Protocol for steam sterilizer and load Pattern

March 22, 2024February 12, 2020 by Pharmaceutical Guidanace

Protocol for steam sterilizer and load Pattern

MODEL :HORIZONTAL-RECTANGULAR

LOCATION :VIAL WASHING ROOM

TABLE OF CONTENTS

Protocol Approval
Objective
Scope
Equipment Description
Responsibilities And Identification Of Execution Team
Test Procedures
Recording Of Observations
Discrepancy And Corrective Action Report
Compilation, Review And Summary Report
Appendix
Revalidation Criteria
Schematics of Stem Quality Test Apparatus

PROTOCOL APPROVAL

Signing of this approval page of protocol indicates agreement with the validation approach described in this document. This protocol cannot be executed until approved by following personnel.

OBJECTIVE

The objective of this Validation Protocol is to obtain a high degree of assurance that the steam sterilization process is capable of sterilizing the specified loads using High Pressure and High Vacuum Steam Sterilizer (Bung Processor) supplied by M/s Machine Fabrik, Mumbai.

To establish a documented evidence that the Steam sterilizer, as commissioned, will produce acceptable goods when operated in accordance with the process specification.

The sterilization process consistently provides a specified degree of sterility assurance level (SAL) of min 10^-6 for each type of load selected for sterilization.

Also the utilities supplied will meet the predefined specifications and capable of supporting the sterilization process consistently.

Different test procedures to be conducted as described in the preceding sections along with the acceptance criteria to meet for each type of test will qualify the equipment.

SCOPE

This protocol is applicable to P.H.V. Steam Steriliser installed in the vial washing room of Production (sterile) area.

To be performed after the completion and authorization of Performance Qualification as per the validation schedule.

EQUIPMENT DESCRIPTION

H.P.H.V. Steam sterilizer is installed in vial washing room.

Purpose of The Equipment

Purpose of this H.P.H.V. Steam sterilizer is for sterilization and drying of the garments, rubber stopper holding canisters, filling machine parts, filtration accessories and previously washed and siliconized rubber stoppers.

Basic Mode of Operation

The main action of H.P.H.V. Steam sterilizer on all the materials placed within the chamber is that, the steam penetrates into the microorganisms present on and within the material thereby transferring the latent heat, which in turn will coagulate the proteins, imparting a lethal effect on the micro organism. This will lead to the destruction of living organisms.

The H.P.H.V. Steam sterilizer is made-up of SS sheet, which is welded with a ‘U’ profile SS jacket.

The sterilization chamber is provided with two SS sliding doors reinforced with MS support structure. The door is operated with the help of pneumatic cylinders. When both the doors reach the end position the gaskets are pushed out automatically with the help of compressed air ejectors. When the gaskets retract the sterilizer chamber, door slides automatically.

The door sealing is made effective with the help of tubular silicon gasket to ensure proper sealing.

These gaskets are activated with compressed air and retracted with the help of vacuum.

The sterilization chamber is insulated with resin-bonded glass wool, which helps in reducing the heat loss to the environment and ensures uniform distribution of temperature inside the chamber. The insulation is covered with SS cover plate.

To ensure leak tight partition between the sterile area and the loading side a SS flush panel is provided on the partition wall and the outer cover of the Bung processor.

All the joints, crevices are filled with silicon sealant to prevent Leakages.

The autoclave holds the vacuum with a rate of NMT 13 mbar per 10 minutes.

All the automatic valves are closed when autoclave chamber is at the atmospheric pressure. During the vacuum cycle except the vacuum valve all other valves are closed.

The sterilizing grade filters are suitable for integrity testing and steam sterilization.

Both the horizontal sliding doors have an interlocking system in which the door doesn’t open simultaneously. The doors doesn’t open in any case during sterilization.

The Bung processor has the ability to withstand maximum of 3.0 kg / cm² pressure. The system is having an emergency stop so as to stop all the physical movement and operation of the machine immediately, which is located along with the control panel.

H.P.H.V. Steam sterilizer, option between four pre-programmed cycles is available.

Vacuum leak test
Bowie-dick test
Standard sterilization cycle
P.H.V. sterilization cycle / H.P.H.V. Sterilization cycle for bungs

Process Description

The sterilizer is supplied with pure (clean) steam for the chamber and plant steam to the jacket.

