Fermentation

Strain selection is the first step for both biomass and precision fermentation processes. In theory, microbial fermentation encompasses an enormous variety of species with vast biological diversity, ranging from fungi to bacteria to microalgae. However, very few microbial species have so far been commercialised for food use.

As biomass fermentation focuses on cultivating microbial biomass as a protein source, the selected strain must grow rapidly and produce high protein levels and essential nutrients. Common microbes used in biomass fermentation include fibrous fungi like Fusarium venenatum, other mycoprotein sources, and algae like Chlorella and Spirulina species. These microbes can vary in nutritional profile, growth and yield efficiency, final texture, and function. 

Precision fermentation harnesses microorganisms (e.g., yeast, bacteria, fungi) to produce specific ingredients that can be used in various food and agricultural products. The ingredients produced can include egg and dairy proteins as well as specific enzymes, flavours, colours, fats, and oil. Strains are therefore selected for their ability to produce the target molecule efficiently and at scale. In the target selection process, the molecule or molecules of interest are called the target. The target can be a protein, a lipid, a flavour compound, a fragrance, an enzyme, a growth factor, a pigment, or another class of molecule. The chosen microbe, also referred to as the host organism, must efficiently grow these molecules in the upstream process and then be separated and purified from the target molecule in the downstream process. Scientists must carefully choose the ideal microbe for the entire precision fermentation process. Common microbes include fungi like Trichoderma reesei and Komagataella phaffii and bacteria like Escherichia coli.

Strains can be sourced from various culture collections and commercial suppliers. Comprehensive strain discovery programmes rely on massive data sets and specimen libraries, or biofoundries, to ensure broad capture of sufficient biological diversity, genomic data, and distinct growth conditions. 

In both types of fermentation, the strain selection process is crucial for optimising yield, scalability, and the final product’s functionality. The chosen strains are often subjected to genetic modifications to enhance their performance and ensure that they meet the specific demands and safety requirements of alternative protein production.

Strain engineering involves genetically modifying microbial strains to enhance their protein production capabilities, which is crucial for both biomass and precision fermentation. Advanced biotechnological tools like CRISPR-Cas9, homologous recombination, and transposon mutagenesis are employed to introduce specific genetic changes.

CRISPR-Cas9 allows for precise genetic modifications by creating targeted breaks in DNA. The process begins with identifying the gene or genetic sequence that influences traits like protein yield or metabolic efficiency. Guide RNA (gRNA) is designed to match the sequence adjacent to the gene of interest, directing the Cas9 enzyme to the exact location for editing. Once the Cas9 makes a cut, the cell’s natural DNA repair mechanisms are leveraged to introduce new genetic material, delete segments, or alter the existing sequence.

Identifying target genes involves analysing metabolic pathways using bioinformatics tools to determine which genes to knock out, insert, or modify for enhanced protein production. Scientists might target various features, such as:

1. Hydrophobicity: Modify surface proteins to improve hydrophobicity, facilitating easier separation of the product from the fermentation broth.
2. Growth rate: Enhance genes regulating cell division to increase growth rates, thus boosting overall yield.
3. Metabolic efficiency: Optimise pathways for more efficient use of carbon and nitrogen sources, thus reducing waste and increasing productivity.
4. Stress tolerance: Introduce genes that confer resistance to environmental stresses like pH fluctuations, temperature changes, or osmotic pressure, to ensure consistent production under industrial conditions.
5. Secretion efficiency: Engineer the secretion pathways to ensure that target proteins are efficiently secreted into the medium, simplifying downstream processing.
6. Product stability: Modify enzymes or proteins to enhance their stability during production and storage, which reduces degradation and maintains quality.
7. Cellular longevity: Extend the productive life cycle of the microbial cells to reduce the frequency of culture replacement, which improves production process efficiency.

After engineering, strains undergo extensive testing to ensure that the modifications enhance performance without compromising growth or stability. The goal is to create microbial strains that produce high yields of target proteins under industrial fermentation conditions, making them more efficient and reliable for alternative protein production. Successful strain engineering bridges the gap between basic research and practical application, which ensures that the strains meet the specific demands of industrial-scale production.

