Lewis.