BEER MASHING AND CHILL PROOFING

CLASS: BSc. Biotechnology 3rd Year

SUBJECT: Industrial Biotechnology

SUBJECT TYPE: Major I

COMPILED BY: Asst. Prof. Debosmita Saha

In industrial biotechnology, Mashing and Chill Proofing are the two most critical enzymatic stages of beer production. While mashing prepares the fermentable "food" for the yeast, chill proofing ensures the final product remains clear and stable for the consumer.

---

1. Beer Mashing: The Enzymatic Conversion

Mashing is the process of steeping ground malt (grist) in warm water to activate endogenous enzymes (natural to the grain) or exogenous enzymes (added industrially) to convert starch into fermentable sugars.

Step-by-Step Process & Reactions

A. Acid/Phytase Rest 35-52℃

• Purpose: To lower the pH of the mash to the optimal range (5.2-5.5) and break down minerals.

• Reaction: The enzyme Phytase hydrolyzes Phytin (inositol hexaphosphate) found in the grain.

• Chemical Reaction: 

Phytin + H₂O —Phytase—> Inositol + Phosphate salts + Phytic Acid

• Result: The release of phytic acid naturally acidifies the mash, which is essential for the stability of later enzymes like α-Amylase.

B. Protein Rest (45-55℃)

• Purpose: To break down large, complex proteins into smaller nitrogenous compounds.

• Reaction: Two main enzymes, Protease and Peptidase, work here.

1. Protease: Breaks large proteins into medium-sized polypeptides (important for beer "head" or foam).

2. Peptidase: Breaks polypeptides into Free Amino Nitrogen (FAN).

• Biochemical Reaction:
  
  

Proteins —Protease—> Polypeptides —Peptidase—> Amino Acids (FAN)

• Result: FAN is vital "yeast food" required for healthy fermentation later in the process.

C. 𝛃-Glucanase Rest (45-50℃)

• Purpose: To break down 𝛃-glucans (gummy cell-wall polysaccharides) that cause high viscosity.

• Reaction: 

𝛃-glucanase hydrolyzes the (1→3) and (1→4) linkages.

• Result: Reduces the "gumminess" of the wort, making filtration (lautering) much faster and preventing "stuck mashes."

D. Saccharification Rest (60-72℃)

This is the most important stage where starch is converted into sugar. It involves two types of amylases:

1. β-Amylase (60–65°C): An exo-enzyme that chops two-sugar units (maltose) from the ends of starch chains.
   
   

Starch —β-amylase—> Maltose (fermentable sugar)

2. α-Amylase (67–72°C): An endo-enzyme that randomly breaks internal α-(1,4) bonds to "liquefy" the starch into shorter chains called dextrins.

Starch —α-amylase—> Dextrins (unfermentable, adds body)

---

2. Chill Proofing: Ensuring Clarity

Chill Haze occurs when beer is cooled to 0°C . Proteins and polyphenols (tannins) in the beer bind together via hydrogen bonds to create visible "clouds." When the beer warms up, these bonds break and the haze disappears—but consumers find it undesirable.

The Industrial Biotech Method: Enzymatic Treatment

To prevent this, industries use Proteolytic Enzymes during the cold storage (lagering) or filtration phase.

Step 1: Enzyme Addition (Papain)

The most common industrial enzyme used is Papain (extracted from papaya). Other enzymes include Bromelain or Pepsin.

Step 2: The Reaction

The added protease attacks the specific haze-forming proteins that are prone to binding with polyphenols.

• Reaction:
   

Haze-Forming Proteins —Papain/Protease—> Soluble Peptides/Amino Acids

• Mechanism: By hydrolyzing these proteins into smaller fragments, they no longer have the structural complexity required to bond with polyphenols and form a visible precipitate at low temperatures.

Step 3: Stabilization

Once the proteins are broken down, the beer is "chill proofed." It can be chilled to near-freezing temperatures without becoming cloudy.

---

Summary Table: Industrial Enzymes in Brewing

Stage	Enzyme	Target Substrate	Final Product
Mashing	Phytase	Phytin	Phytic Acid (pH control)
Mashing	Protease	Proteins	Polypeptides (Foam)
Mashing	β-Glucanase	$\beta$-Glucans	Reduced Viscosity
Mashing	α-Amylase	Starch	Dextrins (Liquefaction)
Mashing	β-Amylase	Starch	Maltose (Saccharification)
Chill Proofing	Papain	Haze-Proteins	Soluble Peptides (Clarity)

The Full Industrial Workflow

In a modern industrial brewery, here is how this looks in sequence:

1. Milling: Grains are crushed into Grist to expose the starch.

2. Mashing: The grist is mixed with water. Computers control steam jackets to precisely hit the Acid Rest, Protein Rest, and Saccharification temperatures described above.

3. Lautering: The sugary liquid (Wort) is filtered away from the solid grain husks.

4. Boiling: The wort is boiled with Hops (for flavor). This also "kills" the enzymes from the mash, so they stop working.

5. Fermentation: Yeast is added. It eats the Maltose and FAN created during the mash, producing alcohol and CO₂.

6. Conditioning (Lagering): The beer is cooled.

7. Chill Proofing: Papain enzymes are added to the cold tank to digest haze-forming proteins.

8. Filtration & Bottling: The beer is passed through a fine filter and put into cans or bottles.