Fermentation Process – Types, Microorganisms, and Applications
- 3. Fermentation ▪ Fermentation is derived from Latin word feivere which means to boil. ▪ Fermentation is to break down of larger molecules into smaller and simple molecules using microorganisms. ▪ In Biotechnology, Fermentation means any process by which microorganisms are grown in large quantities to produce any type of useful materials. ▪ Process of growing a culture of organisms in a nutrient media and thereby converting feed into its desired end product. ▪ This process is carried out in an equipment called as fermenter. ▪ Fermenter is a vessel in which sterile nutrient media and pure culture of microorganism are mixed and fermentation process is carried out under aseptic and optimum condition.
- 5. Basic Requirement Fermentation Equipment • Bioreactors, fermenters, or other vessels. • Provision for mixing/agitation. • Control systems for temperature, pH, aeration, and pressure. • Ports for sampling and monitoring. Monitoring and Control Systems • Substrate concentration. • Product concentration. • Growth of microorganisms. • Environmental conditions (e.g., pH, temperature, dissolved oxygen). Post-Fermentation Processing • Product recovery and purification (e.g., distillation, filtration). • Waste management (e.g., treatment of residual biomass or effluents). Fermentation Medium • Carbon Source: Sugars (e.g., glucose, sucrose). • Nitrogen Source: Ammonium salts, urea, peptone. • Minerals: Magnesium, potassium, calcium, and trace elements. • Vitamins: Required for microbial growth in some cases. • Buffers: Maintain pH stability during fermentation.
- 6. Sterile Environment • Prevents contamination by unwanted microorganisms. • Sterilization of medium, equipment, and air supply. • Maintenance of aseptic conditions during fermentation. Controlled Environment • Temperature: Optimal range for microbial activity (e.g., 30-37°C for many microorganisms). • pH: Maintained within the preferred range for enzyme activity and stability. • Aeration: Ensures oxygen availability in aerobic fermentation (e.g., via sparging or agitation). • Agitation: Uniform distribution of nutrients and temperature. Substrate • A nutrient source or feedstock for microorganisms, such as glucose, sucrose, or starch. • Provides energy and carbon/nitrogen sources for microbial growth and product formation. Microorganism The biological agent (e.g., yeast, bacteria, fungi) responsible for fermentation.
- 8. Microorganisms • Microorganisms that carry out their metabolism using oxygen are referred to as aerobic microorganisms. • Some microorganisms can substitute nitrate, others sulphate or ferric ion, for oxygen and thus grow in the absence of oxygen. These microorganisms are referred as anaerobic microorganisms. • Microorganisms can be classified according to the lowest temperatures at which significant growth occurs. Microorganism Types Growth Temperatures (℃) Psychrophiles <20 Mesophiles 20-45 Thermophiles 45-60
- 9. Microorganisms used in Fermentation 1.Bacteria Examples and Applications: •Lactobacillus → Lactic acid production (yogurt, pickles). •Clostridium → Acetone, butanol, and ethanol production. •Acetobacter → Acetic acid (vinegar production). •Methanogens → Methane in biogas production. 2. Yeasts Examples and Applications: •Saccharomyces cerevisiae → Ethanol (alcoholic beverages, biofuels), bread making. •Candida → Industrial ethanol production and single-cell protein. •Pichia pastoris → Recombinant protein production.
- 10. 3. Fungi (Molds) Examples and Applications: •Aspergillus niger → Citric acid, enzymes (amylase, cellulase). •Penicillium → Penicillin (antibiotic production). •Rhizopus → Lactic acid, fumaric acid, and enzymes. 4. Algae Examples and Applications: •Chlorella and Spirulina → Single-cell protein, dietary supplements. •Dunaliella salina → Beta-carotene production.
- 11. Selection Criteria for Microorganisms •Product specificity: Ability to produce desired metabolites (e.g., alcohol, acid). •Growth conditions: Suitability for specific pH, temperature, and oxygen levels. •Substrate utilization: Ability to metabolize available feedstocks. •Yield and efficiency: High productivity with minimal by-products. •Scalability: Suitability for industrial-scale processes.
