PQ Autoclave

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QUALITY CONTROL Page 1 of 26
PERFORMANCE QUALIFICATION
Equipment Name Autoclave (HPHV Steam Sterilizer) Equipment ID
Protocol Number Effective Date
PROTOCOL FOR
AUTOCLAVE (HPHV STEAM STERILIZER)
Format No:
TABLE OF CONTENTS
1.0APPROVAL SIGNATURE:.....................................................................................................................3
2.0OVERVIEW:.............................................................................................................................................4
2.1 Introduction:....................................................................................................................................4
2.2 Objective:.........................................................................................................................................4
2.3 Purpose & Scope:............................................................................................................................5
2.4 Responsibility:.................................................................................................................................5
3.0EXECUTION TEAM:..............................................................................................................................7
4.0TRAINING RECORDS:...........................................................................................................................8
4.1 Purpose:............................................................................................................................................8
4.2 Scope:................................................................................................................................................8
4.3 Topics:..............................................................................................................................................8
5.0REQUIREMENT FOR PERFORMANCE QUALIFICATION:.........................................................8
6.0SYSTEM / EQUIPMENT DESCRIPTION:...........................................................................................9
6.1 System Equipment Details:............................................................................................................9
6.2 Generic Design of Double Door Autoclave:..................................................................................9
7.0QUALIFICATION PROCEDURE OR METHODOLOGY:.............................................................11
7.1 General Recording Instructions:.................................................................................................11
7.2 General Safety Instruction for Execution:..................................................................................12
7.3 General Steps and Precaution to be followed in Qualification Study:.....................................12
7.4 Selection Criteria for Locations of Thermocouples, Biological and Chemical Indicators for
Loaded Chamber Study:...........................................................................................................13
7.5 Calculation of F0 Values / Sterility Assurance Level:...............................................................13
7.6 Equipment Qualification:.............................................................................................................15
7.7 Steam Quality Test:.......................................................................................................................16
7.8 Vacuum Leak Test:.......................................................................................................................17
7.9 Bowie Dick Type Test:..................................................................................................................18
7.10 Empty Chamber Heat Distribution Study:...............................................................................19
7.11 Loaded Chamber Study:............................................................................................................21
8.0ACCEPTANCE CRITERIA:.................................................................................................................24
9.0REVALIDATION CRITERIA:.............................................................................................................24
10.0QUALIFICATION REPORT:.............................................................................................................24
11.0OBSERVED DEVIATION (IF ANY):................................................................................................24
12.0ABBREVIATIONS:..............................................................................................................................24
13.0LIST OF ANNEXURE:........................................................................................................................26
14.0REFERENCE DOCUMENT (IF ANY):.............................................................................................26
Format No.: QA004-F05-01

1.0 APPROVAL SIGNATURE:

This is specific protocol for Performance Qualification of Autoclave (HPHV Steam Sterilizer) installed
in Media Preparation Room in Microbiology section of Quality Control department. The Author
signature indicates that this document has been prepared in accordance with existing project standards
and adequately reflects the tasks and deliverables necessary for qualification of the facility / equipment
/ process.
Prepared by / Functional area Designation Signature Date
Microbiology Section
Quality Assurance
The reviewer’s signature indicates that, this document has been reviewed and it accurately and
completely reflects the tasks and deliverables necessary for process.
Reviewed by / Functional area Designation Signature Date
Quality Control
Engineering & Maintenance
Validation – QA

The approver’s signature indicates that the documentation and information contained herein complies
with applicable regulatory, corporate, divisional / departmental requirements, and current Good
Manufacturing Practices.
Approved by / Functional area Designation Signature Date
Quality Assurance
2.0 OVERVIEW:
2.1 Introduction:
This protocol defines the methodology for Performance Qualification of Autoclave (HPHV
Steam Sterilizer) installed in the Media Preparation room in Microbiology section of Quality
control department.
2.2 Objective:
The objective of this document is to provide written procedure and/or guidelines and respective
acceptance criteria for the Performance Qualification of Autoclave steam sterilizer /
demonstrate with documented evidence that the equipment produces the required output by
integrating procedures, personnel and systems, by verifying the following parameters –
2.2.1 Steam Quality Test:
To demonstrate that the steam supplied to the autoclave meets the criteria for quality as
defined in HTM 2010 & EN285 w.r.t superheat, dryness & non-condensable gases; and

the steam condensate (tested for physical, chemical and microbial parameters) is as per
the specifications.
2.2.2 Vacuum Leak Test:
To verify & demonstrate that the chamber is integral under vacuum conditions that are /
may be created as a part of air removal stage or drying stage at the end of the cycle.
2.2.3 Empty Chamber Study:
To assess the hottest & coolest locations and identify the locations those are fastest &
slowest to attain the sterilization temperature. This study shall be carried out for standard
steam sterilization and HPHV process.
2.2.4 Bowie – Dick Test:
To demonstrate that the steam penetration of the test pack is rapid, there-by implying
that air and other non-condensable gases are not present.
2.2.5 Loaded Chamber Studies (Distribution & Penetration):
To assess the hottest & coolest locations and identify the locations those are fastest &
slowest to attain the sterilization temperature and demonstrate that every point in the
loaded chamber attains F0 value (lethality) of ≥ 15 minutes.
To establish that the equilibration time to achieve sterilization temperature between the
drain probe and coldest part of the load does not exceed 1 minute (for porous loads
only).
This study shall be done for each defined loading pattern [separate for porous,
unwrapped instruments & fluids] wherein the distribution & penetration study will be
done as a combined cycle. The requirements of automatic control test shall also be
evaluated.
2.3 Purpose & Scope:
After completion of successful Operation Qualification, start the Performance Qualification to
ensure that Autoclave (HPHV Steam Sterilizer) is performing as per its intended use and will
perform reproducibility.
The scope of this Protocol is limited to outline procedure /provide methodology for the
Performance Qualification of HPHV Steam Sterilizer.
2.4 Responsibility:
The validation group comprising of representatives from each of the following departments
shall be responsible for the overall compliance with this protocol:
♦ Quality Control (Microbiology)
♦ Engineering & Maintenance Department
♦ Quality Assurance Department


