Sterilization process in pharmaceutical industry in an autoclave will be considered qualified for consistent and reliable performance (validated) on successful completion of the following tests–
• Bowie – Dick Test for steam penetration (3 trails).
• Empty Chamber Heat distribution studies (3 trails) with temperature mapping probe at different locations of the sterilizer chamber.
• Loaded chamber heat Distribution & penetration studies (3 trails) for each sterilization load of fixed loading pattern –
1) Sterile area garments (20 number Garments packs, Each pack contains 01 Nos. Boiler suit, 01 Nos. Headgear, 02 Nos booties, 01 pairs gloves)
2) Glassware (S.S Mannifold holder (06 holder) 02 Nos, Sampling unit of Compressed air 02 Nos, 500 ml sampling bottles 10 Nos, 250 ml sampling bottles 25 Nos, 04 Nos S.S Bin.)
3) Media (SCDA Medium – 500 ml 09 Nos. Conical flask, SCDM – 100 ml 20 Nos. tubes, FTM – 100 ml 10 Nos. tubes, MSA – 250 ml 01 Nos Conical flask, CA – 250 ml 01 Nos Conical flask, BGA – 250 ml 01 Nos Conical flask, BSA – 250 ml 01 Nos Conical flask, MCA – 250 ml 01 Nos Conical flask, Peptone Water – 500 ml 06 Nos. Conical flask.)
With temperature mapping probes along with Biological Indicator (Geobacillus stearothermophillus spore vials containing 10^6 or more spores per vials) inside the innermost possible layer of the load subjected for sterilization.
• Estimation of the F0 Value achieved during the sterilization hold period at each temperature mapping probe.
To qualify these tests the equipment should fulfill the acceptance criteria described in the individual test procedures. After completion of the qualification tests all the data generated will be compiled together to evaluate ability of the steam sterilizer to sterilize different components at the set parameters and set loading pattern.
• Loaded chamber heat Distribution & penetration studies (3 trails) for each sterilization load of fixed loading pattern –
1) Sterile area garments (20 number Garments packs, Each pack contains 01 Nos. Boiler suit, 01 Nos. Headgear, 02 Nos booties, 01 pairs gloves)
2) Glassware (S.S Mannifold holder (06 holder) 02 Nos, Sampling unit of Compressed air 02 Nos, 500 ml sampling bottles 10 Nos, 250 ml sampling bottles 25 Nos, 04 Nos S.S Bin.)
3) Media (SCDA Medium – 500 ml 09 Nos. Conical flask, SCDM – 100 ml 20 Nos. tubes, FTM – 100 ml 10 Nos. tubes, MSA – 250 ml 01 Nos Conical flask, CA – 250 ml 01 Nos Conical flask, BGA – 250 ml 01 Nos Conical flask, BSA – 250 ml 01 Nos Conical flask, MCA – 250 ml 01 Nos Conical flask, Peptone Water – 500 ml 06 Nos. Conical flask.)
With temperature mapping probes along with Biological Indicator (Geobacillus stearothermophillus spore vials containing 10^6 or more spores per vials) inside the innermost possible layer of the load subjected for sterilization.
• Estimation of the F0 Value achieved during the sterilization hold period at each temperature mapping probe.
To qualify these tests the equipment should fulfill the acceptance criteria described in the individual test procedures. After completion of the qualification tests all the data generated will be compiled together to evaluate ability of the steam sterilizer to sterilize different components at the set parameters and set loading pattern.
A) Bowie – Dick test for Steam Penetration
Objective
Objective of this test is to ensure that the vacuum pulses applied the sterilization hold period are sufficient to remove the entrapped air so as to facilitate rapid and even steam penetration into all parts of the load and maintaining these conditions for the specified temperature holding time (17 minutes at 121 deg.C)
If air is present in the chamber, it will collect within the Bowie – Dick test pack as a bubble. The indicator in the region of the bubble will be of different color as compared to the color on the remaining part of the test paper, because of a lower temperature, lower moisture level or both. In this condition the cycle parameters to be reviewed and the normal sterilization cycles to be modified accordingly.