The Sterilization cycle can be automatically controlled by the PLC, which can be programmed and protected with three different levels of passwords.

The sterilizer has the capability to handle the following types of cycles automatically.

Vacuum leak test.

This cycle is used to assure the sterilizer chamber integrity towards leakage.

Bowie-Dick test.

This cycle is used to ensure steam penetration in to the packs is appropriate.

Standard cycle.

Gravity displacement steam sterilization cycle.

HPHV cycle.

High Pressure High Vacuum sterilization cycle with vacuum pulses.

All the above cycles are PLC controlled and runs automatically when selected.

Operating cycle for a porous load

The selected operating cycle of a porous load sterilizer normally five stages.

Air removal – Sufficient air is removed from the chamber and the load to permit attainment of the sterilization conditions.
Steam admission – Steam is admitted to the chamber until the specified sterilization temperature is attained throughout the chamber and load.
Holding time – The temperature throughout the chamber and load is maintained within the sterilization temperature band for the appropriate holding time.
Drying – Steam is removed from the chamber and the chamber pressure is reduced to permit the evaporation of condensate from the load by prolonged evacuation.
Air admission – Air is admitted to the chamber until the chamber pressure approaches atmospheric pressure.

Area Description

The HPHV steam sterilizer is located in the washing & sterilization room of the sterile powder injectable filling section with restricted access.
As per the specifications for cleanliness the washing area is designed as class D & sterilization room is designed Grade – C.
The equipment is located such that, it can be attended easily for routine operational, monitoring and maintenance purpose.
One door of the sterilizer opens into the washing & sterilization room for loading and the other door opens into the aseptic area for unloading.
The control panel is located in the washing & sterilization room; a modular room physically separates its major components and utility lines from the washing & sterilization room environment.
The other supporting systems such as, water-ring vacuum pump is located near the sterilizer and the pure steam generator is located in the Utility building, in the area dedicated for water system.

RESPONSIBILITIES AND IDENTIFICATION OF EXECUTION TEAM

Responsibilities: The group comprising of representatives from each of the following departments and they shall be responsible for the overall compliance with this protocol.

Production : Execute the validation activity & provide necessary support

Engineering & Utility :Participate & provide necessary support for the validation activity

Quality Control :Testing of samples as per the test procedures

Quality Assurance :Monitoring, sampling & reviewing the validation activities.

Identification of Executors: The personnel involved in the execution of this protocol shall be recorded Name, Designation, Signature and Date.

TEST PROCEDURES

Following test procedures are followed to qualify the equipment for its performance.

Load Details

1.Steam qualification tests (Non-condensable gases test, Steam super heat test, Steam dryness test) – 3 runs of each test.

2.Vacuum Leak test – 3 runs of each test

3. Empty Chamber Heat Distribution study – With temperature mapping probes at different locations of the sterilizer chamber.

4. Bowie –Dick Test – 3 Trials on 3 different days.

5. Loaded Chamber Heat Distribution Studies – Temperature mapping probes shall be placed out side of the load.

Aseptic area Garments [Min Load]

Rubber stopper holding canisters

Filtration accessories

Filling machine parts

Rubber stoppers [Min Load]

Rubber stoppers [Max Load]

Media vessels with WFI water

Loaded Chamber Heat Distribution Studies

Temperature mapping probes shall be placed inside the innermost part (assumed to be difficult to attain sterilization temperature i.e. cold spot) of the load.

Bio-Challenge studies shall use Bacillus stearothemophilus spore strips (containing 10⁶ or more spores per strip) during the heat penetration studies.

Estimation of the F_O value achieved during the sterilization hold period at each temperature-mapping probe.

Aseptic area Garments [Min Load]

Aseptic area Garments [Max Load]

Rubber stopper holding canisters

Filtration accessories

Filling machine parts

Rubber stoppers [Min Load]

Rubber stoppers [Max Load]

Media vessels with WFI water

Steam Quality Tests

Steam Non-Condensable Gas Test

Tools required: Steam quality Testing kit

Test procedure:

This test is used to demonstrate that the level of non-condensable gases in the steam will not prevent the attainment of sterilization conditions in any part of the load. The method described should be regarded not as measuring the exact level of non-condensable gas, but a method by which the provision of acceptable steam quality can be demonstrated.