The Validation team is described through the following Responsibility.
Shall prepare the Qualification Protocol as per the

Executive – QC (Microbiology) guideline of Validation Master Plan in consultation with
QA
Manager – QC, Manager –

Shall review the Qualification protocol for correctness,
Engineering & Maintenance and
completeness and technical excellence.
Executive – QA
Qualification Protocol shall be approved by the QA head

Head – QA
prior to execution
Shall conduct and record the qualification test as per the

Executive QC (Microbiology)
protocol
Engineering & Maintenance Shall identify the instruments for Calibration
Shall review the executed protocol to check the

Manager – QC, Manager –
compliance and corrective action for any discrepancies
Engineering & Maintenance and
found. Also shall prepare the summary and conclusion of
Executive – QA
the qualification study
These personnel shall review the qualification study and

Head – QA its summary report. Approval of the summary report and
conclusion shall be done by this team.
To conduct the qualification study, a team shall be formed. The team shall contain the members
from the, Engineering, Quality control (microbiology) and Quality Assurance departments.
3.0 EXECUTION TEAM:

Following personnel are involved in the completion of this qualification with defined responsibilities
as follows:
Name Designation Department Sign/ Date

4.0 TRAINING RECORDS:

4.1 Purpose:
The purpose of the training is to familiarize the trainees with the principle of working, technical
specifications of Equipment and overall strategy of Performance Qualification.
4.2 Scope:
This Training is applicable to the Autoclave (HPHV Steam Sterilizer) installed in Microbiology
Section of Quality Control department of ABC Pharma Pvt. Ltd. Bhiwadi.
4.3 Topics:
The following topics shall be covered during training:
4.3.1 Principle of working of system
4.3.2 Technical specifications of the system
4.3.3 Overall strategy of Qualification process
4.3.4 General precautions / guidelines to be followed during qualification
4.3.5 Documentation Procedure to be followed
Attach training record with the report as Annexure 01.
5.0 REQUIREMENT FOR PERFORMANCE QUALIFICATION:

5.1 Details of Documents Required Before Performance Qualification:
5.1.1 Purpose:
The purpose of this test is to record and check all the documents which should be ready
before the start of Performance Qualification.
5.1.2 Procedure:
5.1.2.1 Check that the IQ and OQ have been completed. In case of any deviation, the
said deviation should be approved and signed off.
5.1.2.2 Check that all the SOP’s related to the equipment Operation, calibration,
cleaning and maintenance is in place.
5.1.2.3 Check that the training has been imparted to all the persons responsible for
executing the qualification.
5.1.2.4 Complete the check list for Document status as per Annexure 02.
5.1.3 Acceptance Criteria:
All the documents should be in place before executing the Performance Qualification.
5.2 Details of Instruments / Indicators Used During Qualification:
5.2.1 Purpose:
The purpose of this test is to record the details of the instruments and indicators used for
the qualification study & verify their appropriateness for use w.r.t their calibration,
expiry etc as applicable.

5.2.2 Procedure:
5.2.2.1 Following instruments/ indicators shall be required for the Performance
Qualification of Autoclave (HPHV Steam Sterilizer):
5.2.2.1.1 Data Logger
5.2.2.1.2 Thermocouples
5.2.2.1.3 Bowie-Dick test pack
5.2.2.1.4 Geobacillus stearothermophilus ampoules and strips
5.2.2.1.5 Chemical OK indicators
5.2.2.2 Verify the calibration status of the data logger / thermocouples.
5.2.2.3 Record the details of the indicators (chemical / biological etc) & verify their
appropriateness for use.
5.2.2.4 Record the observations & details in the appropriate format and attach (along
with the reports) as Annexure 03.
The data logger should be calibrated as per the procedure; sensitivity of data logger shall
be better that or equal to 0.5°C. The indicators used shall be within their expiry period.
6.0 SYSTEM / EQUIPMENT DESCRIPTION:

6.1 System Equipment Details:
6.1.1 Machine Name : Autoclave (HPHV Steam Sterilizer)
6.1.2 Model No. : PSSD224AS (V)
6.1.3 Equipment ID : QC/AUT-01
6.1.4 Serial No. : 2009223
6.1.5 Manufacturer : Pharmalab
6.1.6 Chamber Size in mm : 600 (W) × 600 (H) × 1200 (D)
6.2 Generic Design of Double Door Autoclave:
The High Pressure High Vacuum Steam Sterilizer (Equipment ID.: QC/AUT-01) consists of the
following features.
The Sterilization Chamber is made up of SS sheet, which is welded with U-Profile SS Jacket.
The Sterilization Chamber is provided with two sliding doors, which are also made up of SS
reinforced with Support structure. The door is operated with the help of pneumatic cylinder.
When the door reaches the end position gaskets are pushed out automatically with the help of
compressed air for sealing. Similarly to open the door, gasket is retracted with the help of
vacuum, which is created by the compressed air ejectors. When the gasket is retracted the
sterilizer chamber door slides automatically. The door sealing is affected with the help of
tubular silicone-rubber gasket. To ensure proper sealing the gaskets are activated with
compressed air and retracted with the help of vacuum.