- 13. Type of Fermentation Process Types of Fermentation process Solid State Fermentation Submerged Fermentation Anaerobic Fermentation Aerobic Fermentation
- 14. 1. Solid State Fermentation: Solid-state fermentation (SSF) is a fermentation process in which microorganisms grow and metabolize nutrients on solid materials in the near absence of free water. However, the materials retain enough moisture to support microbial activity. Key Characteristics: 1. Moisture Content: 1. Low compared to submerged fermentation. 2. The substrate acts as both a nutrient source and a physical support. 2. Microorganisms Used: 1. Fungi (e.g., Aspergillus, Penicillium) are most commonly used because they can thrive in low-moisture environments. 2. Some bacteria and yeasts can also be employed. 3. Substrates: 1. Agricultural residues like wheat bran, rice husk, corn cobs, and sugarcane bagasse. 2. Waste materials from industries (e.g., fruit peels, brewery waste).
- 15. Contd. Applications: •Enzyme Production: Amylase, cellulase, protease. •Secondary Metabolites: Antibiotics, pigments. •Food and Feed: Fermented foods (e.g., soy sauce, miso), animal feed enrichment. •Bioenergy: Production of bioethanol. Advantages: 1.Cost-effective as it uses inexpensive substrates. 2.Low water and energy requirements. 3.Minimal wastewater generation. 4.Simpler downstream processing.
- 16. Submerged fermentation Submerged fermentation is a process in which microorganisms grow and metabolize nutrients in a liquid medium. The nutrients are dissolved or suspended in water, providing an environment conducive to microbial growth and product synthesis. Key Characteristics: 1. Medium: 1. Liquid with dissolved nutrients. 2. Requires high water activity for microbial activity. 2. Microorganisms Used: 1. A wide range of microorganisms, including bacteria, fungi, and yeast. 3. Process Conditions: 1. Controlled parameters like temperature, pH, aeration, and agitation for optimal growth. 4. Equipment: 1. Typically performed in bioreactors (fermenters) equipped with systems for mixing, oxygen transfer, and heat exchange.
- 17. Contd. Applications: •Enzyme Production: Lipase, cellulase, and protease. •Pharmaceuticals: Antibiotics (e.g., penicillin), vaccines, and bioactive compounds. •Industrial Chemicals: Citric acid, lactic acid, ethanol. •Food and Beverages: Alcoholic beverages, amino acids, and flavor compounds. Advantages: 1.Easier control of environmental factors like aeration and pH. 2.Higher productivity due to efficient microbial growth. 3.Suitable for large-scale production in industrial applications. 4.Supports a wide variety of microorganisms and products.
- 18. Anaerobic Fermentation: Anaerobic fermentation is a metabolic process where microorganisms produce energy and desired products in the absence of oxygen. It relies on glycolysis and other pathways to break down organic compounds, producing by-products like alcohol, organic acids, and gases. Key Characteristics: 1. Oxygen Absence: 1. Strictly conducted in environments devoid of oxygen. 2. Microorganisms use other molecules (e.g., nitrate, sulfate) as electron acceptors. 2. Microorganisms Used: 1. Anaerobic bacteria (e.g., Clostridium, Methanogens) and some yeasts. 3. Products: 1. Alcohols (e.g., ethanol, butanol), organic acids (e.g., lactic acid), and gases (e.g., methane, carbon dioxide).
- 19. Contd. Applications: •Alcohol Production: Ethanol for beverages and biofuels. •Biogas Production: Methane from organic waste in anaerobic digesters. •Food Industry: Fermentation of dairy products (e.g., yogurt, cheese) and pickles. •Waste Management: Treatment of wastewater and organic waste through anaerobic Advantages: 1.Cost-effective setup for small-scale and specific applications. 2.Environmentally friendly by-products (e.g., methane for energy). 3.Requires less complex equipment compared to aerobic processes. 4.Suitable for waste valorization and renewable energy generation.
- 20. Aerobic Fermentation: Aerobic fermentation is a metabolic process where microorganisms grow and produce desired products in the presence of oxygen. It involves oxidative metabolism, which generates energy and specific metabolites using oxygen as the final electron acceptor. Key Characteristics: 1. Oxygen Requirement: 1. Requires continuous oxygen supply to sustain microbial activity. 2. Performed in aerated bioreactors with agitation to ensure oxygen transfer. 2. Microorganisms Used: 1. Primarily bacteria, yeast, and fungi that thrive in oxygen-rich environments. 3. Process Conditions: 1. Strict control of parameters like oxygen concentration, temperature, pH, and agitation for optimal product yields.
- 21. Contd. Applications: •Pharmaceuticals: Production of antibiotics (e.g., penicillin, streptomycin) and vitamins. •Food Industry: Citric acid, amino acids, and fermented food products. •Bioremediation: Decomposition of pollutants using aerobic microorganisms. •Biotechnology: Biomass production (e.g., single- cell proteins). Advantages: 1.High energy yield from oxidative metabolism. 2.Efficient production of primary and secondary metabolites. 3.Suitable for a wide variety of microorganisms and high-value products. 4.Well-suited for industrial-scale operations with advanced bioreactors. maintained.