Strain optimisation is a critical process that fine-tunes genetically engineered strains to maximise efficiency and stability in protein production. This stage utilises techniques like Adaptive Laboratory Evolution (ALE) and codon optimisation to ensure that strains function at their most efficient state.

In ALE, strains are exposed to selective pressures—such as temperature, pH, nutrient availability, and oxygen levels—over multiple generations to identify and cultivate those with optimal performance. High-throughput screening and automated systems evaluate environmental conditions and genetic modifications simultaneously.

Codon optimisation is particularly crucial in precision fermentation. It involves modifying the DNA sequence of the target gene to use codons preferred by the host organism, increasing protein expression, efficient protein folding, and post-translational modification without altering the protein’s amino acid sequence. Tailoring gene sequences to the microbial host’s preferred codons significantly enhances protein production efficiency.

Omics technologies—such as genomics, transcriptomics, proteomics, and metabolomics—help understand the strains’ metabolic pathways and regulatory networks, offering comprehensive insights for further refinement.

After optimisation, strains are validated in pilot-scale bioreactors to ensure that they maintain their enhanced traits under conditions mimicking large-scale production. This validation confirms the optimised strains’ ability to deliver consistent, high-yield production in commercially relevant conditions.

Cell banking is a crucial process in fermentation, as it ensures the availability of consistent, high-quality microbial strains for production. This involves creating a Master Cell Bank (MCB) and Working Cell Banks (WCBs) from a well-characterised and optimised microbial strain. The MCB serves as the original source, stored under conditions that preserve genetic and phenotypic stability, while WCBs are derived from the MCB and used for routine production, thus maintaining consistency across batches.

The creation of cell banks begins with the extensive characterisation of the optimised strain, which ensures genetic stability, growth consistency, and freedom from contamination. The selected strain is expanded under controlled conditions, aliquoted, and cryopreserved using cryoprotectants such as glycerol or dimethylsulfoxide at ultra-low temperatures (e.g., -80°C or in liquid nitrogen at -196°C). This process ensures the long-term viability and retention of the desired traits for future use.

Role of Biofoundries: Biofoundries, which are comprehensive data and specimen libraries, play a vital role in modern cell banking. They enable high-throughput, automated processes for strain development and storage and ensure precision and scalability. Biofoundries are also essential for systematic strain discovery, which allows for the broad capture of biological diversity, genomic data, and growth conditions. 

Challenges and Opportunities: Developing and maintaining comprehensive strain libraries and biofoundries is expensive and time-consuming, often restricting access to a few well-funded entities. These limitations can delay the commercial adoption of novel strains and hinder broader innovation. Coordinated efforts in biofoundries can accelerate the discovery and optimisation of strains suitable for alternative protein production by providing extensive data for strain improvement and adaptation, facilitating faster regulatory approval and commercial adoption of new strains. Such efforts can drive significant improvements in nutritional quality, production efficiency, strain robustness, and end-product traits.

To overcome these challenges, the industry must focus on both technological advancements and the adoption of globally recognised standards for feedstock and strain quality. Comprehensive efforts should also integrate systems biology insights to design novel hosts with ideal attributes for fermentation, such as prolonged stability, efficient metabolism, and desirable flavour profiles.

Feedstock and media optimisation are critical in developing nutrient formulations that support the efficient growth and productivity of selected microbial strains in biomass and precision fermentation. The media typically consist of carbon and nitrogen sources, vitamins, minerals, and growth factors essential for microbial metabolism and protein synthesis. Depending on the microbial strain, the media’s composition is adjusted to match specific nutritional needs and optimise the yield of the target product.

