Isoelectric Precipitation of Casein: Protein in Cheese-Making
Milk is an important agricultural product that provides nourishment to humans. It can be
obtained from cows, goats, sheep, and camels. Aside from water, it also contains fat, proteins,
lactose, and a trace amount of minerals.
Casein is one of the most important milk constituents. It is the main protein found in milk
and it is responsible for its white, opaque appearance. Together with calcium and phosphorus,
casein forms clusters called micelles. These micelles are stabilized at a pH of 6.6. Casein has
an isoelectric pH of 4.6. If more acid is added to the protein, the pH falls and casein precipitates.
This principle is used to isolate casein from milk.
The food industry relies on biochemical processes similar to this in the production of
various food types like jelly, aged meat, and cheese etc. One of the earliest examples of
biotechnology, cheesemaking involves coagulating the casein protein in milk and then
separating the milk into solid curds and liquid whey. The liquid whey is drained away, and the
curds will be produced as cheese. The way in which the curds are handled and treated affects
the overall outcome of the cheese and is why there are thousands of types of cheese.
Objectives
1. To precipitate casein from milk with materials found at home.
2. To witness application of the home experiment in large scale food production, and
3. To identify the function of each step of the cheese-making process.
PART A. Home Experiment
Following the procedure below, do the same in the comfort of your own home.
Skimmed milk or the milk brand Alpine may be used.
On your submission, attach a photo showing your progress for each step and take a
selfie with the finished product. It is recommended to use a white cloth or paper as a background
for the photos. Keep the answers to the questions clear and concise.
Procedure:
1. Place 10 tablespoonfuls of lukewarm milk in a glass.
2. Little by little, add white vinegar with constant mixing until there is visible coagulation.
3. Allow the suspension to stand until the casein settles.
4. Decant and discard the supernatant. Filter the remaining mixture.
5. Wash the precipitate with a small amount of distilled water.
6. Dry the precipitate by pressing between paper towels or tissue paper.
7. Air dry the precipitate and take a photo with the finished product.
Questions:
1. What is meant by the isoelectric point of a protein?
2. At what pH is the protein least soluble? Why?
PART B. Application: Protein in Cheese-Making
View this video, read through this article, then answer the following items as required:
1. Break down the cheesemaking into general steps and explain the biochemical
processes involved.
For example:
Step 1 – Heat the milk.
Applying heat to the milk gets rid of most pathogenic bacteria while creating
optimum temperature for lactic acid bacteria and the enzymes that will curdle the
milk.
2. In the video, no live bacteria were used to curdle the milk. What was/were used as
substitute(s) and why?
Isoelectric Precipitation of Casein: Protein in Cheese-Making
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PART A.
1. What is meant by the isoelectric point of a protein?
 The isoelectric of a protein is the point where its pH carries no net electronic charge and its
neuron. The pH of the molecule's surrounding environment influences its net charge, which
can become more positive or negative as protons are gained or lost. The pH at which a specific
molecule carries no net electrical charge is known as the isoelectric point (pI). The pH of the
molecule's surrounding environment influences its net charge, which can become more
positive or negative as protons are gained or lost.
2. At what pH is the protein least soluble? Why?
 The protein is the least soluble at the pH of 5.8 since this is its isoelectric point. The solubility
of a protein is determined by the net charge on the protein molecule's surface. The net charge
is determined by the pH of the solvent as well as the number and identity of the amino acids
that comprise the protein. A protein's isoelectric point occurs at a specific pH when the
positive and negative charges balance out and the net charge is zero. Proteins are likely to
precipitate and aggregate if there is no charge on the surface.
PART B.
1. Break down the cheesemaking into general steps and explain the biochemical processes
involved.
Step 1 - heat milk Heating milk kills any pathogenic bacteria and create an optimum temperature for
the lactic acid.
Step 2 - gradually add vinegar while stirring Adding vinegar causes the protein denature/break apart.
This causes both physical and chemical changes, causing the curdling.
Step 3 - decant and filter When the vinegar was added, new substance has been formed. The
attraction of positively charged hydrogen ions of the vinegar to the negatively charged casein caused
the formation of the solid substance called the curd and the liquid substance called whey.
2. In the video, no live bacteria were used to curdle the milk. What was/were used as
substitute(s) and why?
 They're all working on curdling milk in that video. Turn off the heat once the milk has curdled.
The curdled milk is then collected in a muslin cloth and set aside to firm up. Rennet used to
curdle the milk since the positive charge hydrogen ions can attract the negatively charged
casein of the milk and can curdle.