autoclave validation maliba by x2ky3Ch8

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 Introduction to Validation

 Stages   of qualifications

 Validation   of Autoclave

 Validation   Protocol of Autoclave

 Validation   of Dry Heat Sterilizers And Tunnel
Validation    may be defined as ” Establishing documented
evidence which provides a high degree of assurance that a
specific process     will consistently produce a product
meeting its      pre-determined specifications and quality

It   has been made mandatory by the regulatory bodies to
prove the safety efficacy, Purity & effectiveness of the drug
product, medical devices & biologics in the marketplace &
health system.
    Why Validation of Equipment?

 Equipment validation   is Vital for
Fewer  interruptions of work
Lower repair costs
Elimination of premature replacement
Less standby equipment
Identification of high maintenance cost
Reduction of variation in results
Greater confidence in the reliability of results
           Who should do Equipment
  The vendor or the user
 The user has the ultimate responsibility for the accuracy of
  the analysis results and also for equipment qualification.

   DQ should always be done by the user.

   While IQ for a small and low cost instrument is usually
    done by the user, IQ for large, complex and high cost
    instruments should be done by the vendor.

   OQ can be done by either the user or the vendor.

   PQ should always be done by the user because it is very
    application specific, and the vendor may not be familiar
    with these. As PQ should be done on a daily basis, this
    practically limits this task to the user.
   Part 1. Overview on qualification and validation

   Part 2. Qualification of HVAC and water systems

   Part 3. Cleaning validation

   Part 4. Analytical method validation

   Part 5. Computerized system validation

   Part 6. Qualification of systems and equipment

   Part 7. Non sterile product process validation
Stages of qualification

 Design qualification

 Installation qualification

     Operational qualification

            Performance qualification   Change control

                                                    Results of calibration
Defined schedule                                       maintenance,
                     Frequency based on


   Requalification         After change
                                                       Extent based on
                                                       Risk assessment

                                        Part of
                                Change control procedure
       Equipment qualification
   Equipment qualification / validation includes
    following things:
 Design   qualification (DQ)
 Installation   qualification (IQ)
 Operational qualification    (OQ)
 Performance qualification     (PQ)
      Design Qualification (DQ)
 "Design qualification (DQ) defines the
  functional and operational specifications of
  the instrument and details for the conscious
  decisions in the selection of the supplier".
 List below recommends steps that should be
  considered for inclusion in a design
  Description of the analysis problem
  Description of the intended use of the
  Description of the intended environment
Preliminary selection of the functional and
 performance specifications

Preliminary selection of the supplier

Instrument tests (if the technique is new)

Final selection of the equipment

Final selection of the supplier and equipment

Development and documentation of final
 functional and operational specifications
          Installation Qualification(IQ)
   “Installation qualification establishes that the
    instrument is received as designed and specified,
    that it is properly installed in the selected
    environment, and that this environment is suitable
    for the operation and use of the instrument.”
   The qualification involves the coordinated efforts
    of –
 The   vendor
 The   operating department
 The    project team (which provide input into the
    purchase, installation, operation and maintenance
    of the equipment).
 Operational Qualification (OQ)
 "Operational  qualification (OQ) is the process
 of demonstrating that an instrument will
 function according to its operational
 specification in the selected environment."
 The proper operation of equipment is verified
 by performing the test functions specified in
 the protocol.
 A conclusionis drawn regarding the operation
 of equipment after the test functions are
 checked and all data has been analyzed.
 Following  are the contents of equipment
 operation qualification
1.Application S.O.P’s
2.Utilization List
3.Process Description
4.Test Instrument Utilized To Conduct Test
5.Test Instrument Calibration
6.Critical Parameters
7.Test Function (List)
8.Test Function Summaries
      Performance Qualification(PQ)
 "Performance     Qualification (PQ) is the
  process of demonstrating that an instrument
  consistently performs according to a
  specification appropriate for its routine use ".
 PQ should always be performed under
  conditions that are similar to routine sample
 PQ should be performed on a daily basis or
  whenever the equipment is being used.
 In practice, PQ can mean system suitability
  testing, where critical key system performance
  characteristics are measured and compared
  with documented.
             A. Introduction
 Sterile products have several unique dosage
  form properties, such as
 Freedom from micro-organisms,
 Freedom from pyrogens,
 Freedom from particulates,
 Extremely high standards of purity and
 However, the ultimate goal in the manufacture
  of a sterile product is absolute absence of
  microbial contamination.
   Three principles are involved in the validation
    process for sterile product.
1. To build sterility into a product
2. To demonstrate to a certain maximum level of
    probability that the processing and sterilization
    methods have established sterility to all units of a
    product batch
3. To provide greater assurance and support of the
    results of the end product sterility test
    D value
 “It is time required for a 90% reduction in
  microbial population. Quantitative expression
  of rate of killing of micro organism.”
 In other words, the D value will be affected by
 The type of microorganism used as BI,
 The       formulation       components      and
 The surface on which the micro-organism is
 The temperature, gas concentration, or
  radiation dose of sterilization process.
 D value found by 2 methods,
1) Survivor curve method (log number of surviving
  organism versus time/gas concentration/radiation dose)
2) Fraction negative method