In biomass fermentation, the feedstock must support economically efficient microbial growth, with a focus on high protein content. Refined sugar-based substrates from corn or sugarcane are commonly used due to their availability and well-characterised nature. The fermentation media often includes a blend of carbon sources such as glucose, sucrose, dextrose, and lactose for energy, along with nitrogen sources like ammonium salts, urea, amino acids, and yeast extract for protein synthesis. Essential minerals such as magnesium, calcium, phosphates, iron, and zinc play crucial roles in enzymatic functions and cellular processes. Additionally, vitamins and growth factors, including B vitamins, biotin, folic acid, and trace elements like selenium and copper, are necessary for optimal microbial metabolism and protein yield. As sustainability and cost-effectiveness gain importance, alternative feedstocks—such as agricultural byproducts, waste streams, and gaseous substrates—are being explored. For example, fungi used in biomass fermentation can be grown on lignocellulosic feedstocks, which are abundant and economical, though more challenging to process.

In precision fermentation, where microbes are engineered to produce specific molecules like proteins or enzymes, the media is tailored to enhance the expression and stability of the target molecule. Carbon sources like glucose, methanol, and glycerol, along with precisely formulated nitrogen sources such as ammonium salts and amino acids, are used to support the expression of target proteins. Minerals are carefully balanced, with iron, magnesium, and trace elements being critical for enzyme co-factors and metabolic pathways. Growth factors and vitamins are customised, often including specialised cofactors essential for biosynthesis. The media is optimised for the specific microbial host to maximise yield, efficiency, and product purity. For instance, in producing dairy proteins using yeast, the media might be optimised to increase the secretion of casein or whey protein while maintaining yeast cell health and productivity.

The optimisation process involves empirical testing and computational modelling. Researchers test various feedstocks and media components to identify those that maximise yield and minimise production costs. Computational tools model microbial metabolic pathways, predicting how different media compositions will affect growth and production.

The ultimate goal is to create a scalable, cost-effective, and sustainable fermentation process, ensuring that microbial strains can be grown at an industrial scale without compromising efficiency or product quality.

Media preparation is a crucial step in the upstream process, involving the formulation and sterilisation of growth media used in fermentation. This step ensures that the media is free from contaminants and contains all the necessary nutrients for optimal microbial growth. Large-scale preparation equipment, such as media tanks and autoclaves, are used to mix and sterilise the media. The media composition is carefully optimised based on the results from the feedstock and media optimisation step. Proper pH adjustment and nutrient balance are critical to support the high-density cultures required for protein production. The prepared media is then transferred to sterile storage tanks or directly into the bioreactors for fermentation. This step ensures that the microbial cultures have a consistent and nutrient-rich environment that promotes efficient protein production.

Media recycling is increasingly explored as a strategy to reduce costs and waste, particularly in large-scale fermentation processes. However, recycling presents challenges, such as maintaining nutrient balance and preventing the accumulation of inhibitory byproducts that could hinder microbial growth. Effective recycling requires careful monitoring and adjustment of the media composition to ensure that it continues to support optimal microbial activity.

Supply chain optimisation is another critical aspect, as the availability and cost of raw materials can significantly impact the overall economics of the fermentation process. Sourcing sustainable and affordable feedstocks, such as agricultural byproducts or waste streams, is a priority to enhance the sustainability of fermentation operations. This approach not only reduces reliance on refined sugars but also promotes a circular economy by utilising waste materials as valuable inputs. Ensuring a stable and cost-effective supply of these materials is essential for scaling up fermentation processes and achieving long-term viability.

Cleaning and sterilisation are vital processes in maintaining the safety and efficiency of fermentation operations. Proper sterilisation ensures that all microbial life, including potential contaminants, is eradicated from equipment and surfaces before the initiation of a new fermentation batch. Autoclaving is a common sterilisation method that uses high-pressure steam to achieve this, ensuring that bioreactors, tubing, and other equipment are free from any biological contaminants that could compromise the fermentation process.

In the context of fermentation involving living modified organisms (LMOs), strict regulatory compliance is required to prevent biohazards. The Cartagena Protocol on Biosafety provides international guidelines for the safe handling, transport, and use of LMOs, which ensures that organisms do not pose risks to human health or the environment. Compliance with these regulations is crucial for companies involved in the development and production of alternative proteins, as it ensures that their operations are safe and legally compliant.

