Sterilization - Overview

Sterilization - Overview: 

Sterilization - Overview Objectives Discuss definition of “Sterile” Briefly describe sterilization methods. Describe approaches to be used for the validation of a sterilization process using Moist Heat as an example. Describe requirements for routine monitoring and control of sterilization. Review issues that are specific to other sterilization processes.

Sterile Products - Overview: 

Sterile Products - Overview Certain pharmaceutical products must be sterile injections, ophthalmic preparations, irrigations solutions, haemodialysis solutions Two categories of sterile products those that can be sterilised in final container (terminally sterilised) those that cannot be terminally sterilised and must be aseptically prepared

Sterilization - Overview: 

Sterilization - Overview What is the definition of “sterile”? Free from microorganisms In practice no such absolute statement regarding absence of microorganisms can be proven Defined as the probability of 1 in a million of a container being contaminated (10 -6 ) This referred to as the Sterility Assurance Level (SAL) Organisms are killed in an exponential fashion

Definition of “Sterile”: 

Definition of “Sterile” Resistance of an organism is referred as its “D-value” D-value - Time (or dose) required to reduce the population of organisms by 1 log (or 90%)

Sterilization - Overview: 

Sterilization - Overview Commonly used methods of sterilization Moist Heat Dry Heat Gas (Ethylene oxide) Radiation (Gamma or Electron) Filtration Others - UV, Steam and formaldehyde, hydrogen peroxide

Moist Heat: 

Moist Heat Saturated steam Common cycles: 121°C for 15 minutes 134°C for 3 minutes Other cycles of lower temperature and longer time may be used (e.g. 115°C for 30 minutes) Used for sterilization of: terminal sterilization of aqueous injections, ophthalmic preparations, irrigation & haemodialysis solutions, equipment used in aseptic processing.

Moist Heat: 

Moist Heat Not suitable for non-aqueous/dry preparations Preferred method of sterilization

Dry Heat: 

Dry Heat Lethality due to oxidative processes Higher temperatures and longer exposure times required Typical cycles: 160°C for 120 minutes 170°C for 60 minutes 180°C for 30 minutes tunnels used for the sterilisation of glass vials may use much higher temperatures (300°) for a much shorter period.

Dry Heat: 

Dry Heat Used for: glassware and product containers used in aseptic manufacture, non aqueous thermostable powders and liquids (oils) also used for depyrogenation of glassware ( 250°C) (Pyrogens - substances found in cell wall of some bacteria which can cause fever when introduced into the body)

Ethylene Oxide Gas: 

Ethylene Oxide Gas Either pure or in mixtures with other inert gases Requires presence of moisture Complex process Typical cycles: 1-24hours 25-1200 mg/L gas 25-65 °C 30-85% relative humidity

Ethylene Oxide: 

Ethylene Oxide Used for: heat labile product containers surface sterilization of powders Adequate aeration to reduce toxic residues

Radiation: 

Radiation Gamma rays generated by Cobalt 60 or Caesium 137 radionuclides; or Accelerated electrons from an electron generator 25 kilograys (kGy) usual dose dose dependent on bioburden (resistance of organisms not predictable) process must be properly validated used for: dry pharmaceutical products heat labile product containers can cause unacceptable changes

Filtration: 

Filtration Removes organisms from liquids and gasses 0.2 - 0.22 micron for sterilization composed of cellulose esters or other polymeric materials filter material must be compatible with liquid being filtered used for bulk liquids, gasses and vent filters

Validation - Overview: 

Validation - Overview Selection of sterilzation process must be appropriate for product terminal sterilization is the method of choice moist heat (autoclaving) is the most common process used for terminal sterilization product must not be affected by heat container/closure integrity must be established items being sterilised must contain water (if sealed) or material must allow for removal of air and penetration of steam for steam (moist heat) sterilization

Validation - Protocol: 

Validation - Protocol Requirements for Moist Heat Sterilization Other processes follow similar requirements Validation protocol should include the following details for each sterilization process process objectives in terms of product type, container/closure system, SAL required specifications for time, temperature, pressure and loading pattern description of all equipment and support systems in terms of type, model, capacity and operating range

Validation - Protocol: 

