Step Mash for Customized Worts Part I


Using Enzymes to Break down Glucans and Proteins

By Jim Busch (Brewing Techniques - Vol. 5, No.3)

Enzymes are the catalysts that convert malt proteins, starches, and carbohydrates into fermentable wort. Step mashing offers the brewer tremendous flexibility in creating worts that have specific properties that can ferment to produce exactly the beer you want.

This installment of Home Brewery Advancements begins a two-part exploration of step mashing. Step mashing is a technique by which the brewer ramps the mashing temperature through a series of rests to promote specific enzymatic reactions. Step mashing can be used to compensate for certain characteristics in your grist or to add specific features to your beer. This technique is therefore an important tool in the creative brewer’s tool chest.

In the first part of this two-part series, I provide an overview of the low-temperature enzymes — glucanases and the proteolytic enzymes (proteinases and peptidases). Next issue, I will delve into the amylases, the enzymes that convert starches into sugars.

Enzymes 101

Step mashing has everything to do with malt enzymes, so let’s start with a brief overview of what enzymes are and how they are useful in brewing. Enzymes are merely a specific class of proteins. They occur in the cells of living organisms and function as catalysts in the organisms’ chemical processes. Malt contains many types of enzymes, and it is these enzymes that are the workhorses of mashing, breaking down proteins, starches, and carbohydrates into smaller units that are useful for the production of beer.

Fortunately for brewers, enzymes are water soluble; they can be hydrated by the uptake of water during malt germination and mashing. During mashing, these enzymes become liberated from the malt and are found principally in the water fraction of the mash.

Each malt enzyme becomes active in a specific temperature range. At a certain point above the optimal temperature range, the temperature becomes high enough to deactivate it. When an enzyme is deactivated it is called denatured, which means that the enzyme is no longer capable of performing its function. A denatured enzyme therefore not only ceases to be active, but its activity cannot be restored by lowering the temperature back into its useful range. (You can, however, add more malt to provide additional “fresh” enzymes.) The brewer’s art is a controlled use of time and temperature to activate and then deactivate these enzymes.

By carefully controlling the temperature and duration of each rest, you can activate and deactivate specific enzymes to fix certain parameters in the finished beer. These parameters include viscosity or gumminess in the lauter tun and the wort’s protein spectrum and fermentability.

The creative use of enzymes actually begins in the malthouse, where they play an essential role in the malting process. Maltsters manipulate the germination and kilning processes to create unique malt characteristics, thanks largely to the role of enzymes. Some of these enzymes do not survive the kilning stage, which means the maltster has effectively removed that enzyme option from the brewer. Others remain in the malt and are capable of playing important roles in the mashing process.

To really master the art of brewing, advanced brewers will spend a little time and forethought during recipe formulation to consider each enzyme and how to manipulate target enzymes to meet the needs of a given beer style. Some beer styles (English bitters, for example) show no appreciable benefit from step mashing; for such styles, you need only one rest to convert starch to sugar. Some enzymes are unnecessary or even detrimental for a particular style.

If you have ever brewed a beer with long rests in many temperature ranges only to find out that the finished beer was thin, watery, and lacking head retention, then you have experienced firsthand the problem of excessive enzyme activation. In this case, you may have rested too long in the temperature range of maximum activity for proteolytic enzymes. These enzymes will ultimately break down head-promoting proteins into amino acids, and if left active too long can hurt a beer’s foam stand. Another example includes resting too long in the range of maximum beta-amylase activity and then watching the beer ferment down to a final gravity of 1.006 (1.5 °P).

Table I: Ranges of Enzymatic Activity


Range of Activity


Temperature Tolerance

Beta glucanase

95–105 °F (35–40 °C)


Survives to 122 °F(50 °C)

Proteolytic enzymes

113–140 °F (45–60 °C)


Survives to 158 °F (70 °C)

Beta amylase

140–149 °F (60–65 °C)


Survives to 167 °F (75 °C)

Alpha amylase

158–167 °F (70–75 °C)


Survives to 176 °F (80 °C)

Brewers are most concerned with four enzyme groups: glucanases, proteolytic enzymes (proteinases and peptidases), and the amylases beta-amylase and alpha-amylase (see Table I). Each plays an important role in the mash tun and helps to determine the finished beer’s profile.

