How is Vegan (Animal-Free) Whey Protein Made?

3 Ways

This article describes the 3 different methods of making vegan whey protein.

We cover the main method,

The one used to produce all currently available animal-free whey,

In detail (9 steps).

Let’s go.

This article is based on 47 research papers, 11 patents, and 3 regulatory documents. Last updated: Sep. 16, 2023. See our disclaimer.

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An image that shows that animal-free whey protein can be produced by fungi, plants, or bacteria. Fungi-derived whey protein is already out on the market. Plant-derived whey protein is patented. Bacteria-derived whey protein is in research.

Summary

Vegan whey protein is made by genetically-modified fungi (including yeast), plants (or their cell cultures), or bacteria. The production of vegan (animal-free) whey protein doesn’t directly involve the use of animal milk taken from animals, but it might involve animals in other ways. 1–17

An image that shows that vegan whey protein can be produced by fungi, plants, or bacteria. Fungi-derived whey protein is already out on the market. Plant-derived whey protein is patented. Bacteria-derived whey protein is in research.

Right now (at least in the U.S.) all of the vegan whey protein that you can buy…

Is made by yeast (and other fungi), not plants or bacteria.

So we’re going to focus most of this article’s details on yeast-derived whey protein as a result but we do cover the plant and bacterial methods as well.

In summary:

  1. Scientists take a piece of bovine DNA that encodes the whey protein
  2. They insert that DNA into the yeast
  3. The yeast are fed simple sugars
  4. The yeast use fermentation (“brewing”) to secrete the whey protein
  5. The whey protein is filtered out of the brew
  6. Other ingredients are added to the whey protein (like enzymes and flavors)
  7. The vegan whey protein powder is sold in stores

That’s the gist of it.

Oh,

Just like there’s no single way to make a blueberry pie…

There’s no single way to make animal-free whey protein.

While we cover the steps used to make vegan whey protein without giving you a massive headache…

…Keep in mind that the exact details will vary a little bit from manufacturer to manufacturer and from product to product.

An Asterisk on Terminology

We’re going to use the terms “vegan whey protein” and “animal-free whey protein” interchangeably in this article.

But…

With an asterisk (*) in mind.

We use these two terms only because they have become the popular ways by which to refer to whey protein that’s produced by non-animal life forms.

As you’re about to learn…

That doesn’t automatically mean that animals are not involved in the production of vegan whey.

animal-free whey protein may or may not be considered vegan due to three major factors in products labeled as vegan whey protein: the use of animal DNA, the possible use of animal substrates, and the possible need for animal testing.

More accurately:

It simply means that animal milk, taken from animals, is not the direct source of the whey protein.

Take a look:

We define vegan whey protein and animal-free whey protein…

And discuss our concerns related to the possible misuse of these terms…

In this article:

Vegan Whey Protein & Animal-Free Whey: Terms to Know

That’s for another day.

Let’s talk about how vegan (animal-free) whey protein is made.

How To Make Vegan Whey Protein Using Yeast

Step #1: Select Your Fungus

An image that shows that the biological kingdom of fungi includes yeast, molds, and mushrooms.

Quick note to keep in mind:

There is no one fungus nor one yeast. Fungi are a collection of diverse organisms, and they include many types of yeast, molds, and mushrooms. 18,19

Ok:

Let’s say you’re a chef who cooks in a high-stakes environment…

Would you rather rely on well-known, tried-and-tested, ingredients when it really matters?

Or risk using a completely unknown ingredient?

Probably the tried-and-true.

Companies that make vegan whey protein are no different.

They are more likely to use a fungus that’s familiar and reliable.

Such fungi tend to fulfill a few criteria: 1–11,20

  • They’re productive; not all fungi are good at making whey protein.
  • They’re unlikely to cause allergies or disease in people. Who needs lawsuits?
  • They are well-studied, so you don’t have to do a lot of research from scratch.
  • They’re already used in food applications. This minimizes any regulatory headaches by agencies like the U.S. Food and Drug Administration.

An image that shows that the traits of an ideal fungus for the production of whey protien includes productivity, harmlessness, a high-researched strain, and a fungus already used in food applications.

Given that,

Here are three real-world examples of fungi used to make animal-free whey protein: 1–11,20

  • Trichoderma reesei, a type of filamentous fungus
  • Komagataella phaffii (aka Pichia pastoris), a type of yeast
  • Saccharomyces cerevisiae, also called brewer’s yeast or baker’s yeast

We’ll just say you picked a yeast for the rest of this example.