The apparatus is shown and described in Figure NO.1 (all sizes are nominal).

Connect the needle valve to the steam service pipe as shown in Figure NO.1 (Refer last page).

Assemble the apparatus so that condensate will drain freely from the long rubber tube into the sampling pipe. If the tube is too short, copper or stainless steel tubing may also be used.

Fill the container with cold water until it overflows. Fill the burette and funnel with cold water, invert them and place them in the container. Draw out any air that has collected in the burette.

With the steam sampling pipe out of the container, open the needle valve and allow steam to purge the air form the pipe.

Place the pipe in the container, locate the end within the funnel, and add more cold water until it flows through the overflow pipe.

Place the empty measuring cylinder under the container overflow.

Adjust the needle valve to allow a continuous sample of steam into the funnel sufficient to cause a small amount of “Steam Hammer” to be heard. Ensure that all the steam is discharged into the funnel and does not bubble out into the container. Note the setting of the needle valve. Close the valve.

Ensure that the container is topped up with cold water and that the measuring cylinder is empty. Draw out any air present in the burette.

Ensure that the sterilizer chamber is empty except for the usual chamber furniture. Select and start the operating cycle.

When the steam supply to the chamber first opens, open the needle valve to the previously noted setting, allowing a continuous sample of steam into the funnel sufficient to cause a small amount of steam hammer to be heard.

Allow the steam sample to condense in the funnel. Any non-condensable gases will rise to the top of the burette. Overspill formed by the condensate and the water displaced by the gases will collect in the measuring cylinder.

When the temperature of the water in the container reaches 70-72°C close the needle valve. Note the volume of gas collected in the burette (Vb) and the volume of water collected in the measuring cylinder (Vc).

Calculate the fraction of non-condensable gases as a percentage as follows.

Fraction of non-condensable gases = 100 x (V b/V c).

The test should be done two more times to check consistency. If the results of the three tests differ significantly, then the cause should be investigated before proceeding further.

Record the observations & results in the annexure-1.

Acceptance Criteria: The measured non-condensable gases in the pure steam should not cross 3.2%

Steam Super Heat Test

Tools required: Testing kit

Test procedure:

This test should normally follow a satisfactory test for non-condensable gases.

This test, and the subsequent dryness value test, requires a pitot tube as shown in figure No.2 (Refer last page). The rest of the apparatus is shown and described in figure No.3. All sizes are nominal.

Fit the Pitot tube concentrically within the steam service pipe as shown in figure No.3 (Refer last page).

Fit the sensor entry gland to the steam service pipe. Insert one of the sensors through the gland and position the axis of the pipe.

Insert the second sensor through the gland in the expansion tube and position it on the axis of the pipe. Wrap lagging around the expansion tube. Push the tube on to the pitot.

Ensure that the sterilizer chamber is empty except for the usual chamber furniture.

Select and start the operating cycle.

From the measured temperatures, note the temperature in the steam service pipe (for use in the dryness test) and in the expansion tube (Te) when the steam supply to the chamber first opens. Calculate the superheat in ^oC from the following equation:

Superheat = Te – To

Where:

To – is the boiling point of water at local atmospheric pressure.

Record the observations & results in the annexure-1.

Acceptance criteria: The test should be considered satisfactory if the superheat measured in the expansion tube does not exceed 22^o

Steam Dryness Test

Tools required: Testing kit, balance

Test procedure:

The test is conveniently carried out immediately after the superheat test.

This test requires a pitot tube as shown in Figure No.2. The apparatus is shown and described in figure No.3. All sizes are nominal. A laboratory balance is also required, capable of weighing a load up to 2 kg with an accuracy of 0.1g or better.

If it is not already fitted, fit the Pitot tube concentrically with in the steam service pipe as shown in figure No.3.

If it is not already fitted, fit the sensor entry gland to the steam service pipe. Insert a temperature sensor through the gland and position it on the axis of the pipe.

Connect the rubber tube to the longer of the pipes in the stopper, place the stopper in the neck of the vacuum flask, weigh the whole assembly and note the mass (M1).

