Podcast
Questions and Answers
Immuno-affinity processes are designed to target specific contaminants through binding to antigenic regions.
Immuno-affinity processes are designed to target specific contaminants through binding to antigenic regions.
True
The principle of ion exchange chromatography is solely dependent on the size of proteins.
The principle of ion exchange chromatography is solely dependent on the size of proteins.
False
Sterilization typically involves the complete removal of viruses and prions from therapeutic products.
Sterilization typically involves the complete removal of viruses and prions from therapeutic products.
False
Parenteral administration refers to administering medical products directly into the bloodstream via intravenous methods.
Parenteral administration refers to administering medical products directly into the bloodstream via intravenous methods.
Signup and view all the answers
Adsorption in downstream processing can involve the passage of a liquid phase through a liquid phase.
Adsorption in downstream processing can involve the passage of a liquid phase through a liquid phase.
Signup and view all the answers
Sedimentation speed is solely dependent on the mixing speed of the components.
Sedimentation speed is solely dependent on the mixing speed of the components.
Signup and view all the answers
Micro-filtration uses specified molecular weight cut-off (MWCO) to effectively remove all contaminants.
Micro-filtration uses specified molecular weight cut-off (MWCO) to effectively remove all contaminants.
Signup and view all the answers
Centrifugation applies centrifugal force to separate products based only on their solubility.
Centrifugation applies centrifugal force to separate products based only on their solubility.
Signup and view all the answers
Precipitation can be achieved through changes in temperature and pH.
Precipitation can be achieved through changes in temperature and pH.
Signup and view all the answers
The purification process can increase the risk of side effects while maintaining yield.
The purification process can increase the risk of side effects while maintaining yield.
Signup and view all the answers
The yield percentage in downstream processing is influenced by the number of steps used in the purification process.
The yield percentage in downstream processing is influenced by the number of steps used in the purification process.
Signup and view all the answers
Ion exchange is a type of adsorption method used in purification processes.
Ion exchange is a type of adsorption method used in purification processes.
Signup and view all the answers
Recombinant DNA polymerases are produced without any precipitation processes.
Recombinant DNA polymerases are produced without any precipitation processes.
Signup and view all the answers
UV light is effective for deep water sterilisation.
UV light is effective for deep water sterilisation.
Signup and view all the answers
Chemical indicators change visibly during heat sterilization processes.
Chemical indicators change visibly during heat sterilization processes.
Signup and view all the answers
The biological sterilization indicator used in testing is Geobacillus stearothermophilus.
The biological sterilization indicator used in testing is Geobacillus stearothermophilus.
Signup and view all the answers
Sterility assurance guarantees that there are no survivors of microorganisms after the sterilization process.
Sterility assurance guarantees that there are no survivors of microorganisms after the sterilization process.
Signup and view all the answers
Physical indicators are not used to check sterilization success.
Physical indicators are not used to check sterilization success.
Signup and view all the answers
The chance of a single surviving organism in a sterilized unit is 1 in 1 million.
The chance of a single surviving organism in a sterilized unit is 1 in 1 million.
Signup and view all the answers
Membrane filtration is the preferred sterilization testing method for food products.
Membrane filtration is the preferred sterilization testing method for food products.
Signup and view all the answers
Poor circulation of steam can negatively influence sterilization outcomes.
Poor circulation of steam can negatively influence sterilization outcomes.
Signup and view all the answers
Prions are less resistant to sterilization methods compared to Gram positive bacteria.
Prions are less resistant to sterilization methods compared to Gram positive bacteria.
Signup and view all the answers
Filtration completely destroys microorganisms rather than merely removing them.
Filtration completely destroys microorganisms rather than merely removing them.
Signup and view all the answers
Heat-based sterilization methods include both moist and dry heat techniques.
Heat-based sterilization methods include both moist and dry heat techniques.
Signup and view all the answers
Gas-based sterilization techniques are considered more reliable than heat-based methods.
Gas-based sterilization techniques are considered more reliable than heat-based methods.
Signup and view all the answers
A pore size of 0.22µm is typically used in filtration for removing most microorganisms.
A pore size of 0.22µm is typically used in filtration for removing most microorganisms.
Signup and view all the answers
Ethylene oxide is a safe and non-toxic option for gas sterilization.
Ethylene oxide is a safe and non-toxic option for gas sterilization.
Signup and view all the answers
Ionizing radiation includes ultraviolet light as one of its types.
Ionizing radiation includes ultraviolet light as one of its types.
Signup and view all the answers
All sterilization methods pose no risk of product damage.
All sterilization methods pose no risk of product damage.
Signup and view all the answers
Depth filters trap particles within the filter, while membrane filters retain particles on the surface.
Depth filters trap particles within the filter, while membrane filters retain particles on the surface.
Signup and view all the answers
Moist heat sterilization requires temperatures exceeding 150°C to be effective.
Moist heat sterilization requires temperatures exceeding 150°C to be effective.
Signup and view all the answers
Study Notes
MPharm Programme: Purification & Sterilization
- The presentation covers purification and sterilization processes within the MPharm program.
- Dr Callum Cooper is the lecturer.
- A recommended resource is "Principles and Practice of Disinfection, Preservation and Sterilization" (5th edition) by Russell, Hugo & Ayliffe.
Learning Objectives
- Introduction to downstream processing
- Purification
- Sterilization
- Various sterilization processes
- Sterility checking
- Sterility testing
Recap (MAb Upstream Process)
- The process starts with thawing and expanding cells.
- Cells are further expanded in seed bioreactors.
- The antibody is expressed into the medium in a production bioreactor.
- Filtration and centrifugation processes follow.
- Cell culture broth is harvested.