Let’s move on.

Step #2: Get The Whey Protein’s Code

You can use any dairy animal’s genetic code for this step. We’ll use a cow as our example.

There are two major protein families (collections of proteins) in cow’s milk:

Casein and whey.

Casein accounts for ~80% of milk proteins and whey accounts for ~20%.21–23

An image that shows that casein makes up about 80% of bovine milk proteins while whey makes up about 20% of bovine milk proteins.

There are a lot of different types of whey protein…

The two major forms of whey protein are: 22,24–26

  • Beta-lactoglobulin (BLG). This protein accounts for about 50-58% of all whey protein.
  • Alpha-lactalbumin (ALA). This protein accounts for about 17-20% of whey protein.

Typically:

When you hear that a company or a scientist used fungi to make whey protein, they likely produced BLG and/or ALA. 1–11

We’ll use BLG as our example from now on.

So:

You know how you reference a recipe for a set of instructions on how to make a blueberry pie?

Yeast need a set of instructions too…on how to make whey protein.

The instructions that yeast use to make whey protein aren’t provided by a book. They’re provided by genetic code (DNA).

Not just any genetic code.

But the very specific snippet of genetic code that tells an organism exactly how to make BLG and nothing else.

An image that shows that a very specific sequence of genetic code encodes a very specific whey protein.

Two things:

First:

Yeast don’t naturally possess this snippet of BLG code.

After all…

They are fungi, not a cow.

So the yeast never evolved to make BLG.

Second:

Obviously the yeast can’t access this snippet of genetic code on their own in order to make whey protein.

This is where scientists come in and lend a helping hand.

An old school method of getting this genetic code is by, quite literally, harvesting it from a cow’s mammary (milk-producing) gland using a bunch of fancy lab techniques. 9,17,27,28

An image that shows that scientists can take a sample of animal tissue and extract whey protein DNA from that tissue.

But…

Scientists sequenced (“mapped out”) the code for BLG decades ago.9,17,27–29

Nowadays you can just download…

…Yes you read that right…

Download…

The code for BLG from an online database much like you can download a recipe for blueberry pie.

The code for BLG spells out a very specific sequence of amino acids, unique to BLG. Amino acids are the building blocks of proteins.

For example, here’s the amino acid sequence for bovine BLG. 29

An image of the amino acid sequence of bovine beta-lactoglobulin from Uniprot.

A very specific sequence of 162 amino acids equals one BLG protein molecule.3,4,30

Nerd alert:

Keen eyes will have spotted that the image above actually contains 178 amino acids. Good catch.

178 amino acids encode BLG but 16 amino acids are chopped off before the protein makes its way into the milk. That leaves 162 amino acids per one BLG protein that you consume.

Step #3: 3D Print The Code

An image that shows that a specialized 3D printer can physically make a section of genetic code using nucleotide-based inks.

Ok.

Now you have your BLG code in digital format.

Then what?

You have to create a physical version of this code, one that you’ll then insert into the yeast.

There are multiple ways of doing this. 31,32  But this isn’t Bio 401 class so we’ll only cover a bit of one method here as a highlight.

One of the coolest ways to create this physical code involves 3D printing. 33

Yep.

3D printing.

Physical genetic code (DNA) might be microscopic but it’s still three dimensional.

So you send the 3D printer a digital file of the BLG code and press “print” (we’re simplifying here, of course).

Video credit: @DNAScript

This printer has a set of four inks.

They are not black, red, blue, and yellow as per your inkjet printer at home.

The inks are called Adenine (A), Thymine (T), Guanine (G), and Cytosine (C).

A, T, G, and C represent specific nucleotides.

In a gist, nucleotides are like the “letters” of the genetic code.

If your recipe for blueberry pie reads S-U-G-A-R, you know what those letters spell out and so you understand the instructions and therefore you know how to make a great pie.

Same goes for nucleotides and yeast.

The A’s, T’s, G’s, and C’s spell out what the yeast must produce.

In short,

Using these inks,

The 3D printer transforms the letters of the digital BLG code into the letters of the physical code that the yeast cells then “read” as a set of instructions on how to make BLG.

Here’s another quick video on a few ways by which Step #3 can work, including 3D printing:

Video credit: @GenScriptUSAInc

Step #4: Insert The Code Into The Yeast

Now that you have your physical code, you have to insert that code into your yeast.

