Precision fermentation is a method that uses tiny living things called microorganisms to create specific molecules. This process can make food ingredients usually found in animals and plants, such as proteins, fats, sugars, and other substances. Microorganisms like bacteria or yeast are used to make large amounts of a particular compound. This compound is then taken out and cleaned from the liquid or broken cells where it was made. Examples include proteins from milk or eggs, fats from dairy, sugars that help the body function, and substances that give food flavor, color, or nutrients.

Precision fermentation is different from other fermentation methods because it focuses on making one specific molecule with high accuracy and purity. In other methods, such as traditional or biomass fermentation, the final product includes many different substances, not just one. This precision happens by carefully controlling the growing conditions and using specific types of microorganisms. Sometimes, scientists use naturally occurring microorganisms, but in other cases, they change the microorganisms in the lab through techniques like altering their genes or using special tools to improve their abilities. The cleaning process ensures that any genetically changed microorganisms are not in the final product.

The term "precision fermentation" is not very old, but the technology behind it has been used since the 1980s. For example, proteins like insulin, which helps treat diabetes, and chymosin, used to make cheese, have been made using these methods for many years and are widely used. Precision fermentation uses tools that change genes, science that designs new biological systems, and techniques to improve microorganisms. It is expected to play an important role in future food production that uses natural resources. It may become a key technology as the world moves toward more sustainable food systems, especially as climate change affects farming and when there is not enough land for agriculture.

Principles of precision fermentation

A fermentation bioprocess includes all the steps needed to change raw materials into the desired molecule. This process involves selecting the type of microorganism that will perform the transformation and setting the conditions for its growth.

Feedstocks are the main raw materials that provide carbon, nitrogen, and energy for microorganisms to grow and create products. Choosing the right feedstock is important because it affects the cost and environmental impact of the final product.

• First-generation sugars are often used in precision fermentation. These are refined sugars from food crops, such as glucose. They support strong microbial growth and are safe for food use. However, they are expensive and compete with food supplies.
• Second-generation sugars come from non-food sources, like plant waste. These sugars reduce competition with food and make fermentation more sustainable.
• C1 feedstocks include carbon dioxide, methane, formate, and methanol. These are very sustainable because they help reduce environmental harm. However, using them in large-scale production still has challenges.
• Food industry sidestreams, such as wastewater or byproducts from food processing, can be used as low-cost feedstocks. They can be turned into fermentable sugars for precision fermentation in a circular economy approach.

A cell factory is a biological system, such as microorganisms, plant cells, or mammalian cells, that changes substrates into valuable products like proteins, enzymes, vitamins, medicines, and materials under controlled conditions. Common microorganisms used include Bacillus subtilis, Corynebacterium glutamicum, Escherichia coli, Komagataella phaffii, Saccharomyces cerevisiae, and Yarrowia lipolytica. Each of these has unique traits that make them suitable for specific production tasks. The choice of microorganism depends on factors like the presence of natural pathways to make the desired product, the ability to produce foreign molecules efficiently, the safety of the organism, and its compatibility with growth conditions.

• Bacillus subtilis is a type of bacteria that can survive harsh conditions and use low-cost materials. It grows quickly and is safe for use (GRAS). It can produce many types of biomolecules.
• Corynebacterium glutamicum is a fast-growing bacteria that can use various sugars, acids, and alcohols. It is known for making amino acids and other valuable compounds.
• Escherichia coli is a rod-shaped bacteria. It is widely used because it is easy to modify genetically, has a simple process for adding new genes, is inexpensive, and produces large amounts of desired products.
• Komagataella phaffii (formerly Pichia pastoris) is a yeast that is good at making natural products. It is easy to modify and works well for producing proteins.
• Saccharomyces cerevisiae grows quickly, produces high amounts of products, and can handle tough conditions like low pH or limited oxygen. It is safe (GRAS) and has tools for genetic changes, making it useful in industry.
• Yarrowia lipolytica is a yeast that can use many different materials and adapt to various environments. It is a promising choice for making proteins, lipids, flavors, and pigments.

Microorganisms are often modified to improve the production of a target molecule. Metabolic engineering involves changing the chemical reactions inside cells to increase the production of a specific molecule.