Validation - Protocol Moist Heat continued: performance characteristics of all equipment e.g. pressure gauges, valves, alarm systems, timers, steam flow rates/pressures, cooling water flow rates, cycle controller functions, door closure gasketing and air break systems and filters methodology for monitoring performance of equipment and the process and labatory testing methodology personnel responsible for all stages and final evaluation (should have experience and necessary training and be authorized)

Validation - Calibration: 

Validation - Calibration Laboratory testing should be performed by a competent laboratory, methodology should be documented All instruments must be calibrated e.g. temperature recorders and sensors thermocouples pressure sensors for jacket and chamber timers conductivity monitors for cooling water flow meters for water/steam water level indicators when cooling water is used thermometers including those for thermocouple reference, chamber monitoring and laboratory testing

Validation - Calibration: 

Validation - Calibration Indicators should be calibrated physical and chemcial indicators should be tested to demonstrate acceptable response to time and temperature biological indicators should be tested for count and time/temperature exposure response for commercial indicators - test certificate with count and D-value and exposure response should be available. Results acceptable if verified “in house” periodically. In house indicators must be fully characterized (D-value, identification) and appropriate for sterilization process All indicators should be appropriately stored and within expiry

Validation of biological indicators: 

Validation of biological indicators The reality : We do not use the most resistant organisms The predictive behaviour is generally linear only for one process Manufacturers seldom use more “practical” strains (read : “less linear” strains for more economical, inoffensive, self resistance and stability) There is no such thing as a “universal biological indicator” The choice of any particular strain is therefore a manner of arbitrary choice

Biological Indicators : 

Biological Indicators Biological Indicators device consisting of a known number of microorganisms, of a known resistance to a particular sterilization process in or on a carrier and enclosed in a protective package. Organisms are in the form of endospores (not vegetative state) as these are most resistant to sterilization

Validation - Cycle Development: 

Validation - Cycle Development Concept of F o Lethality factor equivalent to time at 121°C 1 minute at 121°C is equivalent to F o of 1. Lethality can accumulate during heat up and cool down phases F o is calculated using the following equation: F o = ΔtΣ10 (T-121/Z) where: “ Δt ” is the time interval between measurements of temperature (T) “T” is the temperature of sterilised product at time (t) “Z” is a temperature coefficient which measures the number of degrees required to change the D-value of an organism by 1 log

D value: principle and logic: 

D value: principle and logic More it is hot, faster the micro organisms die Faster the bugs die, faster is the process The speed of the process is expressed with the D value D value = exposition time required (i n minutes ) to kill 1 log (90%) of the micro organisms A Dvalue of 6 means that it takes 6 minutes to reduce par a factor of 10 (90%) or 1 log the number of micro organisms.

Z value: principle and logic: 

Z value: principle and logic If the temperature is lowered, bugs are killed more slowly and the D value increases (because it takes more time to kill) Conversely if the temperature is higher, faster the bugs die, and lower is the D value Z value = the number of degree of temperature required to obtain a variation of 1 log of the D value For a given micro organism, A Zvalue is a measure of its resistance to heat because higher the Zvalue , more heat is needed to augment the Dvalue by a factor of 1.

Validation - Cycle Development: 

Validation - Cycle Development Two approaches to sterilization Overkill Probability of survival Overkill approach used when the product can withstand excessive heat treatment without adverse effects Cycle should deliver an F o of at least 12 This will achieve a 12 log reduction of microorganisms with a D-value of 1 minute ( Assuming each product unit contains 10 6 organisms a 12 log reduction will result in 10 -6 organisms per unit or probability of survival (SAL) of 1 in a million ) ( Normal bioburden is usually much lower and the organisms normally much less resistant than this )

Validation of Sterilization: 

Validation of Sterilization Basic Principles Installation Qualification (IQ) Ensuring equipment is installed as per manufacturer’s specification Operation Qualification (OQ) Ensuring equipment, critical control equipment and instrumentation are capable of operating within required parameters Performance Qualifcation (PQ) Demonstrating that sterilizing conditions are achieved in all parts of sterilization load Physical and microbiological

Validation - Equipment: 

Validation - Equipment Installation Qualification Ensuring equipment is installed as per manufacturer’s specification considerations for new and existing equipment specifications for the type of autoclave, construction materials, power supplies and support services, alarm and monitoring systems with tolerances and accuracy requirements for existing equipment documented evidence that the equipment can meet process specifications

