Terminal sterilization

							TERMINAL STERILIZATION

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Contents
Introduction Concepts Thermal sterilization Moist heat sterilization Dry heat sterilization Gamma irradiation Ethylene oxide sterilization Indicators for evaluating sterilization process Validation of autoclave cycle Sterility testing Regulatory requirements
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Introduction
Sterility: Total absence of viable life forms Assurance of sterility: “A satisfactory result only indicates that no contaminating microorganism has been found in the sample examined in the conditions of the test.”

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Introduction
Terminal sterilization: When products are filled and hermetically sealed within their final containers before being exposed to a sterilization treatment
Terminal sterilization Thermal Moist heat sterilization Dry heat sterilization Non thermal Gamma irradiation Chemicals: Ethylene oxide
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Concepts
D value/ DT value: The time taken at a particular temperature T to reduce a population of microorganisms to 10% of its initial number. D = ___U______ log No- log Nu U = Exposure time, under specific conditions No= Initial microbial population Nu= Microbial population after receiving U time
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Concepts
E.g.: After 5 min of product exposure to a temperature of 110°C, the microbial population reduced from 2x106 to 6x103 D 110 = ________5min_________ =3.28 min log (2x106 ) - log (6x103)

Z value: The number of degree of temperature change necessary to change the D-value by a factor of 10.

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Concepts
F value (lethal rate, instantaneous Fo): It is a measurement of sterilization effectiveness. F(T,z) is defined as the equivalent time at temperature T delivered to a container or unit of product for the purpose of sterilization, calculated using a specific value of z. FO - Sterilization Process Equivalent Time The equivalent number of minutes at 121°C delivered to a unit by a sterilization process calculated using a z-value of 10°C. If, FO = 8 minutes - the cycle delivered a microbial lethality equivalent to 8 minutes at 121°C
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Concepts
PNSU - Probability of a Non-Sterile Unit (Nu) The extrapolated number of of living microorganisms per defined unit after a given equivalent heating time U at a specific temperature T. The probability of a unit (product container) being non-sterile after the application of a lethal agent PNSU of 1 in 106 -- the probability that a unit is non-sterile is one in a million F0 = D121 (log No- log Nu) E.g.: If D121 = 1min, No= 102 and Nu= 10-6, Fo = 1min (log 102- log 10-6) Fo = 8 minutes
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Concepts
FzT = Fz121/ L FzT = equivalent time at temperature T to achieve a specific lethality for contaminants with a value Fz121 = Fo when z value of 10°C is valid L= Lethal rate = (10 [T-121]/z) specified z

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Concepts
Over kill sterilization Provides a minimum 12 log reduction of a resistant Biological Indicator with a known D-value of not less than 1 minute. Requires: minimal information on the bioburden

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Concepts
Bioburden/Bioindicator Sterilization Provides a probability of survival of less than 1 in 106 for the bioburden as demonstrated using a resistant Biological Indicator (BI) with a known Dvalue. Biological Indicator may not be inactivated Requires information on the numbers and heat resistance of the BI. Requires ongoing bioburden.
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monitoring

or

control

over

Minimal information on the bioburden

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Thermal sterilization
The lethal effectiveness of heat on microorganisms depends upon: Degree of heat Exposure period Moisture present Mechanism: Coagulation of the protein of the living cells

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Dry Heat sterilization
Two hours exposure to a temperature of 180°C or 45 min at 260°C kills spores as well as vegetative forms of microorganism Ovens used: Natural convection: currents formed by rise of hot air Forced convection: blower to circulate the heated air Factors affecting: Thermal increment time of chamber and product Hold period at max temperature Cooling time
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Dry Heat sterilization
Cellulose materials , chemicals, rubbers, thermoplastic materials cannot be sterilized Used generally in case of glassware, metal-ware and anhydrous oils Dry heat at 250°C is capable of destroying endotoxins within a reasonable time frame.

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Moist heat sterilization
The basis for this process is: “No living thing can survive 10 minutes direct exposure to saturated steam at 121°C, which is attained under ideal conditions at sea level with 15lb in a steam pressure sterilizer or autoclave” Moist Heat causes coagulation of cell protein at a much lower temperature than dry heat Steam liberates thermal energy equivalent to 540 calories per gram at 100°C and 524 calories per gram at 121°C

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Moist heat sterilization
Autoclave: Pressure vessel. Raises the temperature of the product to the specified level and holds it for the specified exposure period. Two types of loads: Fluid loads- e.g.: 2° pharmaceutical products Porous loads- e.g.: materials that do not contain moisture and where steam must penetrate: containers, closures, machine parts, filters, etc.

