Kitchen Laboratory: Proteins and Cheese Making




About: Food Chemist, bug lover and enthusiastic crafter with a passion for papercrafts, specially origami

Chemistry surrounds us, and a Kitchen is a laboratory where a lot of complex chemical reactions take place while we are making our foods. A common task like boiling an egg involves chemical changes.

So I couldnt find a better place to talk and learn about proteins than a kitchen.

In this instructable you are going to learn something about proteins and how they are involved in cheese making. Of course I'll explain how to make fresh cheese, a very simply and easy to make cheese

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Step 1: Proteins and Enzymes

First we are going to review a few concepts about proteins and its characteristics and functions

I'm sure you've heard about proteins, but, what are proteins?

Well, in a few words, proteins are complex polymers composed of amino acid monomers which are arranged in a specific three dimensional pattern held together by a variety of chemical forces and, they are considered to be the primary structure of all living organisms. Some examples of protein are muscle, hair, skin, hormones, and enzymes

Protein structure can be affected by different factors like pH, temperature and the presence of other compounds like enzymes. When an external factor affects the protein structure it losses its biological activity and we call this process protein denaturation. An example of protein denaturation is our boiled egg and of course cheese (In this instructable we are going to denaturate milk proteins using a enzyme, temperature and changing its pH using an acid)

Now that you know what proteins are, let's talk about enzymes

Chemical reactions in living cells are complex and highly controlled. Certain chemicals accelerate reactions without being changes themselves. These agents are called catalysts, and the rate acceleration caused by such substances is called catalysis. Enzymes are specialized proteins uses as catalysts in biological reactions and are produced naturally in plant, animal and microbial cells, and there are thousand of enzymes because they have a specific function and they work at very low concentrations.

In an enzyme catalyzed reaction, the enzyme joins with the substrate (a reactant in a chemical reaction) and helps transform the substrate into the product. As I told before, an enzyme is a catalyst, which means that it participates in the reaction but is not used up during the reaction. The action of the enzyme continues until it losses it biological activity which means is denaturized affected by temperature, pH or other compounds.

Step 2: Milk

As a biological product, milk contains proteins, the principal milk proteins are known as casein but it also contains other proteins. It consists of 80% casein and 20% whey protein. There are four major types of casein molecules: alpha-sl, alpha-s2, beta, and kappa

Because of the negative charge of the casein, it is dispersed in milk so if we want to make cheese we need to denaturate casein to coagulate it.

Denaturation of milk proteins
When an acid is added to milk, the H+ concentration neutralizes the negatively charged casein micelles. When milk is acidified to pH 4.7, it reaches its isoelectric point (the point at which all charges are neutral) and it forms a precipitate known as acid casein. Cottage cheese and cream cheese manufacture involves an acid precipitation of casein with lactic acid or lactic acid-producing microorganisms. Acid casein is used in the chemical industry and as a glazing additive in paper manufacturing.

Casein also can be coagulated with the enzyme rennin, which is found in rennet (an extract from the stomach of calves)
Do you remember which factors affects proteins? If not, go back to step 1 because rennin as a protein is affected by these factors and is enzymatic activity can change.
Rennin works best at body temperature (37 C). If the milk is too cold, the reaction is very slow, and if the milk is too hot, the heat will denature the rennin, rendering it inactive. The mechanism for the coagulation of the casein by the rennin is different from the acid precipitation of casein. The coagulation of the casein by rennin is a two-stage process. In the first stage, rennin (a proteolytic enzyme which means it acts only on proteins) splits a specific bond in the amino acid chain of the kappa-casein macromolecule converting it into a-para-kappa-casein and a glyco-macropeptide. This causes an imbalance in the intermolecular forces in the milk system, and the hydrophilic (water-loving) macropeptides are released into the whey. Unlike kappa-casein, the parakappa-casein does not have the ability to stabilize the micellular structure to prevent the calcium-insoluble caseins from coagulation. In the second stage, colloidal calcium phosphate (this is and example of other compound affecting proteins) bridges within the casein micellular structure, resulting in the three-dimensional curd structure.
The rennin coagulum consists of casein, whey protein, fat, lactose, and the minerals of the milk, and has a fluffier and spongier texture than the acid precipitate. Rennet is used in the manufacture of cheese and cheese products (nowadays, most of the cheese producers do not use calve rennet because is very expensive they use "vegetable rennet" which is made from microorganisms), and rennet casein is used in the plastics industry.