Remove the stopper and tube assembly and pour 620 +_ 20 ml of cold water (below 27oC) in to the flask. Replace the stopper and tube assembly, weigh the flask and record the mass (M2).

Support the flask close to the pitot, and ensure that the rubber tube and flask are protected from excess heat and draughts, don’t connect it to the Pitot tube yet.

Introduce the second temperature sensor through the shorter of the two pipes in the stopper and into the water in the flask. Note the temperature of the water in the flask (To).

Ensure that the sterilizer chamber is empty except for the usual chamber furniture. Select and start the operating cycle.

When the steam supply to the chamber first opens, connect the rubber tube to the pitot discharge and wrap lagging around it. Arrange the rubber tube to permit condensate to drain freely into the flask. Not the temperature in the steam service pipe (TS).

When the temperature of the water in the flask is approximately 80^oC, disconnect the rubber tube from the pitot, agitate the flask so that the contents are thoroughly mixed, and note the temperature of the water (T1).

Weigh the flask and stopper assembly and note the mass (M3).

The initial mass of water in the flask is given by Mw = M2 – M1

The mass of condensate collected is given by MC = M3 – M2.

Calculate the dryness value of the steam from the following equation:

D= (T1-T0) (4.18MW + 0.24) / LMC – 4.18 (TS-T1) / L

Where:

T0 = Initial temperature of the water in the flask (oC);

T1 = Final temperature of the water and condensate in the flask (ºC);

TS = Average temperature of the steam delivered to the sterilizer (ºC);

MW = Initial mass of water in the flask (Kg);

MC = Mass of condensate collected (Kg);

L = latent heat of dry saturated steam at temperature TS (kJ Kg-1).

Record the observations & results in the annexure-1.

Acceptance Criteria

The test should be considered satisfactory if the following requirements are met.

The dryness value is not less than 0.90 (if metal loads are to be processed, the dryness value should not be less than 0.92);

Through out the operating cycle, the temperature measured in the steam service pipe is with in 3ºC of that measured during the superheat test.

VACUUM LEAK TEST

Tools required: Calibrated Data logger

Test procedure:

Operate the Steam sterilized as per the SOP.

Measuring the change of vacuum in the chamber when all valves leading to it have been closed and the vacuum source isolated performs the test.

Operate the steam sterilizer empty. Allow the sterilization chamber temperature to stabilize.

When the chamber temperature is stabilized, the vacuum pump starts.

Achieve chamber pressure (on gauge) equivalent to ≤≥ 100-mbar of absolute pressure. Observe channel No.6 of the multi point recorder for vacuum).

When the pressure in the chamber drops below 20 mbar absolute, close all the valves connected to the chamber and stop the vacuum pump. Note the time and the absolute Pressure P1 (achieved in the sterilizer chamber).

Wait for 2 minutes (± 10 s) to stabilize the chamber pressure (This time allowance is provided to allow evaporation of condensate in the chamber) and record the Pressure again (P2).

Wait for further 10 minutes (± 10 s), and then note the pressure for a third time (P3).

Calculate rate of Vacuum drop in the sterilizer chamber by using the equation given below and record the results in Attachment.

Calculate the vacuum leak rate for minute period from the equation:

Vacuum leak rate = (P2-P3) / mbarmin^-1

Where,

P2 = Pressure (vacuum) observed after 2 minutes of achieving the desire level of pressure.

P3 = Pressure (vacuum) observed after 10 minutes of achieving pressure P2

Record the observations and results in annexure-1.

The printout/ strip chart taken during the test should be enclosed as attachment.

Acceptance Criteria

The absolute pressure (P2) at the start of the 10-minute period is less than 700 mbar.

The vacuum leak rate does not exceed 1.3mbar/min or the rate of vacuum drop at the end of 10 minutes holding time should not be more than 13 mbar.

BOWIE – DICK TEST FOR STEAM PENETRATION

Sterilization is achieved by the rapid and even penetration of steam into all parts of the load and the maintenance of these conditions for the specified holding time. To ensure this, it is essential to remove air from the chamber and load, and to provide a steam supply, which contains a minimal volume of non-condensable gases. Any residual air and non-condensable gases will become concentrated as a bubble in the load and inhibit steam penetration.

