Rennet

Rennet is a complex mixture of enzymes found in the stomachs of ruminant mammals and has been central to cheesemaking for millennia. Its primary function is to coagulate milk by acting on casein proteins, thereby separating solid curds from liquid whey. While traditional rennet was obtained from the stomachs of young calves, modern cheesemaking frequently relies on alternative sources, including microbial and fermentation-produced variants. The science, processes, and applications of rennet continue to play a critical role in both artisanal and industrial dairy production.

Composition and Biochemical Role

Rennet is composed of several enzymes, the most important of which is chymosin, a specialised protease responsible for initiating curd formation. Other enzymes present include pepsin and lipase, each of which contributes additional proteolytic and lipolytic activity during cheese ripening.
The central mechanism of rennet coagulation involves the cleavage of κ-casein, a stabilising milk protein. Casein occurs in micelles, structures whose surface carries slightly negatively charged glycomacropeptides (GMP). These charged regions cause mutual repulsion between micelles, allowing milk to remain liquid. Chymosin removes GMP from the κ-casein chain, eliminating the negative charge barrier. The destabilised micelles then aggregate, trapping milk fat globules, minerals, and other proteins to form a firm curd.
This enzymatic action is further influenced by minerals such as calcium and phosphate. Their presence enhances micellar aggregation and curd strength, which is why cheesemakers may add them, especially when processing milk naturally low in calcium, such as goat’s milk.

Traditional Extraction from Calves

Historically, rennet was sourced from the abomasum, the fourth stomach of unweaned calves whose digestive systems naturally contain high concentrations of chymosin. These stomachs, often a by-product of veal production, were cleaned, dried, and used to produce rennet extracts.
The traditional process involved slicing dried stomach tissue into small pieces and steeping them in salted water or whey. Acidifying agents such as vinegar or wine were added to mimic stomach acidity and activate the enzymes. After several hours or days, the mixture was filtered, leaving a crude rennet solution. Typically, one gram of this solution could coagulate two to four litres of milk, demonstrating the potency of naturally occurring chymosin.
Older animals yield rennet with lower chymosin content and higher concentrations of pepsin, which limits its suitability to certain specialised cheeses. Each ruminant species produces enzymes adapted to its own milk, leading to the use of species-specific rennets, including goat and sheep variants, in traditional dairy regions.

Modern Industrial Production

Industrial rennet extraction employs controlled biochemical and mechanical processes to enhance consistency and potency. Deep-frozen stomach tissues are milled and mixed with extraction solutions to release enzymes. Activation occurs through the addition of acid, as stomach enzymes are naturally produced as zymogens requiring low pH to become active.
The extract then undergoes neutralisation, filtration, and concentration. Standardised preparations typically achieve a potency of 1:15000, meaning one gram of extract can coagulate fifteen kilograms of milk. Only a tiny portion of the final cheese—approximately 0.00003 grams per kilogram—consists of rennet enzymes, the remainder being water, salts, and sometimes preservatives such as sodium benzoate.
Although animal rennet remains available, its use has decreased globally. In the United States, for example, fewer than five per cent of cheeses are now made with traditional animal rennet, with most producers preferring biotechnological alternatives.

Plant-Derived Coagulants

Plants with coagulating properties have been used since antiquity. Ancient Greek sources describe the use of fig sap, while various herbs and plants, such as species of Galium, thistles, Urtica, Malva, Withania coagulans, and Glechoma hederacea, have long been used in specific regional cheeses. Mediterranean cheesemaking traditions, particularly in Portugal and Spain, still use thistle enzymes to produce distinctive soft cheeses with characteristic flavours.
Although referred to as “vegetable rennet”, most commercial products marketed under this term actually contain mould-derived enzymes rather than plant extracts. Industrial-scale production of truly plant-based rennet is limited due to challenges in standardisation and potential for bitterness in long-aged cheeses.

Microbial Coagulants

Microbial rennet emerged as an alternative source when the demand for animal rennet exceeded supply. Certain moulds, particularly Rhizomucor miehei, produce proteolytic enzymes capable of coagulating milk. These cultures are grown in fermentation tanks, and the resulting enzymes are purified to remove undesirable flavour compounds.
Historically, microbial coagulants were associated with bitterness, especially in aged cheeses, due to the presence of secondary proteases. Advances in microbiology and purification processes have significantly improved flavour outcomes and consistency. Microbial rennet is widely accepted for vegetarian and some vegan cheese production, provided no animal-derived substrates are used in the fermentation process.

Fermentation-Produced Chymosin (FPC)

The most transformative development in modern rennet technology is fermentation-produced chymosin. Using genetic engineering, the gene responsible for chymosin production is inserted into microorganisms such as bacteria, fungi, or yeasts. These organisms generate pure chymosin during controlled fermentation. After fermentation, the microorganisms are removed, ensuring that the final enzyme preparation contains no genetically modified material.
First commercialised in 1990 and originally developed by biotechnology researchers associated with Pfizer, FPC became the first bioengineered enzyme approved by the US Food and Drug Administration. Its adoption expanded rapidly due to its purity, consistency, and high performance. By the late 1990s, around 60 per cent of hard cheeses in the United States were produced using FPC, rising to as much as 90 per cent of global rennet use by 2017.
Leading FPC products include CHYMAX, produced using Aspergillus niger, and Maxiren, produced using Kluyveromyces lactis. FPC provides predictable curd formation, higher yields, improved texture, and reduced bitterness. It is suitable for kosher and halal certification and is generally accepted for vegetarian diets when produced without animal-based substrates.

Non-Rennet Coagulation Methods

Not all cheeses require rennet. Many soft cheeses are produced through acid coagulation, either by adding acid directly—such as vinegar, citric acid, or lemon juice—or through the natural production of lactic acid by bacterial cultures. Examples include paneer, cream cheese, rubing, and numerous traditional fresh cheeses. These methods do not produce the elastic curd typical of rennet-coagulated cheeses but instead yield softer, more crumbly textures.
Vegan cheesemaking avoids animal milk entirely, using bases such as soy, nuts, grains, or root vegetables. Coagulation may still involve acids or microbial enzymes, depending on the desired texture and flavour profile.

Cultural and Mythological References

Beyond its scientific and industrial relevance, rennet features in mythology. In Yazidi cosmology, the coagulation of the Earth itself is described through the metaphor of rennet flowing from the White Spring of the celestial realm into the primordial ocean, symbolising the transformation from fluidity to solid form. Such references highlight the ancient recognition of rennet’s transformative properties.