Z value
 Used exclusively in validation of heat sterilization
  process. Z value is reciprocal of slope of plot of log D
  verses T at which D value is found i.e. increase in
  temperature required to reduce D value of organism by
  90 % (1 log reduction)
F value
 Used exclusively in validation of heat sterilization
  process. It is time in min required to kill all spores in
  suspension at 121oC. Measures equivalent time
    Methods of Sterilization of Products
   Moist heat (autoclave)
   Dry heat oven or tunnel
   Ethylene oxide
   Peracetic acid
   Vapor phase hydrogen peroxide
   Chlorine dioxide
   Gamma
   Beta
   Ultraviolet
           B. Qualification and Calibration
1)    Mechanically Checking, Upgrading, and Qualifying the
      Sterilizer Unit

 The    main concern with steam sterilization is the complete
     removal of air from the chamber and replacement with
     saturated steam.

 Autoclaves      can also involve air–steam mixtures for
     Sterilizing flexible packaging systems and syringes.

 When     autoclave system is used, the unit must be installed
     properly and all operations qualified through installation
     qualification and operation qualification (IQ/OQ).
2) Selection and Calibration of Thermocouples

   Thermocouples must be durable for repeated use as
    temperature indicators in steam sterilization validation and

   Copper constantan wires coated with Teflon are a popular
    choice as thermocouple monitors.

   Accuracy of thermocouples should be 0.5°C. Temperature
    accuracy is especially important in steam sterilization

   Thermocouple accuracy is determined using National
    Bureau of Standards (NBS).
     3) Selection and Calibration of BI
Sr.    Sterilization process   Biological Indicator(BI)
1.     Autoclave               B. steriothermophillus spores
                               B. subtilis var. niger spores
                               B. subtilis, 5230 spores
                               B. coagulance spores
                               Clostridium sporogenes spores

2.     Dry heat                B. subtilis var. niger spores
                               B. subtilis, 5230 spores

3.     Ethylene Oxide          B. subtilis var. niger spores

4.     Radiation               B. pumilus spores
                               Micrococcus radiodurans
                               vegetative cells
         C. Heat-Distribution Studies
 Heat-distribution   studies include two phases:

 1) Heat distribution in an empty autoclave

 2) Heat distribution in a loaded autoclave

 The   trips where the wires are soldered should not
 make contact with the autoclave interior walls or
 any metal surface.
 Heat-distribution   studies may employ thermocouples as
  the cool spot in the chamber.
 The   principle is the location of the cool spot and the
  effect of the load size and/or configuration on the cool
  spot location.
 The   difference in temperature between the coolest spot
  and the mean chamber temperature should be not greater
       2.5°C .
  than •
 Greater   temperature differences may be indicative of
  equipment malfunction.
             D. Heat-Penetration Studies

 This   is the most critical component of the entire
 validation process.
 The    main purpose is to determine the F0 value of
 the cold spot inside the commodity.
 The    container cold spot for containers ≥100 ml is
 determined using container-mapping studies.
 Thermocouple      probes are inserted within a
 container and repeat cycles are run to establish the
 point inside the container.
 Thermocouples   will be placed both inside and
 outside the container at the cool spot location(s),
 in the steam exhaust line, and in constant-
 temperature baths outside the chamber.
 F0   value will be calculated based on the
 temperature recorded by the thermocouple inside
 the container at the coolest area of the load.
 F0   value will indicate whether the cycle is
 adequate or alterations are needed.
      Heat-Penetration Studies(Con..)
 Three   critical parameter associated with all
  wet heat sterilization Processes:
1. A minimum F value
2. A design F value
3. A sterilization process time
 Any changes in the load size, load
  configuration, or container characteristics must
  be accompanied;
 To prove that the cool spot location has not
  changed or,
 If it has, that it receives the design F0 time
  exposure from the sterilization cycle used.
                  E. Equipment Qualification
 Prior      to the initiation of process, it is important that the
    sterilizer be suitably qualified to perform its function.