Safety protocols are paramount in cleaning and sterilisation efforts, not only to protect the integrity of fermentation processes, but also to safeguard workers and the environment. Proper handling of biohazardous materials, including the use of personal protective equipment, biosafety cabinets, and containment strategies, is essential to prevent accidental exposure or release of harmful organisms. Regular validation of sterilisation procedures, such as through biological indicators and sterility testing, is also necessary to confirm that sterilisation processes are effective and meet industry standards.

In addition to autoclaving, other sterilisation methods such as steam-in-place (SIP) systems, chemical sterilants, and UV irradiation may be employed depending on the specific requirements of the process and the properties of the materials being sterilised. Maintaining rigorous standards for cleaning and sterilisation not only supports regulatory compliance but also ensures the smooth operation of fermentation processes, minimising the risk of contamination and production downtime.

Monitoring and control are critical components in ensuring the success of fermentation processes. This involves the continuous observation and regulation of key parameters such as temperature, pH, dissolved oxygen, and nutrient levels within the bioreactor. These parameters must be maintained within strict ranges to optimise microbial growth and product formation, as deviations can lead to reduced yields or compromised product quality.

Advanced sensors and monitoring systems are integrated into the bioreactor setup to provide real-time data on these critical parameters. For instance, temperature sensors ensure that the fermentation environment remains within the optimal range for the specific microbial strain, while pH sensors monitor the acidity or alkalinity of the broth, which can significantly impact enzyme activity and metabolic processes. Dissolved oxygen sensors are particularly important in aerobic fermentation processes, as they measure the availability of oxygen, which is essential for the respiration and growth of aerobic microbes.

Nutrient levels are also closely monitored to ensure that the microbial culture is adequately supplied with carbon, nitrogen, and other essential nutrients throughout the fermentation process. Automated nutrient dosing systems can be employed to adjust the feed rates based on real-time data, thereby ensuring that the microbes have a consistent supply of nutrients without the risk of overfeeding, which could lead to byproduct formation or inhibition of microbial growth.

Effective monitoring and control systems not only maximise the efficiency and yield of the fermentation process but also contribute to the overall quality and consistency of the final product. By collecting and analysing data throughout the fermentation process, operators can make informed decisions and adjustments to optimise the process. This continuous feedback loop is essential for maintaining the stability of the fermentation environment, reducing variability, and ensuring that the product meets the desired specifications.

Seed train development is a critical step in scaling up microbial cultures for fermentation, beginning with a small volume of starter culture and progressively expanding it to larger vessels. The process starts with inoculating small flasks, which are then scaled up through a series of small-scale bioreactors. The seed train process has many variations and can involve progressively scaling from 1, 3, 5, 10 to 20 litres to 1, 10, 100, 1000, 10k, and 100k litres. The purpose of the seed train is to increase both the density and the volume of the microbial culture, which ensures that the microbes are actively replicating and producing the target molecule, whether that be a protein, enzyme, or another desired compound.

As the seed train progresses, the microbial cells adapt to increasing volumes and more complex environmental conditions. Throughout this process, cells undergo rapid growth and proliferation, particularly during the log phase, where the population expands exponentially. This expansion is supported by carefully controlled conditions, including nutrient-rich fermentation broth, which serves as the medium in which the microbes grow. The fermentation broth will be a complex mixture of microbes, water, carbon sources, nitrogen sources, vitamins, minerals, and growth factors that provide the necessary nutrients for microbial metabolism and biosynthesis.

By the end of the seed train process, the culture is prepared for transfer into the primary bioreactor, where large-scale production will occur. This step is vital for ensuring that the culture is robust and dense enough to handle the transition to large-scale bioreactors without losing its productivity or stability.