Door interlocks are provided to prevent simultaneous opening of both the doors and process
lock to prevent opening of the door during the operation.
The sterilization chamber is insulated with glass wool, which helps in reducing the heat loss to
the environment and ensuring uniform distribution of temperature inside the chamber. This
insulation is covered with SS plate.
A Stainless steel pipe stand support is provided for the equipment thus requiring no special
foundation. All Joints, Crevices are filled with Epoxy/Silicon sealant to prevent any leakage.
The High Pressure High Vacuum Steam Sterilizer is provided with the following systems and
accessories for the desired functioning.
6.2.1 Water-ring type vacuum pump with suitable electric motor. (SS shell and tube type
condenser for condensation of steam prior to entering the housing of the vacuum pump).
6.2.2 Sterilizing grade vacuum – break filter on the non-sterile side.
6.2.3 One number of Laminar Air Flow workstation is provided for unloading of the
sterilized articles in the aseptic area.
6.2.4 Chambers compound gauge on the sterile area side (Grade B, Buffer Room) and the
washing and sterilization area side (Grade C, Loading side).
6.2.5 Jacket pressure gauge, Gasket pressure gauge for both the doors, Safety valve for
jacket and chamber, Steam trap with strainer and NRV for chamber and steam trap for
jacket, on the washing & sterilization area side (Grade C, loading side).
6.2.6 Control panel consisting of Mitsubishi PLC, Digital Temperature input to the
microprocessor from 5 no’s of temperature transmitters (RTDs), 1no of Pressure
transmitter, pressure and vacuum switches for transmitting digital Pressure/Vacuum
signals to the microprocessor, provision of auto-override for manual operation in case of
microprocessor failure.
The Sterilization cycle can be automatically controlled by the PLC, which can be programmed
and protected for four different types of cycles (i.e. 1st Vacuum Leak test, 2nd Std. Steam
sterilization 3rd HPHV, 4th Bowie Dick cycle). The required cycle can be selected on the
control panel and the selected cycle is automatically governed by the PLC.
The High Pressure High Vacuum Steam Sterilizer (Equipment ID – QC/AUT-01) is located in
the Media Preparation Room of Microbiology Laboratory. The Media Preparation Room is
designed as per the specifications for cleanliness Grade D. The equipment is located such that, it
can be attended easily for routine operational, monitoring and maintenance purpose. The Non-
Sterile door of the sterilizer opens into the Media Preparation Room (Grade D) for loading and
the Sterile Door opens into the Cool Zone (Grade B) for unloading. The major components of
the Control panel and utility lines are located in the Media Preparation side, for better
accessibility.

7.0 QUALIFICATION PROCEDURE OR METHODOLOGY:

7.1 General Recording Instructions:
7.1.1 Read the contents of the document thoroughly before proceeding for Execution of the
activity (in case of doubts / contradictions / contact the approvers of the document for
clarifications).
7.1.2 All the details are to be verified jointly by the validation team members with assistance
of supplier’s representative as applicable.
7.1.3 Recording of all the observations and data shall be as per good documentation practices
and other policies of ABC Pharma Pvt. Ltd., Bhiwadi as applicable.
7.1.3.1 A ball point pen shall be used.
7.1.3.2 Correction fluid and other correction techniques that obscure the original entry
should not be used.
7.1.3.3 Spelling mistakes and other changes shall be corrected as follows:
Perfrmnce Performance (initials & date).
7.1.4 Recording will be done on Annexures of the approved Performance Qualification.
7.1.5 In each of the tests outlined in the protocol the test parameter, specifications, acceptance
criteria and any other requirement are pre-defined. Other columns viz. observations,
verified by and other columns are to be filled in by the member of execution team
manually.
7.1.6 All the columns in the Annexures are to be filled by the concerned persons using a
permanent ball point pen. If a particular detail is not applicable, indicate the same by
putting a ‘––’ or ‘NA’ in the column.
7.1.7 When a minimum or maximum or a range is specified; actual measured values shall be
recorded instead of “meets specification” / “complies” etc.
7.1.8 The concerned personnel shall put sign and date in the assigned space.
7.1.9 Any mistakes / errors discovered in the approved protocol, wrt its content, test
specifications etc before or during the execution are to be recorded as a comment and
not be cancelled. This shall be verified during review of executed protocol.
7.1.10 Follow the specific instructions as mentioned in the particular test for verifying the
parameters & recording the observations.
7.1.11 Recording the requested information in a particular test indicates that it has been
inspected / verified according to specifications by visual examination, product literature,
using a measuring device etc.
7.1.12 All the deviations / discrepancies observed during the activity are to be explained &
justified if acceptable.
7.1.13 Record all deviations / discrepancies observed during the execution in the Observed
deviations format.