Bowie – Dick cycle should be normally preceded by a warm – up cycle, as the effectiveness of air removal may depend on all parts of the sterilizer being at working temperature.
If air is present in the chamber, it will collect within the Bowie – Dick test pack as a bubble. The indicator in the region of the bubble will be of different color as compared to the color on the remaining part of the test paper, because of a lower temperature, lower moisture level or both. In this condition the cycle parameters to be reviewed and the normal sterilization cycles to be modified accordingly.
Bowie – Dick cycle should be normally preceded by a warm – up cycle, as the effectiveness of air removal may depend on all parts of the sterilizer being at working temperature.
Procedure
1. Record the set parameters for the Bowie – Dick test cycle in The Annexure.
2. Place one Bowie – Dick test pack near the drain of the sterilization chamber.
3. Select cycle Bowie – Dick on the control panel & operate the steam sterilizer.
4. The print out taken during the Bowie – Dick test cycle & the Bowie – Dick test indicator should be preserved.
5. Compile the observation made during the qualification test for complete evaluation of the system.
2. Place one Bowie – Dick test pack near the drain of the sterilization chamber.
3. Select cycle Bowie – Dick on the control panel & operate the steam sterilizer.
4. The print out taken during the Bowie – Dick test cycle & the Bowie – Dick test indicator should be preserved.
5. Compile the observation made during the qualification test for complete evaluation of the system.
Acceptance Criteria
The Bowie – Dick Test indicator should show a uniform color change, non – uniform change and/or air entrapment (bubble) spot on the pattern indicates inadequate air removal from the sterilization chamber.
Observation & Results
Record the observation and results in format.
B) Empty Chamber Heat Distribution Studies
Objective
Objective of this test is to ensure that, The sterilizer is capable of attaining a temperature of 121 deg.C during the sterilization hold period with steam pressure of 1.1 to 1.2 kg/cm2.
Temperature spread with in the range of 121 deg.C to 124 deg.C during sterilization cycle will demonstrate the uniform heat distribution within the chamber.
Any location where the temperature indicator is placed, not achieving minimum sterilization temperature of 121deg.C through out the sterilization temperature hold will be considered as cold spot.
Temperature spread with in the range of 121 deg.C to 124 deg.C during sterilization cycle will demonstrate the uniform heat distribution within the chamber.
Any location where the temperature indicator is placed, not achieving minimum sterilization temperature of 121deg.C through out the sterilization temperature hold will be considered as cold spot.
Procedure
1.0 Record the set parameter for the sterilization cycle to be operated during the test for empty chamber heat distribution study, in the Annexure.
2.0 Pass minimum 16 no. Temoerature mapping probe into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place. Suspend the probes in the chamber in different position so that probes do not touch any metallic. Record the position of the probes in a respective schematic form.
3.0 Connect the probes to a suitable autoclave data logger, which can scan and print the actual temperature observed at different locations with respect to time.
4.0 Operate the steam sterilizer and also start the autoclave data logger to record actual temperature within the sterilization chamber with respect to time.
5.0 When the sterilization cycle completes, 1) Collect printout of the sterilizer and preserve as Annexure. 2) Download the data-analysis and printing. Record the temperatures observed at different locations in the Annexure.
6.0 If the empty chamber heat distribution study is acceptable perform three consecutive runs to demonstrate cycle and sterilizer reproducibility.
7.0 Compile the data generated during the qualification test for complete evaluation of the system.
2.0 Pass minimum 16 no. Temoerature mapping probe into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place. Suspend the probes in the chamber in different position so that probes do not touch any metallic. Record the position of the probes in a respective schematic form.
3.0 Connect the probes to a suitable autoclave data logger, which can scan and print the actual temperature observed at different locations with respect to time.
4.0 Operate the steam sterilizer and also start the autoclave data logger to record actual temperature within the sterilization chamber with respect to time.
5.0 When the sterilization cycle completes, 1) Collect printout of the sterilizer and preserve as Annexure. 2) Download the data-analysis and printing. Record the temperatures observed at different locations in the Annexure.
6.0 If the empty chamber heat distribution study is acceptable perform three consecutive runs to demonstrate cycle and sterilizer reproducibility.