The Bowie-Dick test shows whether or not steam penetration of the test pack is even and rapid, and thus by implication that air or other non-condensable gases are not present. It does not confirm that the sterilization conditions in the load have been achieved.

Tools required: The Bowie-Dick test uses a Class B chemical indicator contained within a standard test pack.

Test procedure:

Operate the Steam sterilized as per the SOP set the PLC parameters as per the annexure-3.

The Bowie-Dick test is normally preceded by a warm-up cycle. This cycle is necessary because the effectiveness of air removal may depend on all parts of the sterilizer being at working temperature. A satisfactory sterilizer may give a fail result if this is not done.

Remove the wrapping from a standard test pack and place the indicator paper in the sheet located nearest to the centre of the pack. Reassemble and secure the pack and replace the wrapping.

Place the test pack in the chamber with the bottom of the pack supported 100-200 mm above the centre of the chamber base.

Select the Bowie-Dick on the PLC, to operate the steam sterilizer automatically.

Ensure that the holding time will not be longer than that specified above. (If this time is exceeded, the indicator may change in such a way as to make it difficult to detect the variations that would indicate a fail condition).

Start the operating cycle.

When the cycle is complete, remove the indicator paper from the test pack.

The printout/ strip chart taken during the Bowie-Dick test cycle & the Bowie-Dick test indicator should be preserved along as attachment.

Acceptance Criteria

The test should be considered satisfactory if the following requirements are met:

There is a uniform change throughout the indicator;

The automatic controller indicates that a Bowie-Dick test cycle has just been completed.

No change, non-uniform change and/ or air entrapment (bubble) spot on the pattern indicates inadequate air removal from the sterilization chamber.

It is important to compare the colour of the indicator at the corners of the paper with that at the centre so that any difference can be clearly seen. If there is any discernible difference the test should be recorded as failed, and the paper marked accordingly. A large area of unchanged indicator points to a gross failure.

The indicator paper should be marked with the result and kept for reference for at least three months. (The chemical reaction continues during this time and the paper may be discarded when the indicator becomes unreadable.)

An unsatisfactory test result indicates that the machine should not be used until the fault has been rectified. (It is important to realise that if a sterilizer fails to pass the Bowie-Dick test it cannot be made safe simply by increasing the holding time until a uniform colour change is produced. A failed sterilizer is in urgent need of skilled attention (Ref: HTM2010, Version 2.0).

F₀ VALUE CALCULATION FOR THE BIOLOGICAL INDICATOR USED

Required F₀ Value Calculation For The Biological Indicator Used

Calculate the required F₀value for biological indicator exposed during the sterilization as per the formula given below.

F₀ = D121 (log A – log B) —-(a)

Where,

D121 = D value of the biological indicator at 121ºC

A = Biological indicator concentration or spore population

B = Desired level of non-sterility (PNSU i.e. probability of non-sterile unit).

Example – Biological indicator strips supplied by … Co., Lot # … Expiry … D121 =1.7

Min., spore population (A) = 2.1 x 10⁶ are used for bio-challenge study of a sterilization cycle, which is designed to achieve 12-log reduction, i.e. PNSU 10-6. Biological F0 value for this indicator can be calculated by equation (b) and the results are

F0 = 1.7 x (log 2.1 x 10⁶ –log 10^-6)

F0 = 20.92 min.

Therefore, the minimum F0 value required for more than 6 log reduction of the Bacillus sterothermophilus indicator should not be less than 20.92minutes.

Record the observations.

F₀ Value Calculation For The Biological Indicator Used after Heat Distribution And Penetration Study.

The actual observations obtained during the Heat distribution & Penetration studies at different locations were obtained in the temperature data (printout) and the observed temperatures were subjected for calculation of F₀ values at that particular location.

The calculations are carried out automatically by using the following equation (b) and the lethality factor computed is given in the cumulative lethality chart (printout) and Fo chart (bar graph printout).

F₀ = Dt å 10^(T-121)/Z …………… (b)

F₀ = Dt å (Sum of lethality factors)

Where,

Dt = the time interval between successive temperature measurements (1 min.).

T = the observed temperature at that particular time (as per the actual temperatures recorded)

Z = the change in the heat resistance of Bacillus Stearothermophilus spores as temperature is changed (10ºC).