7.2 General Safety Instruction for Execution:

Safety will be one of the key considerations during the execution of this document. The
following guidelines must be observed during the execution stage.
7.1.1 All personnel involved with the execution shall identify hazards associated with
performance of PQ testing and precautions to be taken.
7.1.2 All personnel involved with the execution shall check that utilities are safely isolated
when energizing or de-energizing.
7.1.3 All personnel involved in the execution shall inform the company management any
hazard, to themselves or others, associated with the materials, equipment, method of
working and the precautions to be taken.
7.3 General Steps and Precaution to be followed in Qualification Study:
7.1.1 Number of load pattern & Validation schedule shall be decided prior to execution the
validation activities.
7.1.2 Thermocouple probes to be inserted through the validation port into the Autoclave. ‘T
type’ thermocouples to be used.
7.1.3 The Drain / Vent sensor should be inserted such that it is as close in proximity to the
sensor located there.
7.1.4 Criteria for selecting the locations for studies shall be as per step 6.2.
7.1.5 In each defined load pattern the thermocouples are to be positioned / attached at strategic
points / locations. Locations least accessible to steam are the areas which are remote
from the steam source or have large air spaces within and outside the loaded articles.
7.1.6 The biological indicators are to be attached to the thermocouple. Determine the initial
spore population for each lot of Biological indicator used (the methodology for
determination of spore population is as per the SOP No. QC026 “Total Viable Spore
Count of Biological Indicator” of Microbiology section. Alternatively the Indicators can
be sent to an external lab for determination of Total viable spore count.
7.1.7 Attach the thermograph, data and chemical indicator strips along with the data of the
relevant cycle.
7.1.8 Record the minimum & maximum pressure inside the autoclave chamber after the
sterilizing condition of the temperature has been achieved.
7.1.9 For calculation of Fo value take measured temperature value for each probe separately.
Refer step 7.5 for details of calculations.
7.1.10 The load patterns shall be validated for combination of maximum number / largest size
of items. During normal operation lesser item of the load patterns can be used but the
position of the item should not be changed from the approved load pattern.
7.1.11 In case of testing being done in succession (wherein a cycle is preceded by another
cycle) allow the chamber to cool to at-least 65°C before starting the next cycle.

7.1.12 If during a thermometric test the thermocouples inserted for profiling purposes are
damaged / short / open / faulty; review & evaluate the total data. If the location where
the sensor was placed is a critical one requiring monitoring, repeat the cycle.
7.1.13 The minimum time for which a cycle should operate in sterilization mode shall be based
upon the validation reports / experience (this will be the time for which the temperature
at the controlling probe / location will be in the range of sterilization temperature band).
7.4 Selection Criteria for Locations of Thermocouples, Biological and Chemical Indicators for
Loaded Chamber Study:
7.4.1 At-least one part/item in the chamber that is fastest to attain the sterilization temperature.
7.4.2 At-least one part/item in the chamber that is slowest to attain the sterilization
temperature.
7.4.3 At-least one part/item in the chamber that is hottest during the sterilization hold period.
7.4.4 At-least one part/item in the chamber that is coolest during the sterilization hold period.
7.4.5 The part of the chamber that is slowest to cool to 90°C (applicable for sterilizers with
thermal door interlock and loads containing Fluids only).
7.4.6 In the active chamber discharge (Drain).
7.4.7 In the centre of the porous load.
7.4.8 Ensure that there is a temperature sensor located in the chamber free space [to be used as
a reference probe in the loaded chamber studies].
7.4.9 In tubes/containers containing fluids the temperature sensors should be placed such that
they are located approximately at the centre of the tube/container.
7.4.10 In silicon tubing the temperature sensors should be placed such that they are located
approximately at the centre of the tubing.
7.4.11 In items like large vessels / tanks, sensors should be inserted inside the item so as to
reach middle.
7.4.12 In item like housings with filter the sensors should be placed in the centre of the
cartridge.
7.4.13 If number of items in the load pattern is less than 6, keep indicators and sensors in all the
items.
7.5 Calculation of F0 Values / Sterility Assurance Level:
7.5.1 Calculate the Fo values (mathematical) at each identified location using the equation:
F0 = Σ L × dt = Σ 10 (TA-121)/10 × dt
Where – Fo = Equivalent sterilization time (in minutes) at 121°C.
L = Lethal rate
TA = Actual temperature measured.

dt = Time duration (in minutes) between- 2 successive

temperature measurements
7.5.2 Calculate the minimum required Fo value for producing a 12 log reduction of the
biological indicator used using the equation
F0 = D121 (log N0 – log Nt)
Where – Fo = Equivalent sterilization time (in minutes) at 121°C.

D121 = D Value of the biological indicator used (as given in the
manufacturers certificate).
N0 = Initial Spore Count of the biological indicator (as determined
experimentally).
Nt = Spore Count of the biological indicator at time ‘t’ (determined by
sterility testing i.e. Nt = 0 ≅ 10-6 for obtaining a 12 log reduction).
7.5.3 SALDesired (Desired sterility assurance level):
SALDesired = 10-6
7.5.4 SLRMeasurable (Spore log reduction that can be measured by the biological indicator used):
SLRMeasurable = log N0 – log SALDesired
Where – SLRMeasurable = Spore log reduction that can be measured

SALDesired = Desired sterility assurance level
N0 = Initial Spore Count of the biological indicator (as
determined experimentally)
The desired spore log reduction is ≥ 12
7.5.5 Calculate the actual spore log reduction achieved (SLRActual) using the equation:
SLRActual = F0
D121
Where – SLRActual = Actual spore log reduction
F0 = Minimum F0 value calculated mathematically
D121 = D value of the biological indicator (as in the certificate of
analysis).
7.5.6 Calculate the actual sterility assurance level (SALActual) using the equation

SAL Actual = 10 [(log N0) – SLRActual]
Where – SALActual = Actual sterility assurance level.