The natural strain of yeast you’ve chosen is called the “host strain” and its genetic code is called the “host strain genome”. 3,4

The genome is the entire genetic code of an organism; the BLG code is just a tiny portion of that. 34

In other words, we’re not changing the yeast cell’s entire genome (entire genetic code) here.

Instead,

We’re modifying a tiny section of it.

In Step #4, scientists use a bunch of techniques to insert the BLG code into the yeast’s genome.

The exact steps all depend on case specifics.

For example:

A combination of chemicals, enzymes, temperature changes, and additional genetic code can be used to “force” an accurate copy of the BLG code into the yeast cell’s genome. These chemicals may (but don’t have to) include substances like glycerol; which can be derived from plant, animal, or synthetic sources. 3–6,8–12,21,35–37

An image that shows a simple image of how whey protein's genetic code is inserted into the yeast cell's DNA.

Nerd alert:

It’s possible to insert more than one copy of the BLG code into the host strain, which may help the yeast produce more of the BLG.3,4,10

Once the BLG code is incorporated into the yeast cell’s genome, then: 3–6,8–12,21,38

  • The host strain is now called the “production strain”, the strain used to produce the whey protein.
  • The production strain is a genetically modified organism (GMO) since we modified the host strain’s genome.
  • The BLG code located inside the yeast cell’s genome is called “rBLG” to distinguish it from the original BLG.

The “r” in “rBLG” stands for “recombinant”.

Recombinant basically means we combined one code (for the BLG protein in this case) with another (the yeast’s genetic code in this case).

So the resultant BLG protein is a recombinant protein (rBLG),

A protein ultimately made thanks to the combination of genetic code from two different organisms. 39,40

However!

This is super important:

The code for BLG inside of the yeast remains identical to the code for BLG inside of a cow.

Think of the yeast’s genome as a gold chain and the code for BLG as a small silver link.

We can combine the silver link and the gold chain to make a “recombinant” chain.

But the silver link doesn’t change just because it’s now inside a gold chain. It’s still a silver link.

An image that shows a silver link inside of a gold chain, with the gold chain representing the yeast cell's genome and the silver link representing the whey protein's genetic code.

In practice,

This means,

The yeast produce an rBLG protein with the same amino acid sequence as the BLG found naturally in cow’s milk. 3,4

Step #5: Make a Seed Culture

Ok.

So now you’ve got your production strain ready to go.

Good.

What do you do next?

Well,

Yeast are already good at fermenting things to make bread, beer, kombucha, and a lot more tasty stuff. 41–45

No reason to rewrite the script here.

An image that shows that yeast are already used to ferment numerous substances in order to make bread, beer, and kombucha.

We’ll use yeast to ferment our way to whey protein as well. In other words, we’ll give the yeast a substrate (basically, a food source) and they’ll use that as a source of energy to multiply and create the whey protein.46

Our first step is to start a seed culture before going into full-scale production.

An image showing a seed culture with yeast inside of a nutrient broth.

Why make a seed culture?

There are numerous possible reasons, depending on specifics…

The gist of it is that a seed culture helps to optimize production and therefore decrease costs.47,48

One way to start a seed culture is to transfer your production strain of yeast into a seed culture vessel, where the yeast cells multiply. 3–5,9–12

The seed culture vessel contains a warm (~30 degrees Celsius) and nutritious broth of compounds…

You shake that broth at 240 rpm for 30 hours and…

You’re done making your seed culture.

The compounds that are used for the seed broth vary from method to method and company to company.

Such compounds can, for example, include glycerol (a type of sugar-alcohol). 10,37

Glycerol is also known as glycerin.

Glycerol can be derived from plant, animal, or artificial (petroleum-based) sources.35,36

Step #6: Produce (Ferment) The Whey Protein

So now you have enough seed culture to start your main fermenter.

Cool.

The basics of it is that you then tip that seed culture into the main fermenter.

And you provide the yeast with a solid food source.

In theory, yeast can use all sorts of stuff as a food source… 49–55

Including everything from plant sugars…

….to animal parts and products…

To industrial waste.

An image that shows that yeast can use many different types of substract (mangoes, prawns, industrial waste, and synthetic compounds) as their source of food.

This is one part of the process where vegan organizations have some qualms, since numerous different animal products might, but don’t necessarily have to be, used.

(Details: Is Animal Free Whey Vegan? What PETA & Surveys Say)

In practice:

The optimal food source depends on the species of yeast, the method of manufacture, and the desired end-product.

That means manufacturers are more likely to use very specific, highly pure, chemicals as a food source for the yeast.