Acceptance Criteria: The Calculated minimum F₀ value by equation (a) (i.e. from the cumulative lethality chart) should be more than the biological F₀value for the biological indicator exposed for the bio-challenge study. i.e., the heat penetration data within the intended loads meets the requirements of the acceptance criteria of minimum 12 log reduction and the process lethality obtained by sterilization cycle is sufficient to provide minimum 6-log reduction of challenged biological spores.

Estimation of Sterility Assurance Level (SAL)

To demonstrate the degree of process lethality in terms of Sterility Assurance Level (SAL) for the sterilization autoclave for individual load configuration based on the minimumF₀

Procedure:

Sterilization Assurance Level is the expected probability of an item being on sterile after exposure to a valid sterilization process.

SAL is a level of microbial inactivation is described by an exponential function.

The SAL of 10^-6 indicates that the probability of a single viable microorganism being present on sterilized item is one in one million after the item has undergone a sterilization process.

A microbial survival probability (SAL) of 10^-6 is considered for steam sterilization process.

The SLR (Spore Log Reduction) of steam sterilization process shall be calculated by using the following equation:

SLR = Log of BI population + 2+ Log of desired SAL

SLR = Log 10⁶ + 2 + log of 10⁶

SLR = 6 + 2 + 6

SLR = 14

The SAL calculation can be done by using following equation:

Identify the minimum F₀ value obtained from the study and determine the D-value. The calculate the SLR and SAL values as follows:

X = SLR – (log of population) – 2

SAL = 10^-x

Example: Minimum F₀ of 28 and D-value of 2 mins considered then the SAL shall be calculated as:

= 28/2 = 14

X = SLR – (log of population) – 2 = 14 – 6 -2 = 6.

SAL = 10^-x= 10^-6

Acceptance Criteria: The calculated minimum F₀ value shall provide minimum SAL of 10^-6 for overkill approach.

Heat Distribution Empty Chamber

Tools required: Calibrated data logger.

Test procedure:

Pass 12 no. Temperature mapping probes 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 as per annexure-2 so that probes do not touch any metallic surface, also place biological indicators along with each temperature-mapping probe in the sterilizer chamber.

Connect the probes to a suitable data logger, which can scan and print the actual temperature observed at different locations with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

Operate the steam sterilizer as per SOP, and also start the data logger to record actual temperatures with in the sterilization chamber with respect to time.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

If the temperature results obtained from the empty heat distribution study are satisfactory, perform (repeat) two more times to check for reproducibility and to establish permitted tolerances as described in the acceptance criteria.

Record all the observations in the annexure-1.

Acceptance criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Distribution Study-Filling Machine Parts

Load material: Following parts are included in the load

Hopper Load Assembling Unit

Powder hopper – 1 number

Rubber stopper hopper – 1 number

Rubber stopper hopper chute – 1 number

Powder wheel with pistons – 10 numbers

1 number of 0.2-micron Catridge filter with housing for N₂/ CO₂ filtration

2 nos. of vent filters of 0.2-micron porosity for nitrogen filtration at dosing stations

Suitably wrapped silicone tubing & Forceps

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam distribution, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

Operate the steam sterilizer as per SOP and also start the data logger to record actual temperatures with in the sterilization chamber with respect to time. All the times set on PLC, Strip chart recorder, Printout and data logger must correspond to each other.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

If the results obtained from the heat Distribution study are satisfactory, perform (repeat) two more times on each load to check for reproducibility and to establish permitted tolerances as described in the acceptance criteria.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Distribution Study – Filtration Accessories

Tools required: Load material as follows.

293 mm diameter membrane holder previously washed with WFI and fitted with 0.2-micron N66 membrane filter and 2 micron pre-filter with suitable silicone tubing for disinfectant filtration.

22-liter pressure vessel previously washed with WFI for storage of sterile filtered disinfectants.

Silicon tubing required for water filtration and clamps required for fixation of the membrane holder and tanks assembly.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam distribution, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

If the results obtained from the heat penetration study are satisfactory, perform (repeat) two more times on each load to check for reproducibility and to establish permitted tolerances as described in the acceptance criteria.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Distribution Studies

Heat Distribution Study of Garment Load

Load material: as per the table below.