The target molecule is usually a lipid, carbohydrate, or other compound made through a series of chemical reactions. These reactions transform materials from the feedstock into the desired product using intermediate steps. Some reactions are part of the microorganism’s natural processes, while others use enzymes from different organisms. Metabolic engineering changes both natural and foreign reactions to maximize product creation. For example, adjusting the activity of key enzymes in the microorganism’s metabolism can increase production and reduce unwanted byproducts. Synthetic biology tools, like promoters of different strengths, help control how much of a molecule is made. Scientists also use math and computer models to predict how changes will affect the process.

In some cases, the target molecule is a protein. To produce it in large amounts, genetic methods are used, such as using strong promoters or timing the protein’s production with an inducible promoter. Techniques may also be used to make the protein leave the cell, which helps in collecting it.

Process optimization in precision fermentation aims to improve how efficiently, sustainably, and productively target compounds are made. This involves improving parts of the fermentation process, such as the culture medium, feeding methods, bioreactors, and growth conditions.

The culture medium’s composition is crucial. It usually contains a carbon source, nitrogen source, minerals, and water. Depending on the microorganism, extra nutrients like growth factors or vitamins may be needed. Alternative carbon sources, such as agricultural waste or industrial byproducts, can be used to turn waste into valuable products.

Feeding strategies include batch, fed-batch, and continuous methods. Each has its own benefits and drawbacks. Choosing the right method based on the fermentation product can improve efficiency.

Different bioreactors, such as stirred tanks, airlifts, wave systems, and membrane reactors, have unique features related to mixing, oxygen transfer, stress on cells, scalability, and cost. Selecting the right bioreactor depends on the microorganism and the product’s needs.

Fermentation conditions, such as pH, temperature, mixing speed, air supply, and the size and age of the initial microorganism population, can be adjusted during production. With the help of artificial intelligence, models can be created to predict and control these conditions, making the process easier, more efficient, and requiring less manual work.

Food applications

Vitamins, antioxidants, and other nutrients are made using microorganisms. For example, vitamin B2 is often produced using a method called precision fermentation, which is widely used in manufacturing. Other compounds are still made in small amounts or only studied in scientific research, and more work is needed to make them suitable for large-scale production.

• Astaxanthin is an orange-red compound with antioxidant and anti-inflammatory properties. It helps prevent health issues like high blood pressure, high blood sugar, and high cholesterol. Microorganisms such as E. coli, Y. lipolytica, and S. cerevisiae are used to make astaxanthin.
• Kaempferol and quercetin are compounds made from naringenin. They have antioxidant, anti-inflammatory, and other health benefits. These compounds have been made using S. cerevisiae, Y. lipolytica, Streptomyces, and E. coli.
• Polyunsaturated fatty acids (PUFAs), such as EPA and DHA, are important for health and help prevent heart disease and support brain development. Microorganisms like E. coli, Y. lipolytica, C. reinhardtii, and Aurantiochytrium are used to produce PUFAs.
• Resveratrol is an antioxidant found in plants like grapes and peanuts. It is used in food, cosmetics, supplements, and medicines. Microorganisms such as E. coli, Y. lipolytica, and S. cerevisiae are used to make resveratrol.
• Vitamin A is a fat-soluble vitamin needed for growth and metabolism. Scientists have created ways to make vitamin A using E. coli, Y. lipolytica, and S. cerevisiae.
• Vitamin B5, also called D-pantothenic acid, is a nutrient needed for body functions. E. coli and C. glutamicum are the best microorganisms for making D-pantothenic acid.
• Vitamin B2, or riboflavin, is used as a supplement, in animal feed, and as a food coloring. Precision fermentation is now used to make it at an industrial scale, replacing older methods. It is made using A. gossypii and B. subtilis.
• Vitamin B12, or cobalamin, is a vitamin the body cannot make and must be obtained from food. Scientists have made 21.09 mg/L of vitamin B12 using E. coli.
• Vitamin C, or L-ascorbic acid, is important for body reactions. Scientists have created a way to make vitamin C using S. cerevisiae.
• Vitamin D is a hormone that helps the body absorb calcium and phosphate. S. cerevisiae is used to make vitamin D.
• Vitamin E, made of tocopherols and tocotrienols, is an antioxidant. It is made in plants and microorganisms.