Don’t get automatically scared by the word “chemicals” in this context…

Glucose, a simple sugar and a source of energy, is found in all sorts of natural foods and it’s technically a chemical…

And that’s mainly what we’re getting at here when we say “chemicals as a food source” for the yeast.

An image that shows a set of three fermentation vessels filled with yeast, glucose, and whey protein.

That being said,

The culture medium (the liquid broth the yeast live in) may contain other chemicals for a huge range of reasons related to growth, production, and safety.

(Related: Is Vegan Whey Protein Safe? What FDA Documents Reveal)

Some of these chemicals are also found naturally in foods…and others…not so much.

Here are some examples of the chemicals that may be added to the fermentation broth during the production of vegan whey protein:10–12,56,57

  • Glycerol, a sugar alcohol that can be derived from plant, animal, or synthetic sources.
  • Glucose, a simple sugar the yeast use as a very important source of food.
  • Galactose, another simple sugar that can help trigger the production of whey protein.

The most important takeaway from this step in the production process,

Is that the yeast use fermentation…

To produce whey protein…

Which the yeast secrete into the broth.

This allows you to separate the yeast away from the whey protein they have produced.

This whole process of using genetically modified organisms (like yeast) to ferment very precise ingredients (like BLG),

Is called precision fermentation.

(Related: Precision Fermentation Statistics.)

Step #7: Purify The Whey Protein

To fully separate the yeast from the whey protein they’ve secreted, you can use a combination of 3 general methods: 10–12,56,57

Spin the broth at high speed, adjust the broth’s acidity, and use a filter.

In our example, we’ve pretended to make the BLG whey protein.

Let’s stick with that.

First:

You spin your broth (sometimes multiple times)…this “spinning” is called centrifugation.

Centrifugation separates the yeast cells away from the rest of the broth that contains the BLG.

An image that shows a centrifuge spinning two vessels with a resultant vessel displaying the separation of yeast cells away from the supernatant (which contains the whey protein).

Second:

After you’ve finished spinning the broth…

You discard the pellet that contains the yeast cells.

What remains of the broth, which contains the BLG, is now called the “supernatant”…

You then adjust the pH (“acidity”) of this supernatant to a pH of 5.2.

(That’s around the acidity of a pickle)58

An image of a glass vessel containing a white liquid within which are suspended whey protein aggregates with green circles representive acidic compounds. Lowering the pH (raising the acidity) of the supernatant undissolves the whey protein within it.

Why change the acidity?

Well…

The BLG is basically dissolved in the supernatant.

It’s like mixing sugar in tea. The sugar “disappears” or “dissolves” into the tea even though it’s technically still there, you just don’t see it.

But when you change the acidity to a pH of 5.2, you reach BLG’s “isoelectric point”. 30

This is the point at which BLG will “un-dissolve” (precipitate or coagulate out) and turn into a solid in the broth. 59

Have you ever seen solid stuff floating in a bottle of spoiled milk? Those solids are milk proteins. As milk spoils, its acidity increases, which causes the proteins to “undissolve” and turn into visible chunks.

Anyways,

Once you reach BLG’s isoelectric point, you then use more spinning and filtration to further purify the BLG.

Once you’ve got a batch of purified BLG, you dry it…

And then you’re left with a white to off-white powder that’s at least 80-90% BLG.

An image that shows that yeast-derived whey protein powder is white to off white in color.

Step #8: Animal Testing?

At this point, you may or may not be able to sell your whey protein.

It all depends on the laws, regulations, and rules set out by governing bodies like the U.S. Food and Drug Administration.

Some governing bodies require animal testing before they let you sell your product.

And some may not.

All of this depends on country-specifics,

As well as the method of manufacture you use,

And the final product in question.

Naturally, animal-rights organizations are worried about this part of the process.

(Details: Is Animal Free Whey Vegan? What PETA & Surveys Say)

An image of a white bunny on a green circular background that says

Note:

When we say animal testing, we don’t mean that every bag of protein powder is tested on animals.

What we mean is that prior to being given the green light for mass production, you might have to test your new product on animals.

Once you get the green light,

Then…

For that specific product, made in that specific way,

You won’t have to test every batch on animals.

Step #9: Create The Final Product

Wow…

We’ve done a lot here.

But we’re not finished.

We have to sell this powder.

As you probably know, pure plant protein or pure dairy-whey protein powder is hard to find…

Most protein supplements have a lot of ingredients added in,

Probably because most people don’t find pure plant or whey protein very tasty or easy on their stomach.