7.0 Compile the data generated during the qualification test for complete evaluation of the system.
Acceptance Criteria
There should be uniform distribution of heat in the sterilizer chamber during the sterilization hold period and the temperature at each temperature mapping probes should be within the range of 121 deg.C to 124 deg.C during the sterilization hold period.
Observations and Results
Record the observations and results in formats.
C) Loaded Chamber Heat Distribution & Penetration Studies
Objective
Objective of this test is to ensure that, the steam is sufficiently penetrating into the innermost portions of the load subjected for sterilization to achieve desired temperature of 121 deg.C during the complete sterilization hold period with steam pressure of 1.1 to 1.2 kg.cm2.
If Sterilization temperature (121 deg.C) is not achieved through out the cycle, load configuration or size of the load has to be reviewed and cycle to be repeated.
Temperature spread within the range of 121 deg.C to 124 deg.C during sterilization hold period indicate that, uniform heating process which is achieved in the empty chamber heat distribution study is not affected by load. There could be the possibility of lag period for attaining 121 deg.C during heat penetration runs as the probes are placed deep into the load.
Any location where the temperature indicator is placed, not achieving minimum sterilization temperature of 121deg.C during sterilization temperature hold period will be considered as cold spot.
If Sterilization temperature (121 deg.C) is not achieved through out the cycle, load configuration or size of the load has to be reviewed and cycle to be repeated.
Temperature spread within the range of 121 deg.C to 124 deg.C during sterilization hold period indicate that, uniform heating process which is achieved in the empty chamber heat distribution study is not affected by load. There could be the possibility of lag period for attaining 121 deg.C during heat penetration runs as the probes are placed deep into the load.
Any location where the temperature indicator is placed, not achieving minimum sterilization temperature of 121deg.C during sterilization temperature hold period will be considered as cold spot.
Procedure
1.0 Record the set parameter for the sterilization cycle to be operated during the test for loaded chamber heat penetration study in the Annexure.
2.0 Pass minimum 16 no. Temperature mapping probe into chamber through the port provided. Seal the port with silicone sealant so that steam leakage does not take place. Place the probes inside the load components, which are supported to be most difficult points for steam penetration, also place biological indicator along with temperature mapping probe (12 Nos.). Record the position of the probes and biological indicators in a representative schematic form.
3.0 Connect the probes to a suitable data logger, which can scan and print the actual temperature with respect to time.
4.0 Operate the steam Sterilizer and also start the data logger to record the actual temperatures within the sterilization chamber with respect to time.
5.0 When the sterilization cycle completes, 1) Collect printout of the sterilizer and preserve as Annexure. 2) Download the data-analysis and printing. Record the temperatures observed at different locations in the Annexure. 3) Aseptically collect the exposed biological indicators and send the indicators to microbiology lab for further incubation and observed the results.
6.0 If the load penetration study is acceptable perform three consecutive runs to demonstrate cycle and sterilizer reproducibility.
7.0 Compile the data generated during the qualification test for complete evaluation of the system.
2.0 Pass minimum 16 no. Temperature mapping probe into chamber through the port provided. Seal the port with silicone sealant so that steam leakage does not take place. Place the probes inside the load components, which are supported to be most difficult points for steam penetration, also place biological indicator along with temperature mapping probe (12 Nos.). Record the position of the probes and biological indicators in a representative schematic form.
3.0 Connect the probes to a suitable data logger, which can scan and print the actual temperature with respect to time.
4.0 Operate the steam Sterilizer and also start the data logger to record the actual temperatures within the sterilization chamber with respect to time.
5.0 When the sterilization cycle completes, 1) Collect printout of the sterilizer and preserve as Annexure. 2) Download the data-analysis and printing. Record the temperatures observed at different locations in the Annexure. 3) Aseptically collect the exposed biological indicators and send the indicators to microbiology lab for further incubation and observed the results.
6.0 If the load penetration study is acceptable perform three consecutive runs to demonstrate cycle and sterilizer reproducibility.