Glucanases work on reducing beta glucans, which tend to add an undesirably gumminess to a mash and can create runoff problems. Proteolytic enzymes break down proteins and peptides to provide essential yeast nutrients and to help prevent haze formation in the finished beer. Amylases convert starches into sugars essential for fermentation. This first installment addresses the first two groups that you might encounter in a step mash schedule, glucanases and proteolytic enzymes, which operate on the lower end of the temperature range.

Understanding the enzymes and their effects on the mash is quite easy. Let’s delve in.


Beta glucans and the action of glucanases are too often ignored in recipe formulation and mashing regimens. At times, it is essential to pay attention to the beta glucan content and the techniques you can use to degrade these compounds. The consequence of ignoring them may be high-viscosity mashes, lauter problems, and even diminished yield.

Glucans arise from the composition of barley seeds. The endosperm of the barley seed consists of starch and proteins that are bound in a matrix. The cell walls within the endosperm contain as much as 75% beta glucans, which are partly degraded during malting and mashing (1). In barley, water-soluble glucans are found in both beta 1–3 and beta 1–4 linkages, the latter comprising 70% of the total glucan content (2).

A malt’s degree of modification roughly determines the extent to which these structures are degraded during malting and therefore the extent to which surviving beta glucans are passed on through the malt. Beta glucan solubilase (as the name implies) is the enzyme that makes beta glucans soluble during malting. Glucanases cleave or cut beta glucans at specific places in their chain structure, thereby degrading them. Several glucanases are active during malting, including endo-1,3-β-D-glucanase and endo-1,4-β-d-glucanase; their names indicate the linkages they cleave (see Table II for a list of enzymes, their activation and denaturation temperatures, and specific activity). Many of these glucanase enzymes are highly temperature labile, and relatively few survive high-temperature kilning, where the total glucanase enzymatic concentration can decrease by as much as 50–60% (1).

Table II: Glucanase Activity Profiles*

Glucanase Type

Optimum pH

Optimum Temperature

Denaturing Temperature

Enzyme Acted Upon



99–113°F (37–45 °C)

131 °F (55 °C)

Beta-1–4 glucans




158 °F (70 °C)

Beta-1–3 glucans




146°F(63 °C)

Mixed beta 1–3 and 1–4 linkages

Beta glucan solubilase



163°F(73 °C)

Proteins/beta glucans (acid carboxypeptidase)

*Data from reference 1.

Active range: Glucanase enzymes are generally most active in the range between 95 °F and 105 °F (35–40 °C), though some become effective at 140 °F (60 °C) (see Table II). Beta-glucanase is rapidly deactivated above 122 °F (50 °C).

Practical applications: Undermodified malt. The influence of the maltster over a malt’s beta glucan content has direct implications for the brewer. It implies, for example, that low-temperature rests with highly modified pale ale malt will not appreciably alter the glucan content of the wort because the enzymes that are active at these temperatures have been significantly reduced during malting (though some brewers still report greater extract yields when using low-temperature rests with these malts [3]). In many continental malts, on the other hand, appreciable amounts of glucanases survive the low-temperature kilning process and are available to the brewer, who can use them to degrade the malt matrix and release some carbohydrate material.

Table III: Percent Total Nitrogen in Classes:

Size of Molecule

Relative Quantity (%)

High molecular weight:


Medium molecular weight:


Low molecular weight:


Adjuncts. Adjuncts have not been through the malting process and therefore have not formed any enzymes of their own. Adjuncts disperse, or gelatinize, their starches, either during pre-cooking or in the mash, freeing them up to be worked on by the enzymes from the malted grains. Thus, it’s best to select for adjuncts with low levels of beta glucans in the first place. To reduce the beta glucans that do exist, a low-temperature rest that maximizes the glucanase activity of the malted grains can be very beneficial.

Wheat malt. Wheat malt contains particularly high levels of gums and proteins, so step mashing is almost mandatory. When brewing classic Bavarian wheat beers, a rest at 111 °F (44 °C) may also be desirable because it will free ferulic acid (4-hydroxy-4-methoxy cinnamic acid); some yeast strains, Bavarian Weizens in particular, decarboxylize ferulic acid into 4-vinyl-guaiacol during fermentation (4), which contributes a characteristic clovey/phenolic flavor. Ferulic acid is bound to pentosans in grains with ester bonds, and beta glucanase degrades both beta glucans and pentosans to free the acid.