SLRActual = Actual spore log reduction
N0 = Spore Count of the biological indicator (as determined
experimentally)
7.6 Equipment Qualification:
The qualification of the Autoclave (QC/AUT-01) shall be done by verifying / testing the
parameters as listed in the following table –
S. No. of
Tests Remarks
No. Runs
1. Steam Quality Testing –
→ Non-condensable gas test 03 May be done simultaneously with
the Empty chamber temperature
profile
→ Superheat test 01
→ Dryness test 01
→ Chemical and 01
Microbiological tests
2. Calibration of the data logger &
temperature sensors used for To be done initially & then after
–
temperature profile / completion of PQ
thermometric tests
Vacuum leak test [Chamber
3. 03 Without probes
integrity testing]
4. Bowie – Dick type test 03 Without Probes
01 (details in
Vacuum leak test [Chamber With probes for profiling / mapping
5. the remarks
integrity testing] inserted in the chamber
column)
This test to be repeated on each day
01 (details in
of PQ
6. Bowie-Dick type test the remarks
column
Empty chamber study [heat The location of probes is to be
7. 03
distribution study] interchanged in each cycle
Loaded chamber studies – Separate for each defined load
8. distribution & penetration studies 03 pattern [porous / unwrapped
[Thermometric tests] instruments / fluid load]
To be done as a part of the
9. Automatic control test –
thermometric tests

S. No. of
Tests Remarks
No. Runs
Vacuum leak test [Chamber With the probes removed; after
10. 01
integrity testing] completion of all the PQ tests
Applicable for the thermometric
F /…./ sterility assurance level
11. 0 – tests, refer step 7.5 for details of
calculation
calculations
12. Loaded Chamber Study (with Of each load pattern except for
03
only BI in form of Paper carrier) Fluid Load
The above-mentioned tests provide a documented verification of the intended performance of
the machine throughout the representative / anticipated range & ensure that the autoclave is
performing satisfactorily & will produce desired results when operated according to SOP & also
provide evidence of appropriate and reliable performance when challenged with worst case
scenario.
The purpose of performing the tests & the methodology of performing / executing of each test
along with the provision of recording the observations is given in the following pages.
7.7 Steam Quality Test:
7.7.1 Purpose:
The purpose of this test is to demonstrate that the steam being introduced into the
chamber is of requisite physical, chemical & microbiological quality such that the
sterilization conditions are achieved and maintained during the sterilization purposes.
The same will be established by performing the following tests:
For Physical Non-condensable gas test
Quality Superheat test
Dryness test
7.7.2 Requirement:
Steam Quality test kit, Sampling bottles, tube etc.
7.7.3 Procedure:
7.7.3.1 Physical Quality Test:
This test shall be performed as per the procedure mentioned in the
Performance Protocol of Pure Steam Generation & Distribution System,
Protocol No. EPQ/ EN/ 008-00.
7.7.4.1 Physical Test:
7.7.4.1.1 Non- condensable gas test – NMT 3.5%
7.7.4.1.2 Superheat Test – NMT 25°C
7.7.4.1.3 Dryness Test – NLT 0.95 (since metal loads are also processed)

7.8 Vacuum Leak Test:

7.8.1 Purpose:
The purpose of this test is to demonstrate that the rate of vacuum drop (air leakage into
the chamber) is within the acceptable limits when the chamber is subjected to the
vacuum.
7.8.2 Requirement:
Stop watch, data loggers, thermocouples etc.
7.8.3 Procedure:
7.8.3.1 The autoclave is equipped with automatic systems that performs the steps
below automatically & calculates the leak rate (pressure rise in mbar/min)
automatically.
7.8.3.2 This test will be performed in the following conditions / situations during this
PQ study:-
7.8.3.2.1 03 run; initially, without sensors for validation study & without
chamber furniture (carriage).
7.8.3.2.2 01 run; with sensors for validation study inserted into the chamber
and without chamber furniture (carriage).
7.8.3.2.3 01 run; on each day of PQ study (to be done before start of the
activities on the day.
7.8.3.2.4 01 run; after removing the sensors inserted for the validation
study (at the end of the PQ study).
7.8.3.3 Set the vacuum leak test cycle on the controller of the machine & operate the
machine as per the SOP No. QC027 - “Operation, Cleaning, calibration and
maintenance of HPHV Steam Sterilizer”.
7.8.3.4 This is an initial test & has to be carried out without the chamber carriage & the
probes for temperature profiling.
7.8.3.5 As per the set cycle parameters when the chamber temperature has stabilized
(the chamber is empty except for fixed furniture and necessary monitoring
sensors), the vacuum leak test cycle is initiated.
7.8.3.6 When the pressure (vacuum) in the chamber is –0.85 bar or below, close all the
valves connected to the sterilizer chamber and stop the vacuum pump (step
performed automatically).
7.8.3.7 Observe and record the time (T1) and the pressure (P1).
7.8.3.8 Wait for 300 sec ± 10 sec and not more than 600 sec ± 10 sec, to allow
evaporation of condensed water in the chamber and then observe and record the
pressure (P2) in the chamber and the time (T2).

7.8.3.9 After a further 600 sec ± 10 sec, observe and record the pressure (P3) and the
time (T3).
7.8.3.10 Allow the cycle to proceed normally to completion.
7.8.3.11 Calculate the vacuum leak rate as per the equation below –
 P - P2 
LR =  3  mbar.min
-1
 10 
Where- LR = Vacuum leak rate in mbar / minute

P = Final pressure in the chamber in mbar
P2 = Pressure in the chamber after stabilization in mbar
7.8.3.12 Record the observations & data in the Annexure – 05.