 Typical       critical requirements that are considered to
    affect      the    sterilization    process     (e.g.“quality”
    requirements) are:

   Accurate temperature and pressure measurement

   Air removal to some predefined level of vacuum

 Temperature         distribution and uniformity        in    the
 The   qualification of a sterilizer should include the
 following :
1.Calibration of temperature and pressure sensors
 (traceable to national or international standard)
2.Air removal (usually measured by vacuum level
 achieved vs. defined requirement)
3.Demonstration of the sequence of operations,
4.Confirmation of alarms and interlocks
5.Precision of temperature control
6.Temperature distribution and uniformity
F. Microbiological Challenge Studies
 Microbiological   challenges studies are employed to
 provide additional necessary assurance that adequate
 lethality has been delivered to all parts of the load.

 Calibrated   BIs used as bioburden models providing
 data that can be employed to calculate Fo.

 The   microorganisms used to challenge moist heat
 sterilization cycles are G. stearothermophilus and
 Clostridium sporogenes.
 After    the sterilization cycle is complete, the
 inoculated items or spore strips are recovered
 and       subjected   to   microbiological     test
 Strips    are immersed in a suitable growth
 medium (soybean casein digest medium is
 typical) and incubated for up to seven days.
        G. Sterilizer Filter Evaluation
 Microbial   filters are employed on most parts of

 sterilizers to ensure that loads are not contaminated

 by air used to vent the chamber as it cools or dries.

 Product     loads   are   protected    from     such

 contamination by their primary containers (vials,

 bags) and many nonproduct loads are protected by

 wraps to provide a microbial barrier.
 For   filters, two issues are of concern:
  Sterility and Integrity.
 If   the load will undergo a bioburden cycle, it may
  be necessary to sterilize the filter in a separate
  phase of the cycle.
 To   ensure that filters will remain functional under
  all expected conditions, the integrity tests should
  be done following the maximum cycle time and
 Triplicate   studies are recommended.
                    A. Introduction
 Mainly      three types of dry-heat sterilization

     systems are utilized in the pharmaceutical

     industry today.

I.     Batch Sterilizer Ovens

II.    Tunnel Sterilizers

III.   Microwave Sterilizers
   PRINCIPLES OF HEAT TRANSFER                    AND
 The    dry heat process must effectively heat the
    article, and air surrounding the article, to achieve
    sterilization or depyrogenation.
 In    moist heat, the condensation of the steam
    sterilizer releases large amounts of heat energy that
    serves to heat the items in the sterilizer.
 In    dry heat processes the hot air carries
    significantly less heat energy than an equivalent
    volume of saturated steam.
Key Process Features to Control Prior to
    Validating Dry-Heat Sterilizer

Batch(Oven)                Tunnel Steriliser
Intake air system          Positive pressure to entrance

Exhaust air system         Even distribution of heat

Internal air circulation   Belt speed recorder

Exhaust HEPA filter        HEPA-filtered cooling air

Static pressure gauge      Exhaust HEPA filter

Heater current             Particulate control
   The four main mechanism through which Heat
    transfer occurs are:



          B. Batch Oven Validation
1. Air balance determination:
 In an empty oven, data are obtained on the flow
    rates of both intake and exhaust air.
 Air should be balanced so that positive pressure
    is exerted to the nonsterile side when the door is
2. Heat distribution of an empty chamber:
 Thermocouples should be situated according to
  a specific predetermined pattern.
 Repeatability of temperature attainment and
  identification of the cold spot can be achieved if
  the temperature range is •   15°C at all monitored
3. Heat-penetration studies:
   These studies should be designed to determine
    the location of the slowest heating point within a
    commodity at various locations of a test load in
    the sterilizer.
 Thermocouples             are placed in the commodities
    located in the areas likely to present the greatest
    resistance to reaching the desired temperature.
   Minimum           and     maximum    temperatures   as
    defined in the process specifications should be
4. Mechanical repeatability:
   During     all    these   studies,     mechanical
    repeatability    in   terms   of     air   velocity,
    temperature consistency, and reliability and
    sensitivity of all the oven and instrumental
    controls must be verified.
         C. Tunnel Sterilizer Validation
1. Air Balance Determination:

 In   this study items being sterilized are moving exposed to

    different air systems (e.g., heating zone and cooling zone).