Validation - Equipment: 

Validation - Equipment Operational Qualification Ensuring equipment, critical control equipment and instrumentation are capable of operating within required parameters Three or more test runs which demonstrate controls, alarms, monitoring devices and operation indicators function chamber pressure integrity is maintained chamber vacuum is maintained (if applicable) written procedures accurately reflect equipment operation pre-set operation parameters are attained for each run

Validation - Performance: 

Validation - Performance Performance Qualification Demonstrating that sterilizing conditions are achieved in all parts of sterilization load Physical and microbiological Physical Heat distribution studies on empty chamber maximum and minimum cycle times and temperatures to identify heat distribution patterns including slowest heating points mulitple temperature sensing devices should be used (thermocouples) location of devices should be documented and ensure that heat distribution is uniform

Validation - Performance: 

Validation - Performance Performance Qualification - Physical (2) Heat distribution of maximum and minimum chamber load configurations multiple thermocouples throughout chamber (not inside product containers) to determine effect of load configuration on temperature distribution temperature distribution for all loads using all container sizes used in production should be tested position of thermocouples should be documented Slowest to heat/cold spots in each run should be documented, inlcuding the drain repeat runs should be performed to check variability temperature distribution profile for each chamber load configuration should be documented

Validation - Performance: 

Validation - Performance Performance Qualification - Physical (3) Heat penetration studies to detect the maximum and minimum temperature within all loads all parts of each load must be on contact with steam need to determine lowest and highest temperature locations and slowest and fastest to heat locations (measured inside product containers) need to consider all variables such as container size, design, material, viscosity of solution and fill volume. Container with maximum fill volume and slowest to heat solution should be used maximum and minimum load configurations for each sterilization cycle using routine cycle parameters

Validation - Performance: 

Validation - Performance Performance Qualification - Physical (4) Heat penetration (2) May be necessary for container mapping for larger volumes - cold spot then used for penetration studies Need to consider effects of packaging e.g. overwrapping Three runs performed once cold spots have been identified to demonstrate reproducibility

Validation - Performance : 

Validation - Performance Performance Qualification - Microbiological Biological challenge studies used when Probability of Survival approach is used may not be necessary when cycle is > 121°C for 15 minutes (except US and Australia) biological indicators (BI) containing spores of Geobacillus stearothermophilus are most commonly used (considered “worst case”). BIs containing other organisms may be used performance studies based on product bioburden require a considerable amount of work indicators should be placed throughout the load, adjacent to thermocouples, at “cold spots” and slowest to heat locations (identified during heat penetration studies) any growth is unacceptable unless processing errors demonsrated

Validation - Performance: 

Validation - Performance Validation report must demonstrate requirements in Validation protocol have been met, any deviations must be justified Requalification must be repeated on an annual basis (usually one run is acceptable) Any changes or modifications must be evaluated may just require requalification any changes to loading patterns, new container/closure systems or cycle parameters require full validation

Other Sterilization Processes: 

Other Sterilization Processes Sterilization using other processes should follow a similar approach as that described for moist heat Validation protocol Equipment calibration Determining the process that will deliver the desired SAL (10 -6 ) IQ, OQ, PQ Requirements for routine monitoring and control

Other Sterilization Processes: 

Other Sterilization Processes Dry Heat Should have air circulation in the chamber Positive pressure in the chamber to prevent entry of non-sterile air HEPA filtered air supplied Biological indicators containing Bacillus atropheus (if used) removal of endotoxin is usually sufficient When removing pyrogens need to validate process using challenge tests

Other Sterilization Processes: 

Other Sterilization Processes Radiation Usually performed by contracting service (need to ensure validation status, responsibilities) Based on bioburden of product being sterilised Biological indicators may be used as additional control but may not be as resistant as naturally occuring bioburden Method defined in International Standard ISO 11137

Other Sterilization Processes: 

Other Sterilization Processes Radiaton (2) Correct dose of radiation (~25 kGy) received by all products (measured with dosimeters) quantitative measurement number, location, within calibration time-limit Radiation sensitive colour discs applied to packaging procedures to distinguish irradiated and non-irradiated materials Variation in density of packaging should be addressed during validation

Other Sterilization Processes: 