Recap (Monoclonal Antibody Downstream Process)
- The process begins with protein A chromatography.
- Polishing chromatography steps remove impurities.
- Orthogonal steps remove viruses (low pH viral inactivation, viral filtration).
- The final step is ultrafiltration/diafiltration to formulate and concentrate the product.
Purification
- Separation of products from production mixtures.
- Removal of unwanted components or contaminants.
- Methods include: sedimentation, precipitation (e.g., heat, pH, organics), centrifugation, adsorption (e.g., ion exchange, immuno-affinity), microfiltration (specified MWCO).
- Purification minimizes side effects while maintaining yield.
- Yield depends on the number of steps and product loss in each step.
Downstream Processing: Sedimentation and Precipitation
- Sedimentation speed depends on cell size, density, and mixing speed.
- Precipitation lowers solute (media) solubility, causing product to precipitate out of solution.
- Methods can be chemical or based on temperature/pH changes.
- Precipitation is used in recombinant DNA polymerase production.
Downstream Processing: Centrifugation
- Application of centrifugal force to separate products based on density differences.
- Denser particles move outward first.
- Requires consideration of component density relative to the medium.
- Factors affecting separation include particle size, density, and relative rotational speed (rpm).
Downstream Processing: Adsorption
- Chromatography-based approach.
- Liquid phase passes through a semi-solid phase.
- Ion exchange binds proteins based on their charge.
- Can be used for capturing or allowing passage of specific proteins.
- Immuno-affinity uses antigenic regions to bind contaminants.
- Common use: removal of bacterial endotoxins (LPS).
Sterilization
- Process that removes or kills all forms of life (bacteria & fungi).
- Can also disable/destroy/remove infectious proteins (e.g., prions).
- Used when a medical product will interact with bodily defenses against infection (e.g., IV administration, broken skin contact, or mucosal surfaces).
Microbial Sensitivity to Sterilization
- Different microbes have varying sensitivities.
- Sensitivity isn't tied to a specific sterilization method but influences method choice.
- Higher resistance is seen in prions, bacterial spores, and gram-negative bacteria, with progressively lower resistance exhibited by other organisms. Prions are notoriously hard to sterilise.
Selection of Sterilization Method
- All methods have some product damage risk; harsh methods impact stability and efficiency.
- The level of microbial reduction must be chosen to minimize product damage.
- 5 common methods recognized by the European Pharmacopoeia (2002): steam, gas, filtration, dry heat, and ionizing radiation.
Downstream Processing: Filtration
- Removes, rather than destroys, microorganisms.
- Filtration efficacy assessed by bacterial count reduction.
- Pore size affects contaminant retention.
- 0.22µm pore size is a widely used standard.
- Other factors: membrane composition, MWCO (molecular weight cut-off).
- Major uses include heat-sensitive solutions, air, gases, and biological products.
Filtration (Methods)
- Membrane filters: particles retained on the filter surface (sieving).
- Depth filters: particles trapped within the filter structure.
- Combinations of depth filters and membranes are possible.
Heat-Based Sterilization Methods
- Moist heat (e.g., autoclave): uses hydrolytic action, typically at >120°C and 1 atm pressure.
- Dry heat (e.g., oven): utilizes oxidative action, with temperatures typically at >150°C.
Gas-Based Sterilization Methods
- Ethylene oxide (EtO): chemically reactive gas.
- Formaldehyde: chemically reactive gas.
- Packaging materials must be EtO permeable.
- Less reliable than heat-based methods, but utilized for temperature-sensitive equipment - (surgical instruments).
Sterilization Using Radiation
- Ionizing (y-rays, accelerated electrons, x-rays) and non-ionizing (UV light).
- Ionizing radiation methods require shielding to prevent damage to surrounding material.
- UV light is best for surface disinfection and certain air/shallow water applications (not ideal for deep water).
Sterilization
- Sterilization process success depends on numerous factors, including equipment maintenance and steam circulation.
Checking Sterilization
- Sterilization success can be verified by physical indicators, chemical indicators, and biological indicators.
Physical/Chemical Sterilization Indicators
- Temperature/pressure record charts.
- Thermometer probes (thermocouples).
- Chemical indicators (e.g., autoclave tape).
Biological Sterilization Indicators
- Standardized bacterial spore preparations (non-pathogenic).
- Spores (e.g., Geobacillus stearothermophilus) are placed in test packs around the sterilizer.
- Post-sterilization, spores grow in nutrient medium; visible changes (like color changes) confirm sterilization process success.
Sterility Assurance
- Microbial populations decrease exponentially with sterilization exposure, independent of initial numbers; sterile means no detectable survivors.
Sterility Testing
- Assesses the absence of microbial contamination after sterilization.
- Membrane filtration is a common method (liquid products).
- Sterile filters collect microorganisms from liquid products.
- Product transferred to media for incubation (long incubation times).
- Turbidity visually confirms contamination.
Issues with Sterilization (Failures)
- Can lead to product recall, product failure, regulatory issues, and economic losses for businesses.
Summary (Presentation Material)
- Subsequent manufacturing stages (purification/sterilization).
- Different purification methods.
- Risks and benefits of purification.
- Sterilization concept and methods.
- Suitable method selection for various product types.
Extra Reading
- "Russell, Hugo & Ayliffe's principles and practice of disinfection, preservation and sterilization".
- "Hugo and Russell's pharmaceutical microbiology".
Studying That Suits You
Use AI to generate personalized quizzes and flashcards to suit your learning preferences.
Related Documents
Description
Test your knowledge on various downstream processing techniques such as immuno-affinity processes, ion exchange chromatography, and sterilization methods. This quiz covers essential concepts related to the purification of products in biomedical fields. Challenge yourself to see how well you understand these important processes.