7.0 Compile the data generated during the qualification test for complete evaluation of the system.
Acceptance Criteria
There should be uniform distribution & penetration of heat in the load subjected for sterilization during the sterilization hold period and the temperature at each temperature mapping probe should be within the range of 121 deg.C to 124 deg.C during the complete sterilization hold period.
Observations and Results
Record the observations and results in formats enclosed as Attachement
D) Bio-challenge studies
Objective
The steam sterilization process in pharmaceutical industry, when challenged with Geobacillus stearothermophillus Biological indicator spore vial, spore population of NLT 106 spores/vial, should reduce bacterial load by mean of Sterility Assurance Level (SAL) 106
On incubation of the loaded biological indicator, if growth is observed, then the sterilization cycle parameters to be reviewed.
On incubation of the loaded biological indicator, if growth is observed, then the sterilization cycle parameters to be reviewed.
Procedure
1.0 Determine the initial counts of biological indicator.
2.0 Collect the exposed indicator (during the loaded chamber heat distribution & heat penetration studies) by using sterile forceps and scissors in a 100 ml beaker and then send to microbiology lab for incubation (Incubate the vial at 55 to 60 deg.C for 48 hours)
3.0 Keep one vial as a negative control provided by the Mfg of biological indicator as well as one vial as a positive control (unexposed vial biological indicator).
4.0 Observe any growth (purple color – sterile, yellow color – Non sterile) in the vial daily. Record the observations on daily basic in the Annexture.
5.0 Compile the data generated during the qualification test for complete evaluation of the system.
2.0 Collect the exposed indicator (during the loaded chamber heat distribution & heat penetration studies) by using sterile forceps and scissors in a 100 ml beaker and then send to microbiology lab for incubation (Incubate the vial at 55 to 60 deg.C for 48 hours)
3.0 Keep one vial as a negative control provided by the Mfg of biological indicator as well as one vial as a positive control (unexposed vial biological indicator).
4.0 Observe any growth (purple color – sterile, yellow color – Non sterile) in the vial daily. Record the observations on daily basic in the Annexture.
5.0 Compile the data generated during the qualification test for complete evaluation of the system.
Acceptance Criteria
No bacterial growth should observed during the incubation period of 48 hours at 55 to 60 deg.C.
Observations and Results
Record the observations and results in formats enclosed as Attachement.
E) Estimation of F0 Value
Objective
The calculated F0 value should not be less than the biological F0 value at all temperature mapping locations during the sterilization hold period.
Procedure
1.0 Record the temperature at all temperature mapping probes during the sterilization hold period in the Annexure.
2.0 Calculate the F0 value at each temperature mapping probe by using the equation as below.
3.0 Record the F0 value (Results) in the Annexure.
4.0 Compile the data generated during the qualification test for complete evaluation of the system.
2.0 Calculate the F0 value at each temperature mapping probe by using the equation as below.
3.0 Record the F0 value (Results) in the Annexure.
4.0 Compile the data generated during the qualification test for complete evaluation of the system.
Calculation
F0 = dt S10(T-121)/z
Where
dt = Time interval between two following temperature measurements (1 minutes).
T = The observed Temperature at that particular time.
Z = The change in the heat resistance of Geobacillus stearothermophillus spores as temperature is changed (10 deg.C).
Where
dt = Time interval between two following temperature measurements (1 minutes).
T = The observed Temperature at that particular time.
Z = The change in the heat resistance of Geobacillus stearothermophillus spores as temperature is changed (10 deg.C).
Acceptance Criteria
The calculated minimum F0 value (by equation) should be more than biological F0 value for the biological indicator vial exposed for the bio-challenge studies.
- The biological F0 value for the specific biological indicator spore vial is calculated as per the following equation
F0 = D121 (Log A – Log B)
Where
D121 = D value of the of the biological indicator at 121 deg.C.
A = Biological indicator concentration or spore population.
B = Desired level of non – sterility. (10 deg.C)
- The biological F0 value for the specific biological indicator spore vial is calculated as per the following equation
F0 = D121 (Log A – Log B)
Where
D121 = D value of the of the biological indicator at 121 deg.C.
A = Biological indicator concentration or spore population.
B = Desired level of non – sterility. (10 deg.C)
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