7.8.3.13 Repeat the above procedure 2 more times without probes & 1 time with probes
inserted for validation study, and attach the data, along with printouts as
Annexure – 05.
7.8.3.14 Repeat the procedure on each day of the PQ study & attach the data as per
Annexure – 06.
7.8.4.1 The measured pressure [vacuum] (P2) should be – 850 mbar (-0.85 bar) or
below.
7.8.4.2 The difference between (P2 – P1) shall not be greater than 20 mbar.
7.8.4.3 The vacuum leak rate should not exceed 1.3 mbar/min.
7.9 Bowie Dick Type Test:
7.9.1 Purpose:
The purpose of this test is to demonstrate whether or not steam penetration in the test
pack is even and rapid, and thus by implicating that air or other non-condensable gases
are not present, by using a Bowie-Dick Type Test pack.
7.9.2 Requirement:
Bowie Dick Type Test pack, Data Logger, Thermocouples etc.
7.9.3 Procedure:
7.9.3.1 Perform this test using readymade test pack, at a set sterilization temperature of
121°C.
7.9.3.2 Place the test pack at a height of 100-200 mm from the drain point of the
chamber.
7.9.3.3 Select the Bowie- Dick test cycle from the PLC. Set the sterilization hold time
as per the values mentioned in the table below and start the cycle.
Sterilization Temperature in °C Holding time

121 17 minutes
7.9.3.4 When the cycle is complete, remove the indicator paper from the test pack and
observe for colour change.
7.9.3.5 Mark the details on the paper viz. the load no., Cycle no., date, etc. & attach
the same to the protocol.
7.9.3.6 In case of improper colour change rectify the machine; repeat the test as per the
procedure mentioned above. Do not use the machine till the fault has been
rectified.
7.9.3.7 Record the observations & data in the Annexure – 07.
7.9.3.8 Repeat the procedure on each day of the PQ study & attach the data as per
Annexure – 06.
7.9.4Acceptance Criteria:
7.9.4.1 Visual indication of the cycle completion should be there at the end of the
cycle.
7.9.4.2 There should be a uniform colour change over the entire pattern of the
indicator sheet.
7.10 Empty Chamber Heat Distribution Study:
7.10.1 Purpose:
The purpose of this test is to
7.10.1.1 Demonstrate that the temperature / pressure conditions required for sterilization
are achieved in all the parts of the chamber i.e. every point in the empty
chamber attains an F0 value (lethality) of ≥ 15 minutes and that the minimum
lethality provided by the sterilization cycle is sufficient to provide at least 12
Log reduction of micro organisms having a D value of 1 minute at 121 °C.
7.10.1.2 Assess the hottest & coolest locations in the chamber.
7.10.1.3 Identify the locations that are fastest & slowest to attain the sterilization
temperature.
7.10.2 Requirement:
Data logger, temperature probes, chemical indicators etc.
7.10.3 Procedure:
7.10.3.1 Insert thermocouple probes through the validation port into the autoclave.
Seal the port with silicone sealant so that steam leakage does not occur.
7.10.3.2 Label all the thermocouples properly with location no. and the indicators
with Batch No., Cycle no., Location no. etc. and then attach/ position the
thermocouples at specified locations (as per Annexure – 10) along with
chemical indicators . Ensure that the thermocouples do not touch any metallic
surface and chemical indicators are within their expiry period.

7.10.3.3
Record the locations in the format of location details. (The locations of the
indicators and thermocouples shall be as per the feasibility of positioning
them).
7.10.3.4 Operate the autoclave for HPHV Sterilization Cycle with a hold time of 15
minutes (for 121°C cycle); record the settings in the format for operational
parameters (as per Annexure – 08). Set the data logger to record
temperatures for all the channels at a frequency not exceeding 5 second
duration for 121°C cycle.
7.10.3.5 After completion of cycle, remove the chemical indicators from the autoclave
and check for the labels. If the labels have faded then properly mark the
location on the same. Observe & document whether the sensors have retained
their original positions.
7.10.3.6 Observe the Chemical indicators for the specified colour change and
document it as per Annexure – 08.
7.10.3.7 Compile the complete data generated during the qualification test for
complete evaluation of the system. Examine the collected data to identify-
7.10.3.10.1 the locations in the chamber that are fastest and slowest to attain
the sterilization temperature
7.10.3.10.2 the locations in the chamber that are hottest and coolest during
the sterilization hold time
7.10.3.10.3 the location in the chamber that is slowest to cool to 90°C
7.10.3.8 If the thermometric data is acceptable perform two additional runs for same
cycle to demonstrate cycle and sterilizer reproducibility. In the next two runs
interchange the locations of the thermocouple, so as to verify the
thermometric data of a particular location. Attach the data, along with
printouts and indicators as Annexure – 08.
7.10.3.9 Similarly, perform the Empty cycle heat distribution study for Gravity
process also and attach as per Annexure – 08.
7.10.4.1 A visual indication of the cycle completion should be there at the end of the
cycle.
7.10.4.2 At the end of the cycle, the sensors should have retained their original
positions.
7.10.4.3 The holding time is ≥ 15 minutes for 121°C cycle / ≥ 3 minutes for 134°C
cycle.
7.10.4.4 All the locations / sensors should achieve the sterilization temperature within
1 minute. (determined w.r.t the 1st location / sensor achieving the sterilization
temperature).