Maximum

06 pairs of primary garments of 06 tye-vak bags

06 pairs of secondary garments in 06 tye-vak bags

02 tye-vak bags consisting 08 mopping dusters in each tye-vak bag

Minimum

01 pairs of primary garments in 01 tye-vak bags

01 pairs of secondary garments in 01 tye-vak bags

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam distribution, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Distribution – Rubber Stoppers Minimum & maximum

Tools required: Load material as follows.

Maximum load: 30,000 nos. of previously washed and siliconised 20 mm stoppers loaded in 8 nos. of SS perforated cassettes (3,720 nos. approximately in each cassette amounting to 30,000 nos.) These cassettes are loaded into the rotating carriage of the bung processor and the carriage is loaded into the chamber of the bung processor.

Minimum load: 2,200 nos. of previously washed and siliconised 20 mm stoppers loaded in 1` nos. of SS perforated cassettes. These cassettes are loaded into the rotating carriage of the bung processor and the carriage is loaded into the chamber of the bung processor.

Note: The rotating carriage must be stopped during the sterilization and drying cycle, as the temperature sensors are introduced into the inner most portion of the rubber stopper load in the cassettes. This simulates the worst-case condition. Similarly 30,000 nos of rubber stoppers as a maximum load also simulate worst-case condition.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam distribution, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Distribution Study – Rubber Stopper Holding Canisters

Load material: 8 nos. of rubber stopper holding canisters (without perforations and with lids) previously washed with WFI, used for holding washed-siliconised-sterilized-dried, rubber stoppers. Arranged 4 nos. in one row and 2 such rows are placed in the sterilizer chamber amounting to 8 nos. of canisters.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam distribution, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Distribution Study – Media vessels with WFI water

Load material: 4 nos. of pressure vessels (used for media) contains WFI water around 17 – 18 liters placed in the sterilizer chamber.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam distribution, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Heat Penetration Studies

Heat Penetration Study of Garment Load

Load material: as per the table below.

Maximum

06 pairs of primary garments of 06 tye-vak bags

06 pairs of secondary garments in 06 tye-vak bags

02 tye-vak bags consisting 08 mopping dusters in each tye-vak bag

Minimum

01 pairs of primary garments in 01 tye-vak bags

01 pairs of secondary garments in 01 tye-vak bags

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam penetration, these placement shall be uniform and as per the annexure-2.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Also place biological indicator strips along with each temperature-mapping probe.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Aseptically collect the exposed biological indicators and send the indicators after wrapping in a sterile enclosure to microbiology lab. For incubation 7 days at 55ºC -60ºC. Record the observations in the annexure-1.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Biological indicators exposed to the steam sterilization cycle When aseptically collected and incubated should show no growth.

The calculated minimum F₀ value should be more than the required F₀value for the biological indicator.

Heat Penetration Study-Filling Machine Parts

Load material: Following parts are included in the load

Hopper Load Assembling Unit

Powder hopper – 1 number

Rubber stopper hopper – 1 number

Rubber stopper hopper chute – 1 number

Powder wheel with pistons – 10 numbers

1 number of 0.2-micron Catridge filter with housing for N₂/ CO₂ filtration

2 nos. of vent filters of 0.2-micron porosity for nitrogen filtration at dosing stations

Suitably wrapped silicone tubing & Forceps

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam penetration, these placement shall be uniform and as per the annexure-2.

Also place biological indicator strips along with each temperature mapping probe.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Biological indicators exposed to the steam sterilization cycle When aseptically collected and incubated should show no growth.

The calculated minimum F₀ value should be more than the required F₀value for the biological indicator.

Heat Penetration Study – Filtration Accessories

Tools required: Load material as follows.

293 mm diameter membrane holder previously washed with WFI and fitted with 0.2-micron N66 membrane filter and 2 micron pre-filter with suitable silicone tubing for disinfectant filtration.

22-liter pressure vessel previously washed with WFI for storage of sterile filtered disinfectants.

Silicon tubing required for water filtration and clamps required for fixation of the membrane holder and tanks assembly.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam penetration, these placement shall be uniform and as per the annexure-2.

Also place biological indicator strips along with each temperature mapping probe.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Biological indicators exposed to the steam sterilization cycle When aseptically collected and incubated should show no growth.

