Untitled Quiz

Microbiological Laboratories

• Microbiological labs focus on strain improvement for increased yields, byproduct suppression, surfactant reduction, and broth modification for easier product recovery.
• Other ongoing research involves culture preservation techniques, stability testing, new procedure development, media optimization, and identification of inducers, repressors, and inhibitors.
• Lab space and equipment requirements include:
  • Glassware and equipment washing area
  • Media preparation area(s)
  • Inoculation room(s)
  • Incubator area(s)
  • Office
  • Laboratories

Analytical Support Laboratories

• Responsibilities include sterility testing, chemical assays for raw materials, materials before sterilization, optimized production batches, fermenter feeds, and waste streams.
• Common lab equipment includes auto-analyzers for product, carbohydrates, phosphate, various ions, specific enzymes, and other tools such as balances, gas chromatography (GC), high pressure liquid chromatography (HPLC), Kjeldahl equipment, titrimeters, UV-visible spectrophotometers, atomic absorption spectrophotometer (AAS), pH meters, viscometer, refractometer, densitometer, etc.

Air Filtration Systems

• Packed-bed or cartridge filters are used for sterile air and humidity reduction.
• Careful filter design extends the filter lifetime beyond 3 years.
• Fine fiber diameter in packed-bed filters increases filtration efficiency.
• Typical bed depth is 10 to 18 inches with 10 microns filter.
• Some plants utilize a separate filter for each sterile vessel, while others have a centralized filter system.

Air Compressors

• Oil-free compressed air is ideal, commonly provided by oil-free screw air compressors.
• Relative humidity should be about 85%.
• Heat exchangers are required to lower air temperature below the dew point and to reheat and control humidity.

Valves

• Gate, diaphragm, ball, and plug valves are common.
• While less of a concern for sterility, diaphragm and ball valves require maintenance.
• Diaphragm and ball valves are ideal for batch sterilizing operations.
• Plug-type valves are preferred for continuous sterilizing operations.
• Plug or diaphragm valves are common for inoculum transfer and sterile feed piping.
• All process valves and piping are 316 S/S.

Pumps

• Peristaltic pumps are a simple method for transferring inoculum between laboratory flasks, seed fermenters, and fermenters.
• Centrifugal pumps (316 S/S) are used to pump non-sterile raw materials, slurries, harvested broth, etc.

Cooling Equipment

• Cooling is required for media cooling, removal of fermentation heat, broth cooling before harvesting, and compressed air cooling.
• Cooling water is provided from cooling towers and chilled water (5º-15ºC) is produced by steam vacuum or refrigeration units.
• The fermentation department should be concerned about cooling water temperature and chloride content.
• Chloride ions above 150 ppm in water at temperatures above 80ºC can cause stress corrosion cracking of stainless steel.
• Conductivity probes are used for monitoring the cooling water line and prevent high salt levels.
• Discharge permits and monitoring may be required by pollution control agencies if cooling water is discharged to waterways.
• The chloride content in cooling water should be determined analytically every two weeks to maintain levels below 100 ppm.

Environmental Control

• Odor:
  • Sterilization of certain raw materials produces odors.
  • High air discharge from a large fermenter house makes odor unavoidable.
  • Wet scrubbing towers with sodium hypochlorite can be used for reduction but are expensive and produce waste.
  • Tall exhaust stacks dilute the released gas before reaching ground but aren't acceptable by pollution control boards.
  • Ozone treatment can be effective.
• Organic Pollution:
  • The fermentation department should monitor and control COD/BOD of liquid waste before discharge to the sewer.

Spargers

• Ring Sparger:
  • A perforated pipe positioned below the impeller in the form of crosses or rings.
  • Size should be ¾ of impeller diameter.
  • Air holes should be at least 6mm diameter.
• Orifice Sparger:
  • Used without agitation in yeast manufacture, effluent treatment, and single-cell protein production in air-lift fermenters.
• Nozzle Sparger:
  • Found in many agitated fermenters, from laboratory to production scale.
  • Consists of a single open pipe or partially closed pipe for air bubble delivery.
  • Low pressure loss and resistance to blockage.
• Combined Sparger-Agitator:
  • Developed by Herbert, Phipps, and Tempest (1965) for small-scale fermentation (1 dm3).
  • Air is introduced through a hollow agitator shaft and released through holes in the disc between the blades.
  • Provides efficient aeration in baffled vessels at varying agitator RPMs.

Aseptic Operation and Containment

• Aseptic Operation: Protection against contamination.
• Containment: Prevention of viable cell escape from fermenters or downstream equipment.
• Containment regulations were introduced in the 1970s.
• Organisms are classified into hazard groups based on potential risks.

Pressure Control

• Regulatory valves and pressure gauges are employed to maintain pressure in various components.
• Safety valves are incorporated in vessels and pipelines to release pressure upon exceeding the working pressure.

Foam Sensing and Control

• Uncontrolled foam formation can lead to oxygen deprivation.
• Antifoam agents are added when foaming reaches predetermined levels.
• Foam sensing and antifoam addition methods depend on the process and economic considerations.
• Mechanical antifoam devices include discs, propellers, brushes, hollow cones, rotating shafts, centrifugal separators, and jets spraying on deflectors.

Troubleshooting in Fermentation Plant

• Problems may stem from electrical, instrumentation, mechanical, physical, or microbial sources.
• Microbial contamination can result in:
  • Unusable inoculum.
  • Production inhibition in fermenters.
  • Contaminated products.
  • Fermentation broth that is difficult to filter or process.

Basic Sources of Contamination in Fermentation Departments

• Contamination of stock cultures due to poor techniques.
• Contamination from raw materials.
• Inadequate sterilization of equipment, air, or media.
• Insufficient procedures or inadequate operator training.
• Bacteriophages.

Six Categories for Regular Inspection to Prevent Contamination

• 1. Microbiological Laboratory: Ensure proper sterilization techniques, media preparation & control, and regular monitoring for contaminants.
• 2. Fermentation Department: Maintain proper air filtration, cleaning procedures, and environmental controls to minimize contamination risks.
• 3. Media Preparation: Strict adherence to sterilization protocols and monitoring for sterility are essential.
• 4. Sterilization: Regular maintenance of sterilization equipment, monitoring of sterilization cycles, and validating sterilization processes are critical.
• 5. Equipment: Strict cleaning and sterilization procedures for fermenters, pipes, and all processing equipment.
• 6. Personnel: Good hygiene practices, proper gowning procedures, and training in aseptic techniques are vital for avoiding contamination.