Step 3: Glossary

Did you feel lost in the lecture because of some or the terms used?, here is a small glossary, hope it helps

Amino acids contain carbon, hydrogen, nitrogen, and sometimes sulfur and serve as the monomers to make peptides and proteins. Amino acids have a basic structure that includes an amino group (NH2) and a carboxyl group (COOH) attached to a carbon atom.
This carbon atom also has a side chain (an " R group). There are twenty amino acids, found in the body. Eight of them are essential for adults and children, and nine are essential for infants.

Casein is a milk protein. There are four major types of casein molecules: alpha-sl, alpha-s2, beta, and kappa.

Coagulation is the transformation of a liquid into a soft semisolid or solid mass. In the coagulation of milk, it refers to the aggregation or clumping together of proteins.

Colloid is a suspension of finely divided particles in a continuous medium in which the particles do not settle out of the substance rapidly, and are not readily filtered.

Denatured means changed from its natural state. In a denatured protein, its characteristics or properties have been altered in some way, by heat, chemicals, or enzymatic action, resulting in the loss of its biological activity.

Digestion is the chemical breakdown of large food compounds into smaller molecules that can be absorbed by the intestines in the human and animals. The smaller food molecules travel in the blood and are used by cells to make other components or produce energy needed by the body. Digestion begins in the mouth as salivary amylase begins to break down starch into simple sugars.

Isoelectric means having equal electric potential.

Kappa-casein is one of the four major types of casein molecules. Kappa-casein selfassociates into aggregates called micelles. The alpha- and beta-caseins are kept from precipitating by their interactions with kappa-casein.

Micelle is a submicroscopic aggregation of molecules, as a droplet in a colloidal system.

pH is the concentration of hydrogen ions in a solution and is expressed in terms of the pH scale. Low pH corresponds to high hydrogen concentration and vice versa. pH is very important because hydrogen ions are positively charged and alter the charge environment for other molecules in solution. By putting different forces on the molecules, the molecules change shape from their normal shape. This is particularly important for proteins because the shape of a protein is related to its function

Peptide bonds are covalent bonds between two amino acid molecules.

Precipitation is the removal of insoluble material from solution.

Rennet is an extract from the inner lining (membrane) of the fourth stomach (abomasum) of the calf. The abomasum is the gastric stomach of ruminant animals such as the cow. The lining is used to make cheese because it contains the enzyme rennin.

Rennin is a proteolytic enzyme that is used to coagulate milk to make cheese. Rennin is typically used in the form of rennet, a commercial preparation taken from the abomasums (fourth stomach) of young calves, but because its demand is great and supply limited, the cheese industry has been increasingly turning to microbial rennin produced from genetically engineered microorganisms. Rennin is also known as chymosin.

Substrate is the name of a reactant molecule for enzymes. A substrate is the substance on which an enzyme acts. Using the analogy of a lock and key, the lock is the substrate, and the key is the enzyme.

After all these reading, let's have fun with proteins making some cheese.

Step 4: Materials

First we need to gather some materials in order to make cheese

3 liters of pasteurized whole milk

Rennet: you can find it liquid or in tablets, you can buy it on grocery stores and of course in, click here to see the product.

Calcium chloride (CaCl2) (found in drugstores)


White vinegar

Pot / Pan: stainless steel, glass ceramic. Do not use aluminum or cast iron

Colander (it will serve as cheese mold)


A big spoon

A wood spatula for stirring (may be a spoon if you do not have a wood spatula)

Some recipients to store the whey

Some cups or glasses



A balance (a kitchen balance is ok if it can measure grams, if you do not have it, check the photos to figure out the quantities I weight)

Gauze (I used sterile gauze to ensure is really clean)

A 10 mL pipette (mine is a 1 mL pipette but still worked) or a graduated dropper if you cannot get the pipette

As you can notice, I use the Metrics International System of Units, in case you want to convert it, here are two links to sites of conversion factors:

Step 5: Testing Our Milk and Rennet

First of all, wash your hands perfectly and be sure all the materials you are going to use are clean.

Believe it or not, not all commercial milks are suitable for cheese making. This is due to several factors (acidity of milk, calcium content, antibiotics present in milk, etc), so is important to test it before making our cheese. Since I do not know which kind of rennet you got, follow the directions of the manufacturer to prepare your rennet.

Most of the tablet rennet dosage is one tablet for 100 liter of milk (like mine), and should be dissolved in 200 ml of cold water (In theory, this means I only need 0.2 mL to coagulate 1 liter of milk). So I broke one tablet in four parts and dissolved 1/4 tablet in about 30 mL of cold water.

This is important: it seems Junket Rennet Tablets is the most common commercial rennet in the USA, but also is a kind of "diluted" rennet (because of the ingredients) so after a long searching in the internet I found that 1 Junket Rennet tablet coagulates about 18 liters of milk, so you have at least to quadruple the dosage to obtain good results.