Microbial pigments and flavors made by bacteria, fungi, yeast, and algae are being used as eco-friendly alternatives to synthetic or plant-based dyes and flavors in many industries.

• Carotenoids: These are yellow-orange pigments with antioxidant properties used in food, drinks, cosmetics, and medicines. Examples made using precision fermentation include β-carotene, zeaxanthin, lycopene, astaxanthin, canthaxanthin, and lutein.
• Heme: This is a red pigment and iron source found in meat. It is now used in lab-grown meat and plant-based alternatives.
• Indigo: A blue pigment used as a sustainable alternative to synthetic dyes in textiles or electronics.
• Limonene: A fragrant compound found in citrus fruits.
• Linalool: A compound with a lavender-like scent found in over 200 plant species.
• Methyl anthranilate: A compound with a grape-like scent and flavor.
• Nerol: A compound with a floral-citrus scent found in plant oils.
• Phycocyanin: A blue pigment with antioxidant and immune-boosting properties used in foods and supplements.
• Prodigiosin: A red pigment used as an antimicrobial and anticancer agent.
• Raspberry ketone: A flavor compound found in raspberries, cranberries, blackberries, and peaches.
• Riboflavin (Vitamin B2): A yellow pigment important for body functions and used in supplements, energy drinks, and animal feed.
• Vanillin: A compound that gives vanilla its flavor.
• Violacein: A purple pigment with antibacterial and antifungal properties.

Sustainable food ingredients made using precision fermentation include carbohydrates, proteins, and fats.

As the world’s population grows, finding high-quality protein sources has become more important. Alternative proteins can come from plants, lab-grown animal cells, or fermentation. Precision fermentation offers a low-carbon, sustainable way to make proteins. Proteins like casein, lactoferrin, lactalbumin, lactoglobulin, and ovalbumin have been made using this method and are used in food.

Dairy proteins such as casein and whey are made by companies for use in milk, yogurt, cheese, and ice cream. Some companies also make human milk proteins, focusing on their immune benefits. Egg white proteins are made using microorganisms and used in food, drinks, plant-based meats, and baking. For example, Impossible Foods uses a soy-based protein called leghemoglobin to improve the taste and look of plant-based meat, like the Impossible™ Burger.

Oligosaccharides like fructooligosaccharides (FOS), galactooligosaccharides (GOS), and human milk oligosaccharides (HMOs) help support gut health and are used in infant formula. Fermented sugar alcohols like erythritol and xylitol are used as sweeteners in healthy foods and are made using engineered yeast or E. coli.

DHA and omega-3 fatty acids are important for brain and eye health and are used in infant formulas and supplements. EPA

Non-food applications

Biofuels made using precision fermentation with renewable resources provide a sustainable option instead of fossil fuels. Examples include alcohol-based biofuels (butanol, isobutanol, and isopropanol), hydrocarbon-based biofuels (alkanes, alkenes, biodiesel, and fatty acid ethyl esters), gaseous biofuels (hydrogen and methane), and advanced biofuels like jet fuels.

Many sustainable biomaterials have been created using engineered microorganisms. These materials are used in textiles, packaging, cosmetics, and medical devices. Examples include biodegradable plastics, alternatives to collagen and gelatine, spider silk proteins, microbial cellulose, and coatings.

Industrial chemicals produced through precision fermentation include solvents (acetone and isopropanol), monomers for polymers and resins (1,3-propanediol and 1,4-butanediol), precursors for biodegradable plastics and specialty chemicals (lactic acid, succinic acid, and itaconic acid), and bio-based polyamides (1,5-pentanediamine). These chemicals offer sustainable alternatives to traditional methods, reducing environmental harm.

Precision fermentation allows for the efficient and large-scale production of animal-based medicines such as insulin, vaccines, antibodies, and recombinant proteins without depending on animal farming. It also enables the creation of plant-derived medicines like alkaloids, cannabinoids, and antimicrobials, reducing land and water use and environmental impact. This method can also produce certain medicines that are typically made through chemical processes. Examples include taxol, anti-malarial drugs such as artemisinin, and lanthipeptides.