Primary USP bioreactor cultivation is where microbial cultures are scaled up to industrial volumes, often ranging from hundreds to thousands of litres, to maximise the production of proteins, enzymes, or biomass. This stage is critical in transitioning from seed train development to full-scale fermentation, where the focus is on maintaining optimal conditions such as temperature, pH, dissolved oxygen, and nutrient supply to ensure efficient microbial growth and target molecule production. The bioreactor setup, microbe cultivation process, and media formulation will differ depending on the fermentation technique used. Types of fermentation include: 

• Submerged Fermentation (SmF): SmF involves microbes growing in a liquid nutrient medium, which is ideal for producing soluble products like enzymes or proteins. The microbes, such as Komagataella phaffii or Escherichia coli, replicate and produce the target molecules within the fermentation broth. SmF is favoured for its scalability and ease of controlling environmental factors like oxygen and nutrient distribution. However, it requires significant energy input for aeration and cooling, making it costlier. It is the common technique used in the precision fermentation process. Companies like Perfect Day use SmF for producing animal-free dairy proteins.
• Solid-State Fermentation (SSF): SSF involves cultivating microbes on solid substrates with minimal free water. This method is particularly effective in biomass fermentation, producing fungal biomass or enzymes. The microbes, including strains like Aspergillus niger or Rhizopus oryzae, grow on substrates such as agricultural byproducts. SSF is advantageous for its lower water and energy use, but it is challenging to scale due to difficulties in maintaining uniform conditions across the substrate. SSF is utilised by companies like MycoTechnology for producing mycoprotein.
• Liquid-Air Interface Fermentation (LAIF): LAIF is a hybrid method where microbes grow at the interface between a liquid medium and air. This method combines aspects of both SmF and SSF and is used for specialised applications. For example, Nature’s Fynd uses LAIF to produce mycelial mats. While LAIF offers efficient oxygen transfer and the benefits of both liquid and solid-state methods, it is complex to manage and scale.
  
  

Aerobic vs. Anaerobic Fermentation: The choice between aerobic and anaerobic fermentation significantly impacts the process. Aerobic fermentation, which requires oxygen, generally yields higher biomass or target molecules and produces byproducts like carbon dioxide. However, it is energy-intensive due to the need for continuous oxygen supply and cooling systems to manage the heat generated. Anaerobic fermentation occurs without oxygen and produces ethanol, lactic acid, or other metabolites as byproducts. This method is typically slower and yields less biomass but is more energy-efficient and often used in specific precision fermentation applications. Each method’s choice depends on the desired product and the microbial strain used, with aerobic processes commonly applied in high-value products like proteins and anaerobic processes in applications like bioethanol production.

Temperature Control and Cooling: Maintaining the correct temperature is vital for microbial growth and product formation. Overheating generated during fermentation can negatively impact yield and process efficiency. Cooling systems are integrated into bioreactors to dissipate heat and maintain the desired temperature. This is crucial as even slight deviations from the optimal temperature can slow microbial activity or lead to the degradation of target molecules. Different fermentation setups may require unique cooling solutions depending on the type of fermentation and the specific process conditions.

Fermentation Times and Yields: The duration of upstream fermentation varies based on the process and organism used. SmF processes typically last from 24 to 72 hours, while SSF and LAIF can extend over several days or weeks. Yield, often measured as grams of product per litre of culture (g/L), is a critical metric for process efficiency. Higher yields reduce production costs and improve scalability, making yield optimisation a key focus in bioprocess development.

Harvesting is the first step in downstream processing (DSP). DSP involves purifying and refining the target product from the fermentation broth, which may include proteins, enzymes, or biomass, depending on the type of fermentation used. Harvesting specifically refers to the initial step where the microbial culture is collected from the bioreactor, setting the stage for subsequent purification and processing. DSP is essential for transforming the raw fermentation broth with the target molecule into a final, market-ready product with high purity and quality.

Harvesting Based on Fermentation Type:

1. Submerged Fermentation (SmF): In SmF, where microbes are cultivated in a liquid medium, harvesting typically involves separating the microbial cells from the liquid broth. Centrifugation or microfiltration is often used to achieve this, depending on the size and density of the cells. After harvesting, the liquid medium containing the target product can be further processed for extraction and purification. In precision fermentation, the protein is secreted into the medium or broth, which is then processed to isolate the product. The first step in this process involves removing the biomass to clarify the medium.
2. Solid-State Fermentation (SSF): SSF involves growing microbes on solid substrates with minimal free water. Harvesting in SSF is more complex, requiring the physical removal of microbial biomass from the solid substrate. This may involve mechanical separation techniques like sieving or scraping. After harvesting, the solid biomass is processed to extract the target molecules, which may involve additional washing and filtration steps.
3. Liquid-Air Interface Fermentation (LAIF): In LAIF, microbes grow at the interface between a liquid medium and air, often forming biofilms or surface layers. Harvesting in LAIF involves carefully removing these surface-grown cells or biofilms, which may be done manually or with specialised harvesting tools. The process must be delicate to prevent damaging the microbial cells, which are then processed similarly to those from SmF or SSF.