Proteolytic Enzymes — Proteinases and Peptidases

The next major area of enzymatic activity occurs as the mash is exposed to the proteolytic enzymes — proteinases and peptidases. These enzymes are active between the temperatures of 113 °F and 140 °F (45–60 °C), typically known as the protein rest. As their names imply, these enzymes break down proteins into less complex compounds, such as peptides, and, ultimately, their amino acid building blocks. A thick mash (1 qt water/lb malt, or thicker) is known to favor the action of proteolytic enzymes.

Proteins can be grouped into three ranges of importance to brewers: high molecular weight, medium molecular weight, and low molecular weight proteins (see Table III). Brewers like to reduce larger proteins during mashing because these are the proteins that cause haze in finished beer. At the same time, it is desirable to retain some medium and smaller sized proteins for head retention and body while breaking down enough to provide the amino acids that are so important to the yeast. Proteinases work at the higher end of the temperature range to break down the larger proteins, while peptidases work on smaller proteins (peptides) at lower temperatures.

Proteins and malting: When we talk about proteins, we are talking about nitrogen. The nitrogen compounds that comprise the protein spectrum of malt are divided into four simple protein fractions (1), including albumin (leucosin), globulin (edestin), prolamin (hordein), and glutelin. The action of proteinases and peptidases during malting degrades the vast majority of these nitrogen fractions, and hence malting again plays a significant role, determining to a large extent a beer’s protein spectrum.

The endosperm of the unmalted barley seed is where we find hordeins and glutelins, where they form a protein matrix around starch granules. Germination hydrolizes these proteins, thus liberating alpha-amylase, which is useful in converting the starch into sugars in the mash tun (alpha-amylase will be discussed in the next issue’s installment). Unmalted grains do not benefit from this enzyme activity, which is one reason unmalted grains contribute little to the protein spectrum in wort.

During malting, the hordein protein fraction declines dramatically and the simple water-soluble albumin fraction increases appreciably. The proteolysis that occurs in malting is mostly performed by endopeptidase, which facilitates amino acid production in the mash by preparing the protein substrate for the action of carboxypeptidase.

In the mash: Peptidases. Peptidases are most active in the lower end of the proteolytic enzyme temperature range, at 113–128 °F (45–53 °C), with 122 °F (50 °C) the optimum. Rests at this temperature favor the breakdown of medium molecular weight proteins into peptides, which are simply building blocks of amino acids linked by amide bonds.

The downside is that all-malt worts already yield abundant quantities of amino acids for healthy fermentation, and extensive rests at the maximum range of peptidases activity can result in excessive degradation of medium molecular weight proteins and can lead to poor foam stability. Often a brewer is better served to avoid long rests at ~122 °F unless necessary for undermodified malt, as might be used for lagers or for adjuncts, and instead spend more time in the range of the proteases, between 131 °F and 137 °F (55–58 °C). Rests in this range will act more selectively on larger proteins, degrading them into head-promoting mid-sized molecular weight proteins while avoiding haze-causing fractions. Note that Table I shows that beta-amylase becomes effective at 140 °F (60–65 °C), so beware of the effects of resting too long at this temperature.

Customize the Wort to Your Specifications

Time, temperature, and pH are the main factors that brewers have at their disposal to alter the glucan and protein content of sweet wort. The challenge is to begin with the agricultural raw ingredients of malted barley, malted wheat, and raw grains and optimize the mash program to yield the desired wort constituents to meet a given beer style.

The maltster’s role in modifying the proteins and glucan structure of raw grains during malting cannot be overemphasized. Maltsters work with many of the same enzymes as brewers, but maltsters have more flexibility because much of their action occurs before heating; after heating, the enzyme concentration is radically altered.

By carefully considering the raw ingredient profile and commanding a modest understanding of the action of each enzyme, you can manipulate the mash program to maximize yield and lauterability and fix the protein spectrum of the wort.

In the next issue, I will examine the function of the amylases, which have the all-important job of breaking down starches into sugars.

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