7.10.4.5 The temperature of an individual sensor does not fluctuate by more than ±
1°C.
7.10.4.6 The indicated and recorded chamber temperatures are within 2°C of the
temperature measured in the active chamber discharge.
7.10.4.7 The hottest & the coolest locations are identified.
7.10.4.8 The autoclave door should not open till the cycle is complete.
7.11 Loaded Chamber Study:
7.11.1 Purpose:
The purpose of this test is to-
7.11.1.1 Assess the hottest & coolest locations and identify the locations that are fastest
& slowest to attain the sterilization temperature.
7.11.1.2 Demonstrate that the temperature / pressure conditions required for sterilization
are achieved in all the parts of the chamber and the load articles i.e. every point
in the chamber and load attains an F0 value (lethality) of ≥ 15 minutes and that
the minimum lethality provided by the sterilization cycle is sufficient to
provide at-least 12 Log reduction of micro organisms having a D value of 1
minute at121 °C.
7.11.1.3 Assess the requirements of automatic control test, by verifying the cycle of
each type of load Vis-à- Vis the requirements.
7.11.2 Requirement:
Data logger, temperature probes, chemical and biological indicators etc.
7.11.3 Procedure:
7.11.3.1 The distribution & penetration studies shall be done as a combined cycle.
7.11.3.2 Ensure that there is no leakage from the validation port. Ensure that there is a
temperature sensor located in the chamber free space.
7.11.3.3 Select the appropriate cycle from the HMI and set the sterilization parameters
& hold time as per the requirements of the load pattern.
7.11.3.4 Load the autoclave & attach / position the thermocouples at points / locations
along with biological and chemical indicator strips after appropriately
marking the same. Ensure that the thermocouples are in good thermal contact
with the fluid / device which is being monitored by it.
7.11.3.5 At a minimum the thermocouples are to be placed in –
7.11.3.5.1 the active chamber discharge
7.11.3.5.2 one each in the items known to be the fastest and slowest to
attain the sterilization temperature
7.11.3.5.3 one each in the items known to be the slowest to cool to 90°C
7.11.3.5.4 if the load consists of less than six items, place a sensor in each
item

7.11.3.6 The locations shall be as per Annexure – 10. (The locations of the indicators
and thermocouples shall be as per the feasibility of positioning them in the
loaded articles).
7.11.3.7 Operate the autoclave for sterilization hold time as per the requirements;
record the settings in the format for operational parameters. Set the data
logger to record temperatures for all the channels at a frequency not
exceeding 10 second duration for 121°C cycle.
7.11.3.8 During the holding time of the cycle observe the details w.r.t the
temperatures & pressure in the chamber & the active chamber discharge –
drain.
7.11.3.9 Remove the biological indicator strips & chemical indicators from the
autoclave and properly mark the location on the same again, in case the
earlier marking has been erased. Observe & document whether the sensors
have retained their original positions, and that the items in which the sensors
were placed are intact.
7.11.3.10 Observe the Chemical indicators for the specified colour change and
document it as per Annexure – 09.
7.11.3.11 Send the biological indicators for incubation. In no case there should be a
time lag of more than 2 hrs between completion of cycle and incubation of
Biological indicators. Also, incubate one positive control along with each lot
of tested biological indicators. The tested biological indicators are to be
incubated at 55-60°C for a period of 7 days, while the positive control
indicator should be incubated for a period of 2 days. Attach the report as
Annexure – 09.
7.11.3.12 The initial spore population is also to be determined for each lot of indicator
used, as per SOP QC026 “Total Viable Spore Count of Biological Indicator”
of Microbiology section. Alternatively the Indicators can be sent to an
external lab for determination of Total viable spore count.
7.11.3.13 If the thermometric data is acceptable perform two additional runs for the
said load pattern to demonstrate reproducibility.
7.11.3.14 Compile the complete data generated during the qualification test for
complete evaluation of the system.
7.11.3.15 Examine the collected data to identify-
7.11.3.15.1 the locations in the chamber that are fastest and slowest to attain
the sterilization temperature
7.11.3.15.2 the locations in the chamber that are hottest and coolest during
the sterilization hold time
7.11.3.15.3 the location in the chamber that is slowest to cool to 90°C.

7.11.3.16 Repeat the above procedure for each defined load pattern, as per Annexure –
10. Attach the data, along with printouts and indicators as Annexure – 09.
7.11.4.1 During plateau period[determined from the recorded chamber
temperature]-
7.11.4.1.1 The indicated and recorded temperature should be in the range
121°C – 124°C.
7.11.4.1.2 The difference between indicated and recorded chamber
temperature shall not exceed 2°C.
7.11.4.2 During the holding time-
7.11.4.2.1 The holding time is ≥ 15 minutes for 121°C cycle / ≥ 3 minutes
for 134°C cycle.
7.11.4.2.2 The temperature is within 121-124°C for 121°C cycle.
7.11.4.2.3 The temperature measured in each load item does not fluctuate
more than ± 1°C, and does not differ from that in other load
items by more than 2°C.
7.11.4.2.4 The indicated and recorded chamber temperatures are within
2°C of the temperature measured in the active chamber
discharge.
7.11.4.2.5 Pressure in the chamber shall be between 1.0 – 1.5 kg/cm 2 (for
121°C cycle).
7.11.4.3 On completion of cycle-
7.11.4.3.1 A visual indication of the cycle completion should be there.
7.11.4.3.2 The temperature sensors should have retained their original
positions.
7.11.4.3.3 The items containing sensors are intact.
7.11.4.3.4 The chemical indicators show a uniform color change as
specified by the supplier at end of cycle.
7.11.4.3.5 The biological indicator exposed to sterilization cycle shows no
growth at the end of the incubation time and the unexposed
inoculated carrier shows growth within 48 hours.
7.11.4.3.6 The actual SAL (Sterility Assurance Level) calculated should be
≥ 10-6.
7.11.4.3.7 The F0 at all points shall be ≥ 15 minutes.
7.11.4.3.8 The temperature of fluids in the load should be less than 90°C
(applicable for fluid loads only)
7.11.4.3.9 The autoclave door shall not open till the temperature falls to
less than 90°C.