   Air flow must be balanced in order to provide a gradual

    decrease in air temperature as items move along the


 In   the absence of a critical balance of air dynamics, either

    the items will not be cooled or they will be cooled too

    quickly, causing contamination of the entire tunnel area.
 2. Heat-Distribution Studies:

 Thermocouples       used in tunnel sterilizer validation
 must be sufficiently durable to withstand the extremely
 high (≥300°C) temperatures in the heating zone area of
 the tunnel.

 Heat-distribution   studies should determine where the
 cold spots are located as a function of the width of the
 belt and height of the tunnel chamber.

 Peak                                             10°C
         temperature readings should remain within •
 across the belt for at least three replicate runs.
3. Heat-Penetration Studies:
 Prior   to microbial challenge testing of the tunnel
 sterilization,     heat-penetration   studies   must     be
 completed in order to identify the coolest container in
 the entire load.
 Three   to five replicate runs for each commodity size
 and every loading configuration should be              done
 using 10 to 20 thermocouples distributed throughout
 the load.
 Careful    analysis of the temperature data after each
 run will be invaluable in the determination of the cool
4. Mechanical Repeatability:
 Tunnel     sterilizers   must    demonstrate
 mechanical repeatability in the same manner
 as batch ovens.
 Air   velocity, air particulates, temperature
 consistency and reliability of all the tunnel
 controls (heat zone temperatures, belt speed)
 must be proved during the physical validation
  D. Biological Process Validation of Dry
         Heat Sterilization Cycles
 If   the dry-heat process is claimed to produce both
  sterile and pyrogen-free commodities, validation
  studies must be done using both micro-organisms
  and microbial endotoxins.
 The    goal is to validate a heating cycle that can
  produce a 12-log reduction in the biological
  indicator population.
 The    most widely used biological indicators for
  dry heat have been spores of B. Subtilis.
   Procedures for the validation of a tunnel sterilization:

 The    overkill approach is selected for the validation

 Select     the type of biological indicator to be used.

 Run     a complete cycle using the desired loading

 Determine      the number of survivors by plate-counting
    or fraction negative Methods.

 Determine      the number of spore log reductions (SLRs)
  E. Endotoxin challenge in Dry Heat
 Inoculate      commodity samples with a known
 amount of endotoxin. (e.g., 10–100 ng Escherichia
 coli lipopolysaccharide)
 Thermocouples       should be placed in commodities
 adjacent     to     those   containing   endotoxin   for
 temperature monitoring and correlation with LAL
 test results.
 Endotoxin      destruction should be ascertained at the
 coolest location of the load.
 Several   endotoxin challenge samples should be
 done per cycle, and the studies must be adequately
 Following    the dry-heat cycle, aseptically transfer
 the units containing endotoxin to an aseptic area
 for extraction procedures.
F   values required for endotoxin destruction at
 various       temperatures   and/or    cycle    time–
 temperature variations can be determined using a
 Z value of 54°C.
Equipment   required to conduct the IQ, OQ and
 PQ are discussed here.
All   temperature equipment employed to
 perform the validation studies must be
 traceable and calibrated to the International
 Temperature Scale
   The equipments used for validation testing of dry heat
    processes are discussed here:
 Resistance Temperature    Detectors
 Thermocouples

 Data   Loggers
 Wireless Temperature     Logger
 Infrared Thermometer

 Constant Temperature     Baths
 Stopwatch

 Voltmeter   or Ammeter
 Optical Tachometer
 The   IQ is designed to compare the system against the
 manufacturer’s specifications for proper installation.

