Microbiology_6_Sterilization

MPharm Purification & Sterilization

• The presentation covers purification and sterilization within the MPharm program.
• The lecture notes provide a useful reference for purification and sterilization in the MPharm program, as well as Micro courses.
• Learning objectives include introduction to downstream processing, purification, sterilization, different sterilization processes, sterility checking, and sterility testing.
• A diagram outlines the mAb upstream process, from cell bank storage to production/seed bioreactor and primary recovery.
• Purification separates products from production mixtures by removing unwanted components, using methods like sedimentation, precipitation, centrifugation, adsorption, and microfiltration (specified MWCO).
• Purification reduces risks of side effects while maintaining yield, which depends on the number of steps and product loss at each.
• Different yield percentages are shown at various stages of product recovery.
• Downstream processing involves techniques like sedimentation and precipitation, which rely on cell size, density, and mixing speed.
• Sedimentation and precipitation methods lower the solubility of the solute in the media and causes the product to fall out of solution.
• This is useful in production of recombinant DNA polymerases via chemical, temperature, or pH changes.
• Centrifugation uses centrifugal force to separate products, with denser particles moving outwards.
• Adsorption chromatography is used for protein separation. Ion exchange binds proteins to the charge, while immuno-affinity uses antigenic regions to bind unwanted components.
• Sterilization removes all microorganisms, including bacteria, fungi, and viruses (including those in biologically derived therapeutics such as monoclonal antibodies and plasma components). It can also include disabling/destruction/removal of infectious proteins (e.g., Prions, or Transmissible Spongiform Encephalopathies).
• Microbial resistance to various sterilization methods is evaluated; prions show exceptional resistance to most sterilization agents.
• Sterilization methods include steam (autoclave), gas sterilization, filtration, dry heat (oven), and ionizing radiation.
• Filtration removes microorganisms rather than destroying them by size using varying filter grades, pore size, and membrane composition.
• Heat-based sterilization methods use either moist heat (hydrolytic action at >120°C and >1 atm pressure) or dry heat (oxidative action at >150°C) for a wide spectrum of antimicrobial use in inactivating biohazard waste.
• Gas-based sterilization uses ethylene oxide or formaldehyde, often for more temperature-sensitive items like reusable surgical instruments or medical devices.
• Sterilization using radiation includes methods such as ionizing or non-ionizing radiation. Ionizing forms like Gamma or X-rays are common in industrial sterilizing while non-ionizing ultraviolet light has uses in surface disinfection and other forms of sterilization.
• Sterilization success is verified using a combination of physical, chemical, and biological indicators. Physical indicators provide temperature & pressure records. Chemical indicators, such as autoclave tape, visually react to process conditions, and biological indicators demonstrate sterilization success by growing bacteria after the sterilization process if living microorganisms remain.
• Sterility testing verifies that the sterilized product is free from contamination by incubating it in a nutrient medium. Membrane filtration is a common technique.

Sterility Assurance

• Sterility testing involves assessing if a sterilized product is free from microbial contamination by incubation in an appropriate sterile nutrient medium.
• Sterility assurance relies on the understanding that microbial populations decrease exponentially with exposure to the sterilization process, regardless of the initial contamination level.
• Achieving true sterility is theoretically possible but practically takes an infinite time to ensure zero survivors. Testing can minimize microbial contamination but doesn't guarantee 100% sterility.
• The presence of different microbial contaminants will affect the sterilization efficiency.

Additional Information

• Useful references for the study material include the textbook Principles and Practice of Disinfection, Preservation and Sterilization by Russell, Hugo & Ayliffe and the other reading materials from the pharmaceutical microbiology text.

This quiz on MPharm Purification and Sterilization covers essential concepts in the field of pharmaceutical sciences. It includes downstream processing, different sterilization methods, and various techniques involved in purification and sterility testing. Assess your understanding of these critical processes in the drug manufacturing industry.