Then, if you are using Junket Rennet Tablets dissolve a whole tablet in about 30 mL of cold water.

To test the milk, take about 30 mL of milk and warm it to 35C (remember, rennin is a enzyme and works at body temperature, if you try it on cold milk nothing is going to happen and if you heat the milk over 38C you will inactivate the rennin and also nothing will happen, so be sure the temperature is between 32 - 38C and keep it at this temperature, you can achieve it by immersing the cup in warm water), then add 3 mL of rennet, stirr and take the time it needs to form the curd.

If everything goes well, it only takes lees than 5 minutes to form the curd (remember, we formed the curd by using an enzyme to modify the casein which is the protein of the milk so it become insoluble and form a curd)

If after 10 minutes nothing happens, and you are sure you kept the milk at 35C and you did everything well, then something went wrong, try other milk, or other rennet, but first try other milk (is not common that rennet doesn't work)

If your test is successful, you can continue to the next step. Look at the photos to see examples of two different milks (one is good for cheese making while the other never reached coagulation)

Step 6: The Calcium Chloride

Weight 1g of calcium chloride and dissolve it in 2 or 3 mL of cold water. We are going to use calcium chloride to reinforce the calcium present in the milk because commercial pasteurized milk looses calcium during pasteurization and calcium plays and important role in the curds forming. So if the milk does not contain enough calcium we are not going to get a nice curd. (I read Junket tablets contains calcium, so if you couldn't find calcium chloride, maybe you can skip it if you use Junket Rennet but I'm not sure because I've never tryed Junket Rennet, so is better to try to get calcium chloride)

Step 7: Making the Curd

Warm your 3 milk liters to 32C - 36C (remember do not overheat or you are going to destroy the enzyme and nothing is going to happen) and let in the stove to keep it warm (of course the stove is turn off now we got the temperature we wanted), add the calcium chloride solution, stir gentle and add 1.5 mL of rennet (or 6 mL if you are using Junket Rennet). Stir gentle and let it undisturbed from 1 to 3 hours (mine took about 55 minutes to be ready but as I told you, the rate depends of the rennet and the milk) so, be patient and just be sure to keep temperature between 32C - 36C.

After one hour, test for a clean break, insert your knife into the milk and if it is ready, it should be clean, if not let it and test it again after 20 or 30 minutes.

When it's done, you should have a firm but soft gel, this is called "curd".

Step 8: Cutting the Curd

So now we are going to cut the curd. Using the knife, make vertical and horizontal cuts to the curd, we are trying to cut the curd into cubes like about 2 cm wide. I think the videos are the clearest explanation I can give of to how make it.

After you finished the vertical and horizontal cuts, continue cutting as shown on the next video

Step 9: Cooking the Curds

Warm until 38C and stir the curd to help it release the whey. The stirring must be gentle and slow, do not do it fast because you can destroy the curds.

When you reach the temperature, turn off the stove and continue stirring for 5 minutes more, then let the curds sit for a few minutes.

Now we are going to retire the whey from the curds. You can try to decant it but if you have not practice, you can loose a lot of curd in this process, so is best to retire the whey using a spoon. Retire up to 2/3 of the whey. (Keep the whey; we are going to use it later)

Step 10: Adding Salt to the Curds

The next step is to salt the curds. A typical cheese contains from 0.5 to 2% of salt, so I decided to add 1% of salt, this means 3 g of salt (if you want, you can add more or less salt)

Add the salt to the curds and stir gentle to distribute the salt among the curds.

Then, we are going to past the curds to the colander which also is going to serve as mold in this case. Cover the colander with the gauze and pass the curds to the colander using the spoon. Be sure to have a recipient under the colander to receive the whey.

Step 11: Finishing the Cheese

When you are done, press the curds gently with the spoon to help it to drain the whey.

Your cheese is almost done, now you just need to wait, let it drain for two or three hours at room temperature, then carefully, turn it over cover with the gauze keep it on refrigeration and let it continue draining for 24 hours. After 24 hours, your cheese is ready. Remember this kind of cheese is fresh cheese so you have to eat it quickly (it should last a week). This kind of cheese is called basket cheese (queso canasta) in my country because the mold is a basket or a colander and the cheese takes the shape of the basket or colander. It is very common here in Mexico.

Also is fat reduced because it contains about 20% fat while other cheeses contains 40% fat or more. These is also the basics for cheese making, all the other type of cheeses follow a similar process the difference is that most of the cheeses need cultures and the treatment of the curds is different.
Is excellent for salads and also you can eat it as a snack.