So at this point you’ll need to decide what you want to mix in:

  • Sweeteners
  • Other proteins
  • Digestive enzymes
  • Colors

Your choice.

After that,

You’re done.

An image showing that a final whey protein product contains the whey protein mixed with sweeteners, enzymes, and other ingredients.

Depending on the manufacturing choices you make, 3,4,10–12,33,60

Like the (one or more) genetic modifications you make and the fungi you select,

Your finished product will be made from:

One or more types of whey protein…

Secreted by one or more types of fungi.

Is Animal-Free Whey Protein Vegan?

Animal-free whey protein (sometimes called vegan whey protein) may or may not be truly vegan and animal-free, depending on a combination of your personal views and the exact manufacturing details that are specific to each product.

To recap:

We demonstrated in the 9 steps above that there are several phases whereupon an animal, or animal-derived product, may be introduced or used for the manufacture of animal-free whey protein.

This includes:

1. During research and development: the possible use of animal tissue to derive the necessary genetic code for the whey protein.

2. During manufacture: the possible use of substances and chemicals derived from animal sources.

3. Before regulatory approval for sale: the possible need for animal testing.

The only way to know for sure…

Whether any animals or animal-derived products were used in any way…

Is to ask the manufacturer of the product in question.

Plant-Based Whey Protein

The general idea behind making whey protein from plants is very similar to everything we’ve described so far for making yeast-secreted whey protein.

Except,

The ending here can be very different from yeast-derived whey protein.

So,

Here’s how plant-based whey protein is made: 13–15,61–63

Step #1:

Select your plant of choice.

This can be an actual plant, like soy that will later grow in a field…

Or it can be a plant’s cells (like carrot cells) that’ll grow in lab-like conditions during manufacture.

An image that shows soybean plants and soybeans, as well as a lab beaker and petri dish, to signify that genetically engineered plants can created whey protein as well.

Step #2:

Select one or more whey proteins of choice, like beta-lactoglobulin (BLG).

Step #3:

Obtain the genetic code for your whey protein of choice…

Step #4:

Insert that code into the plant’s genome.

Video credit: @US_FDA

So far that’s pretty much what we did with the yeast so those steps should look familiar to you.

Of course…

You know that the details of each step vary a bit since yeast are not plants.

What comes next though…

Is the really interesting part.

Possible Ending #1:

As the genetically modified plant grows, it produces whey protein inside one or more of its parts,

Like its seeds or roots.

Then,

You harvest the plant and use a combination of methods to separate the whey protein from the plant compounds.

For example,

You can use enzymes (bioactive proteins),

Chemicals,

Or physical processes (heat, presses, centrifugation, etc)…

To separate the whey protein away from the plant itself.

Often, you’ll use a combo of many such methods.

And so that purifies the whey protein into a powder you can sell as a supplement.

Possible Ending #2:

You don’t bother with possibility #1.

Maybe you think it’s too messy or it takes too long.

Instead, you simply eat the genetically modified plant as is.

For example:

Let’s say you modified soy to produce whey protein inside the soybean.

You harvest the genetically modified soybeans and eat them as normal.

That means you get all the benefits of the plant:

Like antioxidants, vitamins, minerals, and healthy plant oils…

As well as the benefits of the whey protein inside the genetically modified soybean.

Some plants can even be modified to secrete the whey protein in liquid form, kind of like the syrup that comes out of a maple tree.

So, in theory, you could drink the whey protein without having to mix a powder with water.

(But that’s only an idea at this point.)

Bacterial Whey Protein

An image that shows the pros and cons of bacterial whey protein production. Pros: cost-effective and easy. Cons: low protein yield and hard to purify.

Although researchers have made it clear that bacteria can produce whey protein and even patents to that effect exist, 1,7,8,16,17,64–66

We’re not aware of any vegan whey protein powder,

Made by genetically modified bacteria,

That’s available for sale in the U.S.

Why not?

Although the steps involved in the production of bacterially-derived vegan whey protein are,

At a high level,

Similar to that of yeast-derived whey protein described above…

Challenges remain.

Here’s a couple of them:

It’s actually pretty cheap and easy to modify bacteria in order to produce whey protein.

But:

As things stand right now, 1,7,8

The bacteria can’t produce enough of the whey protein to justify their relatively low cost…

And it’s quite difficult to purify the whey protein produced by bacteria.

That doesn’t mean scientists aren’t working to overcome these and other challenges, they totally are.

Still,

It may take some time before we see a whey product, made by genetically-modified bacteria, available for sale.

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