Efficient harvesting is crucial as it determines the yield and quality of the final product. The choice of harvesting method impacts the purity of the product and the efficiency of the subsequent downstream processes. Properly conducted harvesting minimises the loss of valuable target molecules and reduces contamination risks, ensuring that the downstream processing can proceed smoothly and efficiently.

Extraction is first phase of separation, which removes the target product—such as proteins, enzymes, or other molecules—from the harvested microbial biomass or fermentation broth, as well as cellular components, impurities, and residual media, thereby setting the foundation for further purification and refinement.

Different types of extraction techniques can be used as follows: 

• Cell Disruption: For intracellular products, microbial cells must be disrupted to release the target proteins. Common cell disruption methods include:

• Mechanical Disruption: Techniques such as high-pressure homogenisation and bead milling physically break the cell walls.
• Chemical Lysis: Uses chemical agents like detergents to dissolve cell membranes.
• Enzymatic Digestion: Employs specific enzymes (e.g., lysozyme) to break down cell walls.
• Sonication: Uses ultrasonic waves to create cavitation bubbles that implode, causing the cells to rupture and release their contents. This method is effective but can generate heat, which may denature sensitive proteins.
• Solvent Extraction: This technique is used when the target product is soluble in organic solvents. The fermentation broth or disrupted cell mixture is mixed with a solvent that selectively dissolves the target compound, which is then separated from the aqueous phase. The solvent is later evaporated to recover the purified product.
• Aqueous Two-Phase Extraction: A two-phase system is created where the target product preferentially partitions into one of the phases, typically an aqueous phase. This method is advantageous for purifying proteins and enzymes while minimising denaturation or loss of activity.
• Affinity Extraction: Utilises specific interactions between the target molecule and a ligand attached to a solid support or resin. This technique is highly selective and is often used for extracting proteins or other biomolecules with a known affinity to a particular ligand, such as antibodies or metal ions.

Efficient extraction is crucial for maximising protein yield and purity. Proper extraction methods ensure that the maximum amount of the target product is recovered from the fermentation broth or biomass while minimising contaminants. This step also preserves the activity and stability of the product, which is essential for its functionality in end-use applications. The extraction process must balance efficiency, cost, and product quality.

Separation and purification followed by extraction, aims to isolate and purify the target product—such as proteins, enzymes, or other molecules—at high specificity and purity levels. These processes ensure that the final product meets the stringent quality requirements necessary for food applications or other end uses.

The level of purity required depends on the intended application of the product. For example, food-grade proteins need to be free from contaminants like other proteins, nucleic acids, lipids, and residual solvents. High specificity is essential to ensure that the target molecule is effectively separated from other components in the mixture. 

Different types of purification techniques can be employed including: 

1. Centrifugation: This technique uses centrifugal force to separate components based on their size and density. It is often the first step in purification to remove large particles, such as cell debris, after extraction.
2. Filtration: A physical separation process that uses a membrane or filter to separate particles based on size. Microfilitration is used to remove larger particles, bacteria, and yeast cells. Ultrafiltration is used to remove smaller molecules, such as proteins and nucleic acids. However, membrane fouling can occur, leading to reduced efficiency and increased costs.
3. Chromatography: A highly specific technique that separates molecules based on differences in their chemical properties, such as size, charge, or affinity for certain ligands. This is commonly used for purifying proteins, enzymes, and other biomolecules to high purity levels. Chromatography is often expensive due to the cost of resins and the need for precise control over the process.
   • Types:
     • Ion-Exchange Chromatography: Separates molecules based on their charge.
     • Size-Exclusion Chromatography: Separates based on molecular size.
     • Affinity Chromatography: Utilises a specific interaction between the target molecule and a ligand.
4. Precipitation: Involves adding solvents or salts to the mixture to precipitate the target molecule out of the solution. This technique is typically used for bulk protein purification where high purity is not required. It may require multiple rounds to achieve desired purity, and there is a risk of product loss during washing steps.
5. Dialysis: A separation technique where molecules are separated based on their ability to pass through a semi-permeable membrane. This technique is used to remove small contaminants or to change the buffer in protein solutions. Dialysis can be time-consuming and may not be practical for large-scale operations.