Economic and environmental considerations

Microbial products made through precision fermentation can help reduce the environmental impact of industries, farming, and waste management. These products, such as alternative proteins, are green alternatives to animal-based proteins from farming, which produces 20% of the world’s greenhouse gas emissions. Using microorganisms to create materials like biomaterials, medicines, or biofuels instead of chemicals from fossil fuels can lower emissions. Engineering microorganisms to produce valuable products using gases like methane and carbon dioxide is a major scientific advancement.

Producing high-value microbial products from agricultural waste through a circular bioeconomy method can reduce harmful pollutants and toxic substances released into the environment. These products can also reduce freshwater use by replacing items made in farming, which uses 70% of the world’s freshwater. Microbial products made in bioreactors need much less land than farming or animal agriculture. This freed-up land could be used for reforestation, supporting biodiversity, and reducing the need to import animal feed.

Although precision fermentation is better for the environment than farming or fossil fuel-based products, solving some environmental challenges will be important to fully use its potential.

Precision fermentation needs a lot of energy for heating, cooling, mixing, and processing. Ways to make the process more energy-efficient include: i) using renewable energy sources on-site, ii) improving fermentation conditions (like oxygen levels, pH, and temperature) to speed up production and save energy, and iii) designing better microorganisms that work well at low temperatures, need less oxygen, or produce purer products, which reduces energy use.

A key environmental factor in precision fermentation is the type of materials used to grow microorganisms. Many products now rely on sugars from sugar cane or corn, which can reduce the environmental benefits due to high water and land use, competition with food production, and the need for fertilizers and pesticides. To fix this, industries should use non-food sources like food waste, plant-based materials, or industrial gases like carbon dioxide. Fermentation also creates waste that must be handled properly to follow environmental rules.

The growing demand for sustainable, animal-free foods and products, along with the need to reduce environmental harm, has caused the precision fermentation market to grow rapidly. It was worth $2.1 billion in 2023 and is expected to reach over $100 billion by 2034. To lower costs, methods include: i) improving production efficiency through genetic changes, ii) using low-cost materials, and iii) making better methods to recover and purify products to save money.

Regulatory status

Although the term "precision fermentation" is new, the technology has been used for many years. As of 2024, there is no official or legal definition of "precision fermentation" in any country or region, and no specific rules or laws govern it. This means that products made through precision fermentation are regulated based on their type and how they are used, using existing rules in different areas.

In the United States, some ingredients made through precision fermentation have been reviewed under the GRAS (Generally Recognized as Safe) system. Chymosin, produced using a genetically engineered microorganism, was the first genetically engineered food product to receive GRAS status in 1990. In 2020, the FDA approved β-lactoglobulin (a type of whey protein) made by fermenting Trichoderma reesei, as submitted by Perfect Day, Inc.

In some regions, products made through precision fermentation may be classified as "novel foods." If this classification applies, these products may need to follow special approval processes that differ from those for traditional foods. Precision fermentation may use genetically modified microorganisms during production, but these organisms are often not present in the final food product. However, their use might still require rules related to genetically modified materials in food manufacturing. In both the United States and the European Union, food products made with genetically modified microorganisms but that do not contain any DNA do not need to be labeled as genetically modified.

Products made through precision fermentation are already available in many countries. Some have been sold for many years, such as vitamins, food colorings, and enzymes. For example, chymosin made through precision fermentation makes up more than 90% of the global market for rennet used in cheese production. More recently, new products like human-like milk oligosaccharides, soy leghemoglobin, and whey protein have been introduced in some countries.

Future perspectives in precision fermentation

Precision fermentation is expanding the possibilities of biotechnology, providing a way to make products that helps the environment in many industries. Growth in areas like alternative proteins and foods made from microbes is expected to increase quickly because more people want natural, organic, and sustainable choices. Governments and companies are also investing in research and innovation in these areas. Examples include the UKRI Engineering Biology Mission Hub for Microbial Food in the United Kingdom, the Centre for Precision Fermentation and Sustainability (PreFerS) in Singapore and the United States, and the three Bezos Centres for Sustainable Protein at Imperial College London, NC State University, and National University of Singapore. These developments have been helped by AI technology that assists in designing microbes, improving processes, and making production more efficient.