It is very common to find someone who understands everything about malt and temperature ramps when talking about mashing beers. However there is another determining factor that acts on your wort and determines the final result of your beer: they are the enzymes!

Enzymes are organic substances (usually of a protein nature), which act as catalysts in some chemical reactions. They convert starches (during the mashing stage) into fermentable sugars and dextrins, so that yeasts consume (during the fermentation stage) those fermentable sugars transforming them in alcohol.

For a chemical reaction to occur between two organic substances that are in the same solution, it is necessary to provide a certain amount of energy for the connection between them to take place. In the process of making beer, this energy is supplied through heat. During mashing, enzymes bind to starch molecules, "break" their chains and shut down, then bind to other starch molecules. Therefore, an enzyme is not consumed during the chemical reaction that it catalyzes.

The increase in temperature causes more agitation of the molecules, therefore, the greater the chances that they will collide to react. However, if the temperature exceeds the limit that each enzyme supports, the agitation of the molecules becomes so intense that the chemical bonds of the enzyme itself break and it denatures and changing its molecular structure, it is no longer effective.

Each type of enzyme acts within a certain temperature range, where the reaction speed is maximum, allowing the largest possible number of molecular collisions without the enzyme being denatured. It is up to the brewer to pay attention to the temperature ranges in which each type of enzyme is active, so that he can make his beer within the previously planned parameters.

The most important enzymes in the mashing process are alpha-amylase and beta-amylase.

However, it is important to remember that there are other enzymes, also responsible for several products and sub-products that are part of the process and that contribute to the excellence of beers.

Below you can see the enzymes...

General tips:

• The unique infusion for mashing at 65°C is targeted by many brewers, with optional mash out.

• There are temperatures and pH levels that favor the performance of one enzyme or another. Temperature being the determining factor of which enzyme will be predominant.

• The inactivation of beer enzymes occurs after 75°C, so make sure that during the mash out the temperature does not exceed 78°C. Above that, there is a risk of extracting tannins from the malt skins , which will cause astringency to the beer.

• The pH (hydrogen potential) affects the structure of proteins, each one has its "own" optimum pH for maximum activity. There is conversion of starch with a different pH than indicated, however, it is less than it could be and it isn't ideal. When added to the boiler the malt naturally lowers the pH of the water. However, each brewer must know the pH of the water, so that it can stay within the appropriate parameters for a good mash. Generally speaking, if the pH of the water is between 6 and 7, after adding the malt and waiting for 5 to 10 minutes, the pH of the mash should be between 5 and 6. The pH range of the brewing water most desired for making beer is between 5,2 and 5,4.

• Following the recipe, respecting the indicated temperature (s) and keeping the pH range between 5,2 and 5,8 is the best way to keep the enzymes working the way we want.

• Beta amylase enzyme denatures at 65°C. Therefore, once the grist is heated above this temperature, the Beta will be completely inactivated. There is no point in lowering the temperature afterwards.

• So for a more fermentable wort and a lighter beer (with less body) possible, it is necessary to make the mash between 60°C and 65°C, taking care not to let the temperature pass 65°C, avoiding denaturing the Beta Amylase enzyme. If it is necessary to connect the wort to

• For a less fermentable wort and a fuller beer it is necessary to make the mash above 65°C. And for a fuller beer (outside the carbohydrate range) above 67°C.

• One point about temperatures that doesn't have much to do with enzymes is that a stop of at least 15 minutes at 60°C helps to solubilize the starch and enzymes in the wort. That is, even if you want a beer with more body and less attenuated, a short stop at 60°C helps a lot to increase the conversion efficiency.

Alpha and Beta Amylase

Alpha and beta amylase are the main enzymes responsible for converting starch into sugars during mash. The bigest doubt of most brewers, especially beginners, is how each one acts and what the expected result is if one or the other is strengthened during mashing.

• α-amylase breaks starch down into maltose and dextrin, by breaking down large, insoluble starch molecules into soluble starches (amylodextrin, erythrodextrin, and achrodextrin) producing successively smaller starches and ultimately maltose.

• β-amylase catalyses the hydrolysis of starch into maltose by the process of removing successive maltose units from the non-reducing ends of the chains.

Each of the two enzymes acts on a type of bond between the sugar molecules that form the starch molecules (carbohydrates).

Beta amylase breaks bonds 1-4 close to the ends of the starch molecules. It does not break 1-6 links or 1-4 links next to broken links.

Alpha amylase breaks down any 1-4 bond of the carbohydrate molecule.

Debranching enzyme is a molecule that helps facilitate the breakdown of glycogen (multibranched polysaccharide of glucose).

Bonds 1-6 can be broken by the carbohydrase (debranching) enzymes (limit dextrinase) activated during the malting process.

Enzymes are proteins that act by controlling speed and regulating the reactions that occur in the body. They catalyze specific chemical reactions acting on specific substrates and at specific locations on those substrates.

The action of enzymes can be influenced by some factors, such as high temperature.

Enzymes are specialized globular proteins that act by controlling speed and regulating the body's chemical reactions. It is important to note that some RNA molecules , known as ribozymes , act as enzymes. These still have a catalytic role, that is, they act by increasing the speed of chemical reactions.Enzymes have a three-dimensional structure, and their activity depends on the characteristics of the environment in which it is found.