 All   equipment, utilities, and connections must be
 checked          against      the       manufacturer’s

A. Structural:

 Check   dimensions, presence of identification plates,
 correct leveling, proper insulation, presence of seals,
 and inspect for structural damage.
B. Filters:
 All filters used within the system must be
  recorded, such as those used with air (supply, re-
  circulating) or in other utilities (e.g., steam,
 Some HEPA filters may need to be checked
  periodically by performing an integrity test or
C. Electrical:
 Ensure conformance to National Electrical Code
 Ensure the system provides the RH, temperature,
  and pressure differential required.
E. Air Supply:
 Identify source (direct from the HVAC system
  or room air), duct size, duct material of
  construction, and air classification.
F. Ventilation:
 Check that the ventilation exhaust duct
  exhausts to an appropriate area (not to an
  aseptic environment), and identify the method
  used to prevent back-flow.
G. Door Gaskets:
 Check integrity of gaskets and materials of
H. Heaters:
 Record the manufacturer’s model number, the
  number of heating elements, and the voltage,
  amperage, and wattage of the elements for the
I. Lubricants:
 Make certain that any lubricants used cannot
  contaminate the material being sterilized or
J. Blowers:
 The blower must be mechanically sound, the
  volute in place and correctly balanced, and that
  the blades rotate in the correct direction.
A. Temperature Monitors:
 The temperature controllers, recorders, and
  sensors on the process equipment must be
  calibrated before the unit can be operated
B. Cycle Timer:
 The accuracy of the timer must be determined,
  so that assurance is provided for cycle length.
C. Door Interlocks:
 If a unit is equipped with double doors, the
  interlocks must operate such that the door
  leading to the aseptic area cannot be opened.
D. Heaters:
 All of the heating elements must be functional. It
  is preferable to have them monitored
  continuously with ammeters in order that burned-
  out elements can be immediately detected.
E. Cooling Coils:
 To enable a faster cool-down cycle, the air is
  often circulated across coolant coils.
F. Belts:
 The belt speed is a critical operating parameter in
  both continuous hot-air tunnels and flame
 Recorders for charting the belt speed are
  recommended for units with adjustable speed
G. Particulate Counts:
 Particulate   counts should be checked within
 the containers before and after sterilization to
 quantitate the particle load.
H. Chamber Leaks:
 The   perimeter of the doors for batch sterilizers
 should be checked for air leakage while
 Upon    completion of IQ and OQ efforts and
 approval of the protocol, testing may begin.
 The   testing will include empty-chamber testing
   Heat distribution studies,
   Loaded-chamber testing consisting of heat
    distribution and heat penetration studies.
1) Component Mapping Studies
 Before conducting the loaded-chamber heat
  penetration studies, component mapping
  should be conducted.
 The studies help to determine the coolest point
  within a specific load and item.
2) Empty-Chamber Testing
 The initial testing is performed on an empty
  oven or tunnel to establish the uniformity of
  temperature distribution.
 The thermodynamic characteristics of the
  empty unit are depicted in a temperature
  distribution profile.
 3) Loaded-Chamber Studies

 For   validation purposes, the loads tested must be
  representative of standard items and quantities.

 Ideally,   each size and type of material should be
  tested by penetration studies.

 For   ovens, the time and temperature set points
  should be reduced. For tunnels, the temperature set
  point should be reduced and the belt speed
  increased if possible.
4) Bio-Challenge/Pyro-Challenge Studies
   The challenge should demonstrate the lethality
    delivered by the cycle with either microorganisms
    or endotoxin.
   The challenge can be accomplished using
    commercial strips or suspensions of B. subtilis
    spores for sterilization or E. Coli endotoxin for
   The concentration of the challenge for overkill
    processes must demonstrate adequate sterility
 After  the empty and loaded-chamber studies and bio-
  challenge studies have been completed, the data must
  be analyzed to ascertain that all testing requirements
  have been achieved.
 The results of the biochallenge studies and F value
  computation must demonstrate the required degree of
  lethality according to the protocol.
 The following information should be provided in the
  process qualification validation report:
1. Protocol achievement
2. Summary of data
3. Deviations
4. Diagram
 Berry   I.R., and Nash R.A., ”Pharmaceutical Process
 validation” second edition, revised and expanded;
 Marcel Dekker series; 83-110.
 Agalloco      J.A,   Carleton   F.A,   ”Validation   of
 Pharmaceutical Process, Third Edition, 175,223.

 Wood,     R.T; Journal of Parental drug association;
 volume 34; 286-294
 Groves,    M.J.; Journal of Parental Technology; 2nd
 edition; 432
Topic is open for Discussion


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