- 22. Comparison between types of Fermentation Feature Solid-State Submerged Anaerobic Aerobic Medium Solid Liquid No oxygen Oxygen Product Types Enzymes, flavors Enzymes, drugs Alcohol Antibiotics Energy Requirement Low High Low High By-Products Minimal Variable Gases Organic acids
- 24. Fermenter (Bioreactor) A fermenter is a vessel used for the cultivation of microorganisms or cells under controlled conditions to produce desired products like enzymes, antibiotics, alcohol, or biofuels. It ensures optimal conditions for microbial growth and product formation through precise control of parameters like temperature, pH, aeration, and agitation. Structure of a Typical Fermenter A standard fermenter can be visualized as: 1.Cylindrical Vessel: Vertical orientation with a conical or flat bottom for efficient mixing and drainage. 2.Top Lid: Contains ports for air inlet, nutrient addition, and sensors. 3.Internal Components: Includes the agitator, sparger, and baffles for optimal mixing and aeration. 4.External Components: Includes temperature control systems, pumps, and a control unit.
- 25. Ensures uniform mixing of the medium, enhances mass transfer, and prevents cell clumping Introduces sterile air or oxygen into the medium, crucial for aerobic fermentation Monitors and adjusts pH by adding acid or alkali. Vertical strips inside the vessel to prevent vortex formatio during agitation and enhance mixing efficiency Body: Contains the culture medium and microorganisms. Provides a sterile environment Central unit to monitor and regulate parameters like temperature, pH, oxygen levels, and agitation speed. Temperature sensor: Maintains optimal temperature for microbial growth. Removes gases produced during fermentation (e.g., CO₂) while preventing contamination. Supplies oxygen for aerobic processes and maintains sterility of incoming air.
- 26. 1.Body/Vessel: 1. Material: Stainless steel (industrial), glass (lab-scale). 2. Function: Contains the culture medium and microorganisms. Provides a sterile environment. 2.Agitator/Stirrer: 1. Types: Impellers (e.g., Rushton turbine, marine propeller). 2. Function: Ensures uniform mixing of the medium, enhances mass transfer, and prevents cell clumping. 3.Sparger: 1. Function: Introduces sterile air or oxygen into the medium, crucial for aerobic fermentation. 2. Types: Porous sparger, nozzle sparger. 4.Baffles: 1. Function: Vertical strips inside the vessel to prevent vortex formation during agitation and enhance mixing efficiency. 5.Temperature Control System: 1. Components: Heating/cooling jackets, coils, or plates. 2. Function: Maintains optimal temperature for microbial growth. 6.pH Sensor and Controller: 1. Function: Monitors and adjusts pH by adding acid or alkali. 7.Aeration System: 1. Components: Air pump, sparger, and filters. 2. Function: Supplies oxygen for aerobic processes and maintains sterility of incoming air. 8.Sampling Port: 1. Function: Allows aseptic withdrawal of samples to monitor growth and product formation. 9.Harvest Port: 1. Function: Facilitates the collection of the final product at the end of the fermentation process. 10.Nutrient Feed System: 1. Function: Introduces additional nutrients into the fermenter as required during the fermentation process. 11.Exhaust System: 1. Components: Exhaust outlet with filters. 2. Function: Removes gases produced during fermentation (e.g., CO₂) while preventing contamination. 12.Control Panel: 1. Function: Central unit to monitor and regulate parameters like temperature, pH, oxygen levels, and agitation speed. Parts of a Fermenter and Their Functions
- 27. Working of Fermenter • The vessel and medium are sterilized to maintain a contamination-free environment. Preparation and Sterilization: • Introduction of microorganisms into the sterilized medium under aseptic conditions. Inoculation: • Continuous monitoring and regulation of temperature, pH, aeration, and agitation to optimize microbial growth. Control of Parameters: • For aerobic processes, air is supplied and distributed via the sparger and agitator, ensuring even oxygen availability. Mixing also ensures uniform nutrient distribution. Aeration and Mixing: • Microorganisms grow and metabolize the substrate, producing the desired product (e.g., alcohol, enzymes). Product Formation: • Samples are periodically taken to track microbial growth and product concentration. Sampling and Monitoring: • The product is extracted, and the remaining medium and biomass are disposed of or reused. Harvesting:
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