Enzymes are highly specific , each of which acts on a specific substrate in a reaction. Currently, more than 2,000 enzymes are known, and each one acts in a specific reaction.

Enzyme nomenclature

Enzyme nomenclature occurs in several ways. The three most used forms are:

• Classic or recommended name : usually names, adding the -ase termination to the name of the substrate on which the enzyme acts. This is the form most used by those who work with enzymes. For example, the amylase enzyme acts in the hydrolysis reaction of starch in glucose molecules , and urease catalyzes the urea hydrolysis reaction in ammonia and CO _2.

• Usual name: uses names consecrated by use, such as trypsin and pepsin.

• Systematic name: more complex form and instituted by the International Union of Biochemistry and Molecular Biology (IUBMB), it presents more information than the others regarding the functionality of the enzyme. The systematic name usually has three parts: the name of the substrate, the type of catalyzed reaction and the suffix -ase . For example, the reaction of converting glucose-6-phosphate to fructose-6-phosphate is catalyzed by the enzyme called glucose phosphate isomerase. In addition to the systematic name, the enzyme also receives a number, which must be used for accurate identification. This numbering follows the model: EC XXXX. The acronym EC represents the Enzyme Commission (Enzyme Commission) of the International Union of Biochemistry and Molecular Biology, and the sequence of four numbers refers to its classification.

Classification of enzymes

Enzymes can be classified according to the International Union of Biochemistry and Molecular Biology and according to the type of reaction they catalyze , as follows:

• Class 1. Oxide-reductases: oxide-reduction reactions or electron transfers (hydride ions or H atoms). Examples: dehydrogenases and peroxidases.

• Class 2. Transferases: reactions of transfers of functional groups between molecules. Examples: aminotransferases and kinases.

• Class 3. Hydrolases: hydrolysis reactions, in which a molecule breaks down into smaller molecules with the participation of water. Examples: amylase, pepsin and trypsin.

• Class 4. Liases: reactions in which the addition of groups to double bonds or the removal of groups leaving a double bond may occur. Example: fumarase.

• Class 5. Isomerase: reactions in which the formation of isomers occurs. Example: epimerase.

• Class 6. Ligase: synthesis reactions in which molecules join with energy expenditure, usually from ATP . Example: synthetases.

Mechanism of action of enzymes

Enzymes work by binding to specific substrates at specific locations. At the end of the process, they are released to catalyze new reactions.

The energy required for a reaction to start is called activation energy . Enzymes work by reducing this activation energy and causing the reaction to occur more quickly than in the absence of it. This catalytic capacity of the enzymes increases the speed of the reactions by about 10 ¹⁴ times.

The action of enzymes occurs through their temporary association with the molecules that are reacting, bringing them closer together. As a result, enzymes can also weaken existing chemical bonds, facilitating the formation of new bonds. They bind to specific molecules, called substrates , and in specific places, the activation sites, forming a transient complex. At the end of the process, this complex decomposes, releasing the products and the enzyme, which usually recovers its shape and can be used again to catalyze reactions.

The enzymes act in a chain, and several of them can act in sequence, in a certain set of reactions, forming the so - called metabolic pathways . A cell has several metabolic pathways, each responsible for a specific function, for example, the synthesis of substances, such as amino acids .

Connecting sites

As stated, enzymes bind to substrates at so-called binding sites . They have specific amino acid residues arranged in a three-dimensional form, forming the binding sites, places where the substrates bind during the reaction.

In addition to this three-dimensional arrangement , the enzymes present, in these sites, an appropriate arrangement of hydrophilic (interact with water) and hydrophobic (do not interact with water) regions , charged (with electrical charges) and neutral (with no electrical charges).

The substrate must have an adequate , structural and chemical configuration , in order to be lodged in the connection site. This perfect fit model is known as the key-lock model , due to the relationship with the fact that each key fits a specific lock. However, it is known that the approach and binding of the substrate to the binding site induces a conformational change in the enzyme, making it ideal. This model is known as the induced fit model .

Factors that regulate enzyme activity

The enzymes can have their activity influenced by many factors. Among these we can highlight the temperature, the pH and are regulatory enzymes.

• Temperature : Most enzymes increase their reaction rates as the temperature at which they act increases by 10 ºC. However, this rate starts to drop as soon as the temperature reaches 40 ºC. From that temperature, it is observed that the enzymes start to undergo denaturation, an unfolding of their structure.

• pH : Changes in the pH of the medium in which the enzyme is found leads to changes in its loads. The maintenance of the form of the enzymes is due to the attraction and repulsion between the charges of the amino acids that constitute it. Changes in these charges change the shape of the enzyme, affect the bond between it and the substrate and, thus, its functionality.

• Regulatory enzymes : These act by regulating the rate of metabolic pathways. Often, they occupy the first place in the sequence of the metabolic pathway and increase or decrease the activity through some signals, such as the substrate levels or the energy demand of the cell .

Enzyme Summary

• Enzymes are proteins that act as biological catalysts.

• Catalysts are substances that act by decreasing the activation energy of reactions, increasing the speed at which they occur and not being consumed in the process.

• Enzymes have high specificity, acting only on specific substrates.

• Enzymes work by decreasing the activation energy of reactions in cells.

• Temperature, pH and regulatory enzymes are factors that influence enzyme activity.