Advantages of home made cheese is that it has a different flavor than the ones sold in the stores, and you know exactly how is made off and which ingredients contains. Commercial made cheeses often have other ingredients (cheeses with other ingredients different of milk are called analog cheeses) to help the producer to get a better cheese yield and improve costs, but real cheese is only made with milk, sodium chloride, rennet and cultures.

Step 12: Whey Proteins

This is optional, your cheese is done, just remember we are making a kitchen laboratory, so as scientists we need to have a little curiosity and always ask what happen if...?

Remember we told milk proteins are 80% casein and 20% whey proteins? In fact, whey is very nutritious and is rich in proteins (these proteins were not affected by rennin which is a specific enzyme for casein, remember, enzymes are very specific catalysts) and vitamins (is greenish-yellow tint is because of the rivoflavin it contains)

Well, these proteins still soluble in the whey so we can try to denaturate it and try to make a kind of ricotta cheese (I say try because I didn't got ricotta but an acid, creamy, soft and spread able cheese-like curd.

First we are going to acidify the whey using the white vinegar, add two or three spoons of vinegar (about 30 -50 mL) to the whey. Then warm it and stir it gentle until temperature is near boiling (92 -95C it depends of your location) Foam will build up and if it boils, it can boil over, so be careful at this point Stop warming and let it cool until it reaches room temperature.

The curds should look like clouds in the bottom and whey now is clear.

Step 13: Whey Curds

Carefully remove the curds, if you can, retire the whey without disturbing the curds and then gently transfer the curds to the cloth (just like we did it before with the fresh cheese). You will notice this curds are different from the one we got from the casein, and is obvious, these curds are not casein but whey proteins, and they were denaturalized by heating and acid instead of using an enzyme.

Then let it drain and you will get your ricotta, remember you also need to consume it fast, but it may be no trouble because the quantity you get is low.

Step 14: Unexpected Trouble

If happens that the curds are very light (like in my case) and you cannot avoid disturbing it, nor take them with a fine colander, then you have to improvise (like I did) and filter the whey to separate the curds.

I used a clean cloth and a fine colander to filter it, the draining was very slow because the curds covered the cloth and it made very slow the whey draining. However, as I told you before, I got an acid creamy, soft and spread able curd, is not ricotta because but you can see the whey proteins now :).

My theory is that I got this kind of curd because I added too much vinegar to the whey (by accident I added like about 250 mL instead of 50 mL, this is why I recommend you to use a spoon or your dropper). However the objective was accomplished because I could demonstrate three ways of denaturate proteins: by enzyme action, by temperature and by changing pH.

If you want to help me to confirm my theory, then add about 250 - 300 mL of vinegar to your whey and let me know if you get the same type of curd, and please be patient while the curds drains(at the end, I helped it by gently squeezing the cloth and I used my spatula to remove it from the cloth, so make sure your cloth is really clean to avoid contaminating it)

Step 15: Want to Learn More? Then Read...

To learn more about cheese making, and milk chemistry, you can try a book, there are plenty of them. I recommend you the following (the first one is very good, but it is a scientific book, I never read the second one, but it seems is good and easy to understad even if you don't know about food chemistry.

Fundamentals of cheese science

And that's how you make cheese!

Also, I found a kit for making mozzarella and ricotta; the reviews says the mozzarella is hard to do but that ricotta was a success, so if you do not get ricotta using my method, maybe you should try this kit:

That's all, if you liked this instructable do not forget to rate it with a + and thank you for looking



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      12 Discussions


      5 years ago on Introduction

      I really enjoyed the quality of your instructable; you put in so many pedagogical aids...infographics, glossary, multiple well selected photos on and on. Question; how does pastuerization reduce the calcium content of milk as you state in Step 7?

      beehard44lil jon168

      Reply 9 years ago on Introduction

      heat the milk till it's near boiling and add lemon. kill the flame and cover for 30 minutes, cut up the curds and continue with step 10


      9 years ago on Introduction

      Great 'ible. I've been reading about the health benefits of whey protein (claims for muscle building/maintaining, insulin regulation, etc.) Instead of, or in addition to, ricotta, could the whey liquid be processed into a "sports drink"? How much protein would it contain?


      11 years ago on Introduction

      Thank you for the instructable! Quick question: Where can I find calcium chloride locally? I know it's available online, but where can I go pick it up in a store? I live in Northern California, of course, my town probably looks very much like your town. Your anticipated wisdom and knowledge is greatly appreciated! :)

      2 replies

      11 years ago on Introduction

      An amazing job- I especially like the use of a glossary for individuals like myself that are a bit lacking in the necessary knowledge base. :P


      12 years ago on Introduction

      That was an intense tutorial! I'll have to try it.

      Check here for info on inserting clean-looking links.

      1 reply