7.11.4.3.10 The equilibration time between the drain / reference probe and
coldest part of the load for achieving sterilization temperature
should not exceed 1 minute (applicable for Porous loads only).
8.0 ACCEPTANCE CRITERIA:

The HPHV Steam sterilizer should meet all the acceptance criteria of each specified test as per
section 7.0.
9.0 REVALIDATION CRITERIA:

Revalidation shall be carried out in case of
• Major maintenance of critical parts
• Change of cycle program
• Inclusion of new load
• Change of existing load pattern
• Scheduled validation (Frequency: 6 months)
10.0 QUALIFICATION REPORT:

Qualification report shall be prepared in summarized format separately. It shall contain the results in
compiled & evaluated form.
11.0 OBSERVED DEVIATION (IF ANY):

Allot a sequential number starting from 01 for each deviation / discrepancy observed during the
execution of the activity. Record the details of the test / parameter during which the deviation /
discrepancy was observed, along with a brief description of the deviation / discrepancy, as per
Annexure – 11.
The deviation/ discrepancy shall be addressed as per the SOP QA006 – “Deviation Control”.
12.0 ABBREVIATIONS:
14.1 HPHV : High Pressure High vacuum
14.1 GA : General Arrangement
14.1 P&ID : Piping and Installation Diagram
14.1 FAT : Factory Acceptance Test
14.1 PLC : Programmable Logic Controller
14.1 DQ : Design Qualification
14.1 MOC : Material of Construction
14.1 HMI : Human Machine Interface
14.1 PW : Purified Water

14.1 VF : Vent Filter

14.1 BI : Biological Indicator

13.0 LIST OF ANNEXURE:

Annexure Annexure Title
No.
01 Training Record
02 Document Status Report
03 Calibration Certificates of Instruments / Indicators Used For Qualification
04 Pure Steam Report
05 Report for Vacuum Leak Test
06 Report for Daily Vacuum Leak Test and Bowie – Dick Test
07 Report for Bowie – Dick Type Test
08 Report for Empty Chamber Heat Distribution Study
09 Report for Loaded Chamber Study
10 Load Pattern Diagram with Probes & BI Location
11 Observed Deviation / Discrepancies
12 Cycle printouts and Reports of Loaded chamber study with BI Strips
14.0 REFERENCE DOCUMENT (IF ANY):

14.1 HTM 2010- Part III
END OF DOCUMENT

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QUALITY CONTROL Page 1 of 26

Format No.: QA004-F05-01

1.0 APPROVAL SIGNATURE:

Engineering & Maintenance

Format No.: QA004-F05-01

Approved by / Functional area Designation Signature Date

Format No.: QA004-F05-01

Format No.: QA004-F05-01

Format No.: QA004-F05-01

The Validation team is described through the following Responsibility.

Shall prepare the Qualification Protocol as per the

Manager – QC, Manager –

Qualification Protocol shall be approved by the QA head

Shall conduct and record the qualification test as per the

Engineering & Maintenance Shall identify the instruments for Calibration

Shall review the executed protocol to check the

These personnel shall review the qualification study and

3.0 EXECUTION TEAM:

Format No.: QA004-F05-01

4.0 TRAINING RECORDS:

5.0 REQUIREMENT FOR PERFORMANCE QUALIFICATION:

Format No.: QA004-F05-01

6.0 SYSTEM / EQUIPMENT DESCRIPTION:

Format No.: QA004-F05-01

Format No.: QA004-F05-01

7.0 QUALIFICATION PROCEDURE OR METHODOLOGY:

Format No.: QA004-F05-01

7.2 General Safety Instruction for Execution:

Format No.: QA004-F05-01

Format No.: QA004-F05-01

dt = Time duration (in minutes) between- 2 successive

F0 = D121 (log N0 – log Nt)

Where – Fo = Equivalent sterilization time (in minutes) at 121°C.

SLRMeasurable = log N0 – log SALDesired

Where – SLRMeasurable = Spore log reduction that can be measured

Format No.: QA004-F05-01

SAL Actual = 10 [(log N0) – SLRActual]

Where – SALActual = Actual sterility assurance level.

Format No.: QA004-F05-01

Format No.: QA004-F05-01

7.8 Vacuum Leak Test:

Format No.: QA004-F05-01

Where- LR = Vacuum leak rate in mbar / minute

7.8.3.12 Record the observations & data in the Annexure – 05.

Sterilization Temperature in °C Holding time

Format No.: QA004-F05-01

Format No.: QA004-F05-01

Format No.: QA004-F05-01

Format No.: QA004-F05-01

Format No.: QA004-F05-01

Format No.: QA004-F05-01

8.0 ACCEPTANCE CRITERIA:

9.0 REVALIDATION CRITERIA:

10.0 QUALIFICATION REPORT:

11.0 OBSERVED DEVIATION (IF ANY):

Format No.: QA004-F05-01

14.1 VF : Vent Filter

Format No.: QA004-F05-01

13.0 LIST OF ANNEXURE:

14.0 REFERENCE DOCUMENT (IF ANY):

Format No.: QA004-F05-01

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