Other Sterilization Processes Gasses and Fumigants e.g. ethylene oxide, hydrogen peroxide vapour Only when no other method is suitable Must demonstrate that process does not adversely affect product Packaging must be able to permit ingress of gas and humidity Ensure product load is adequately heated and humidified prior to sterilization (called “conditionning”) need to take into account validation performed in summer or winter

EO VALIDATION OVERVIEW: 

EO VALIDATION OVERVIEW Process development Product compatibility Commissioning PQ – Physical PQ – Microbiological Certification Revalidation

PROCESS CONTROL: 

PROCESS CONTROL Must assure that validated process parameters are met Temperature RH Gas concentration Biological indicators are used to demonstrate lethality Microprocessors are used to control process

RELEASE MECHANISMS: 

RELEASE MECHANISMS Documentation showing that processing specification are met Successful results of tests Sterility of BI EO residues Packaging Pyrogens

PARAMETRIC RELEASE: 

PARAMETRIC RELEASE BIs not used in release Validation more involved Routine control more rigorous AAMI TIR20:2001 “Parametric release for ethylene oxide sterilization”

PRODUCT COMPATIBILITY: 

PRODUCT COMPATIBILITY Post sterilization testing for Device functionality Package integrity and strength Residue dissipation rates Impact of re-sterilization

INITIATING VALIDATION: 

INITIATING VALIDATION Determine the standard Insure appropriate packaging Determine worst case load Determine challenge device Internal Process challenge device (PCD) Select Validation Method BI release Parametric

CHALLENGE DEVICES: 

CHALLENGE DEVICES Internal Challenge Device (ICD) Most difficult to sterilize devices seeded with a BI in the most difficult to sterilize location PCD An external BI test pack that replaces the internal challenge device Should be an equal or more difficult challenge to the process than the ICD Developed using comparative resistance studies

PARAMETRIC RELEASE: 

PARAMETRIC RELEASE Benefits Faster TAT Useful if extended aeration not required Considerations More complicated validation Minimum of 6 or 7 sub lethal cycles Direct measurement of EO, RH and temp. Load configuration becomes more critical

BI RELEASE: 

BI RELEASE BI Overkill (most common) Demonstrate 10 -6 SAL Assume bioburden has lower population & resistance than BI Need a > 12 Spore Log Reduction (SPL) of BI Combined BI/Bioburden Absolute Bioburden (rarely used)

SAMPLE PLACEMENT: 

SAMPLE PLACEMENT Protocol must detail the number and location of all samples in load BI’s Product sterility (if applicable) ETO residuals Product functionality Package integrity LAL

VALIDATION CYCLES: 

VALIDATION CYCLES Fractional cycles Half cycles Full cycles

FRACTIONAL CYCLE: 

FRACTIONAL CYCLE Must be run when bioburden >100 and no comparative resistance studies are performed Desired cycle time must results in some positive BI and sterile product in sterility tests A minimum of 20 product sterility samples (10 TSB, 10 FTM) Product sterility samples must be placed adjacent to BI

HALF CYCLES: 

HALF CYCLES Three half cycles must be run in production chamber with a gas dwell time half the full cycle dwell time The following must be placed in load Temperature and humidity sensors Internal BI External BI (optional) Product sterility samples if comparative resistance studies not done or inconclusive

FULL CYCLE: 

FULL CYCLE A minimum of one full cycle is required for the Micro PQ Three cycles are required to meet residual requirements The following samples are included EO residual Product functionality Packaging integrity External BI (routine release BI) LAL

EO RESIDUAL TESTING: 

EO RESIDUAL TESTING 1 - 3 samples of each type should be tested at a minimum of 3 time intervals from processing (Ex. 1, 3, & 5 days) This must be done after 3 full cycles Testing for EO and ECH Samples must be shipped frozen

ACCEPTANCE CRITERIA: 

ACCEPTANCE CRITERIA Bioburden must be in control Product sterility all neg after half cycles Acceptable B&F test BI Testing Fractional cycle - some should grow Half cycle - all negative Full cycle - all negative

ACCEPTANCE CRITERIA (cont.): 

ACCEPTANCE CRITERIA (cont.) Temperature sensors <10°C Humidity sensors <30% EO residual Product functionality Package integrity LAL