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Moist heat sterilization
Steps in autoclaving: Air removal: Removed completely as steam displaces air from autoclave. Trapped air prevents penetration of steam and hence sterilization in that part. Heat up: To the required temperature by passing steam Exposure/Hold time: Temperature and time achieved within the product. Measured generally using thermocouples

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Moist heat sterilization
Drying: Usually by application of vacuum Cooling: It is important to control. Uneven cooling may result in some containers retaining higher internal pressures than those in the vessel. This leads to loss of sterility by displacement of the closures. Cooling to about 80°C.

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Moist heat sterilization
Factors affecting Autoclave cycles: Product characteristics Volume of solution Type of bioburden Container types Lag time before the sterilization temperature is reached in product

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Moist heat sterilization
Types of Autoclave cycles: Compendial cycles: USP, BP and EP quote conditions as 121°C for 15min in absence of prior knowledge of the actual numbers and types of contaminants. Overkill cycles: Where 121°C is unsuitable and in absence of pertinent information, USP suggests 12-log inactivation's of Bacillus stearothermophilus.

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Moist heat sterilization
Assumed D-value cycles: With knowledge of probable number of µorganism contaminating each unit, choose spores Defined cycles:Calculated based on knowledge of numbers and thermal resistances of µorganism likely to contaminate a product a sterilization cycle assuming that all contaminants are as heat resistant as the USP ref

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Moist heat sterilization
Other considerations: Air steam mixtures: To control pressure in the chambers This mixture needs mixing continuously by means of blower Prolonged heat is detrimental to products Thus reduction of cycle time achieved by rapid cooling usually by spraying with cooling water while pressure is reduced

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Moist heat sterilization
Advantages: Rapid, Inexpensive, Effective, Large volumes The processes are predictable and controllable by temperature and time as long as the steam is saturated and air is absent from the product Disadvantages: Cannot use for oily preparation (oil base ointment) Cannot use for moisture sensitive preparations

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Gamma Irradiation
Cold process . Temperature rarely rising above 60°C Critical parameters: The radiation source Method of exposure (i.e., movement through the irradiator), and Type and location of dosimeters used to monitor routine production loads. No microbiological controls are required, neither test for sterility or biological indicators to certify sterility Monitoring the absorbed dose by physical dosimetry
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Ethylene oxide sterilization
Admixed with inert gas such as CO2 It penetrates readily into plastics, paperboards and powder Alkylates essential metabolites in microorganisms particularly affecting reproductive process Pressure chambers Sterilization Cycle: Prehumidification, gas concentration, vacuum and gas pressure cycles, exposure time and temperature, humidity, degassing, aeration, and determination of residuals
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Indicators for evaluating sterilization process
Thermocouple Wax or chemical pellets that melt at 121°C Paper strips impregnated with chemicals that change colour Resistant bacterial spores in sealed ampoules or impregnated in dry paper strips as biological indicators E.g.: For Steam sterilization, the biological indicator is Bacillus stearothermophilus The resistance of the Biological Indicator Relative to that of bioburden needs to be determined
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Validation of sterilization process
In absence of a meaningful end product test, sterility is assured by conformance of the process within its validated tolerances Thus validation of the sterilization process is essential

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Validation of sterilization process
Steps in Validation: Certify that sterilizer is mechanically checked and qualified Select most appropriate biological indicator Experimentally determine D value and Z value of selected biological indicator Determine distribution of heat of defined loading size and configuration. Identify coolest location. Determine heat penetration into product units at coolest points and at any such suspected locations
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Validation of sterilization process
Steps in Validation: Evaluate effect of such cycle parameters as time, temperature and load configuration on the destruction of biological indicator and magnitude of F0 value Determine sterilization process time for achieving desired F0 value and/or desired probability level of biological indicator destruction Repeat process until desired replication is obtained Periodic monitoring of cycle
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Sterility testing
Carried out under aseptic conditions Tests are performed on a sample selected to represent the entire lot of material. Types of tests mentioned in USP 30<71>: Membrane filtration method Direct Inoculation of Culture medium method Sampling sizes are defined in USP 30 <1211>

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Regulatory requirements
European Agency for the Evaluation of Medicinal Products (EMEA): “Where a choice is made not to utilize a method of terminal sterilization, … proper scientific explanation and justification should be provided in the dossier.” Heat labiality of a packaging material should not in itself be considered as adequate justification for not utilizing terminal sterilization, for otherwise heat stable products. Manufacture by aseptic processing only when terminal sterilization is not feasible

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Regulatory requirements
US FDA’s Guidance for industry: Sterilization Process Validation in Applications for Human and Veterinary Drug Products “Regardless of whether the applicant uses terminal sterilization or aseptic processing to manufacture a drug product that is purported to be sterile, certain information about the validation of that process should be submitted for both of those types of sterilization.”

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Regulatory requirements
USP: By prior approval from FDA, Parametric release USP 30<1222>: Defined as the release of terminally sterilized batches or lots of sterile products based upon the compliance with the defined critical parameters of sterilization without having to perform the Sterility Test requirement.

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Thank You
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