Purification steps can be costly, especially with techniques like chromatography that require specialised resins and equipment. The choice of method must balance cost with the required purity and specificity. High costs can be mitigated by optimising the process, such as by reusing chromatography resins, or by developing more cost-effective alternatives.

The efficiency of the separation and purification steps directly affects the overall yield of the process. High yields are critical for ensuring the economic viability of the production process, particularly in large-scale operations.

The final step in DSP is drying, preservation, and storage, where the purified product is converted into a stable form suitable for long-term storage and transport. This step is crucial for maintaining the quality and functionality of the target molecule or biomass, ensuring it remains viable for its intended application in food products or other uses. Different techniques include: 

1. Spray Drying: Spray drying rapidly converts liquid products into dry powder by atomising the product into a fine mist and introducing it to hot air. This method is efficient and widely used for producing stable, powdered proteins or microbial biomass. However, the high temperatures can risk denaturing sensitive proteins, affecting their functionality, which makes it less suitable for delicate or high-value products requiring maximum bioactivity.
2. Freeze Drying (Lyophilisation):  Freeze drying is a dehydration process where products are frozen and then subjected to a vacuum, causing ice to sublimate directly from solid to gas. This method preserves the structural integrity and bioactivity of proteins, making it ideal for sensitive and high-value products like enzymes. However, the process is energy-intensive, time-consuming, and costly, often reserved for applications where product quality cannot be compromised.
3. Vacuum Drying: Vacuum drying involves reducing pressure around the product to lower the drying temperature, which is particularly beneficial for heat-sensitive materials. This method allows for gentle drying that minimises thermal degradation, making it useful for preserving the quality of delicate proteins and other sensitive biomolecules. However, it is slower than other drying methods, and its scalability can be limited by the equipment’s capacity and cost.
4. Air Drying: Air drying uses warm air to evaporate moisture from the product, often employed for drying microbial biomass. This method is straightforward and cost-effective, making it suitable for large-scale applications where some loss of activity is acceptable. However, the lack of precise control over temperature and humidity can lead to inconsistent product quality, making it less ideal for high-value or sensitive products.

Once the target ingredient has been dried, the following preservation techniques can be used to ensure long-term shelf life. 

1. Use of Preservatives: Incorporating preservatives like salts, sugars, or specific chemical agents to inhibit microbial growth and prolong shelf life. This is used for food products where long shelf life is required without refrigeration. The choice of preservative must ensure that it does not alter the taste, texture, safety, or nutritional value of the product.
2. pH Adjustment: Modifying the pH of the product to create an environment that inhibits microbial growth. This is used for both  biomass and precision-fermented products, particularly in acidic or alkaline environments that microbes find inhospitable. pH adjustments must be carefully managed to avoid degrading the product.
3. Packaging in Inert Atmospheres: Removing oxygen and replacing it with nitrogen or other inert gases to prevent oxidation and spoilage. This is common for preserving the quality of dried protein powders and sensitive biomolecules. This requires specialised packaging equipment and materials, adding to production costs.

Finally, storing the product at appropriate temperature, moisture, and choosing the right packaging are key for creating the final product. 

Efficient drying and preservation processes maximise product recovery and ensure that the target proteins or biomass retain their desired functional properties.

Drying and preservation are energy-intensive processes, with costs depending on the method used. Balancing cost with the need for high-quality, stable products is a key challenge. Freeze drying, while producing high-quality results, is costly, whereas spray drying offers a more cost-effective solution but may compromise some product quality.