The calculated minimum F₀ value should be more than the required F₀value for the biological indicator.

Heat Penetration – Rubber Stoppers

Tools required: Load material as follows.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam penetration, these placement shall be uniform and as per the annexure-2.

Also place biological indicator strips along with each temperature mapping probe.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Biological indicators exposed to the steam sterilization cycle When aseptically collected and incubated should show no growth.

The calculated minimum F₀ value should be more than the required F₀value for the biological indicator.

Heat Penetration Study – Rubber Stopper Holding Canisters

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam penetration, these placement shall be uniform and as per the annexure-2.

Also place biological indicator strips along with each temperature mapping probe.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select HPHV sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 2 minutes.

Biological indicators exposed to the steam sterilization cycle When aseptically collected and incubated should show no growth.

The calculated minimum F₀ value should be more than the required F₀value for the biological indicator.

Heat Penetration Study – Media vessels with WFI water

Load material: 4 nos. of pressure vessels (used for media) contains WFI water around 17 – 18 litres placed in the sterilizer chamber.

Test procedure:

Pass 12 no. Temperature mapping probes into chamber through the port of the sterilizer. Seal the port with silicone sealant so that steam leakage does not take place.

Place the probes inside the load components, which are supposed to be most difficult points for steam penetration, these placement shall be uniform and as per the annexure-2.

Place biological indicator Ampoule along with each temperature-mapping probe in side media vessels in such away covers the top, middle and bottom of the vessel.

Connect the probes to suitable data logger, which can scan and print the actual temperature with respect to time.

Select sterilization cycle on control panel and set the parameters as per annexure-3.

When the sterilization cycle is complete

Collect Strip chart from the Strip chart Recorder of the sterilizer and enclose as attachment.

Download the data from data logger into the computer for data-analysis and printing. Enclose the data printouts as attachment. Review and calculate the F₀value for each temperature probe.

Record all the observations in the annexure-1.

Acceptance Criteria

The measured temperatures in the sterilizer chamber should be uniform.

The temperature in the Sterilisation hold time should be within the sterilization temperature band of 121ºC to 124ºC.

The hold time shall be 30min with a lag period of NMT 12 minutes.

The calculated minimum F₀ value from assumed z – value should be more than the required F₀value for the biological indicator.

RECORDING OF OBSERVATIONS

Record the observations of after execution of each test procedures, in the annexure –1 (Recording of Observations For Revalidation).

DISCREPANCY AND CORRECTIVE ACTION REPORT

Document any discrepancies observed during the Revalidation of the equipment in annexure -1. Include the corrective actions of the same. When all the discrepancies are satisfactorily resolved or an approved action plan is developed which ensures that the discrepancy will be resolved.

COMPILATION, REVIEW AND SUMMARY REPORT

Compile and review that all test functions have been completed, reconciled and attached to this protocol. Verify that the approvals for deviations have been taken and are resolved appropriately to the satisfaction.

Revalidation shall be considered acceptable when all the conditions specified in the test procedures have been met.

Prepare the summary report in the annexure –2 (Revalidation Report) and submit this for review, approval and authorisation to Validation Core Team.

APPENDIX

Abbreviations and definitions

RVP : Revalidation Protocol

PLC : Programmable Logic Controller

F_O value: A quantity, measured in minutes, used to determine the efficacy of an operating cycle and equivalent to a continuous period at a temperature of 121°C.

D-value: Decimal reduction value (for Biological Indicators). The time in minutes required to secure the inactivation of 90% of the test organisms under stated exposure conditions.

References

Enclosures

Annexure – 1 (Recording Of Observations For Revalidation)

Annexure – 2 (Load pattern & Justification)

Annexure – 3 (PLC Set Parameters For Different Loads)

Annexure – 4 (Revalidation Report)

REVALIDATION CRITERIA

Revalidation shall be carried out in case of

Change of cycle program
Inclusion of new load
Major modification in the existing equipment / system / utility.
Shifting of the equipment / system from one location to another.
Heat Distribution study for Empty chamber, Heat Penetration Study for Load Chamber [Min 1 trial for each load] every 6months + 1 Month.

Protocol for steam sterilizer and load Pattern

Operating cycle for a porous load

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