Steam sterilization uses high-pressure steam to kill microorganisms. It is the most effective sterilization method when heat and moisture do not damage products. An autoclave uses steam to heat items inside a pressurized chamber, reaching temperatures over 121°C. Validation of a sterilization process involves qualification of the autoclave design, installation, operation, and performance through studies measuring temperature distribution, heat penetration, and using biological indicators to confirm a sterilization log reduction factor is achieved.

1. Khushboo Pasbola
M.Pharm (QAT)
SGRRITS

2.  Steam Sterilization under pressure is the
most efficient method of sterilization
 It is the method of choice when heat and
moisture is not a problem to the product
 Steam sterilization is carried out in an
autoclave
What is steam sterilization?

3. HOW DOES AN AUTOCLAVE WORK?
 Steam enters the chamber
jacket, passes through an
operating valve and enters
the rear of the chamber
behind a baffle plate. It
flows forward and down
through the chamber and
the load, exiting at the
front bottom. A pressure
regulator maintains jacket
and chamber pressure at a
minimum of 15 psi, the
pressure required for
steam to reach 121ºC
(250ºF). Overpressure
protection is provided by a
safety valve.

4. HOW TO VALIDATE?
Validation Involves
DESIGN
QUALIFICATION
INSTALLATION
QUALIFICATION
OPERATIONAL
QUALIFICATION
PERFORMANCE
QUALIFICATION

5. CYCLE DEVELOPMENT
 There are Several Factors that affect Sterilization
cycle e.g. Nature of load, type of sterilizer, container
and closure used
 If the material to be sterilized is heat stable , then
overkill approach may be employed.
 The material which can be sterilized by overkill
approach include filters, containers, closures, hoses
and other goods.
 Some soft goods such as gowns can also be
sterilized by overkill approach.
 Generally accepted probability of survival of
microorganisms is 10-6.

6. DESIGN QUALIFICATION
 The Purpose of design qualification is to demonstrate
that the sterilizer is designed properly and to
demonstrate that it perform according to its design
criteria.
 Modern sterilizer design is the result of several year
experience of the manufacturer of the sterilizers and the
users requirement
 Usually well designed steam sterilizer are available in
market.
 The Design can be modified by the manufacturer ,if
specified by the user
 The manufacturer of sterilizer can be asked for design
qualification document of the sterilizer.

7. INSTALLATION QUALIFICATION
IQ should include
 A brief description of sterilizer and its ancillary
equipment.
 A schematic drawing of the system
 List of other associated drawings like structural and
layout drawings and electrical schematics.
 List of detailed specifications
 Attestation sheets stating that the system has been
properly installed

8.  List of the equipment which make up the sterilizer
system
 List of system instruments specifying vendor,
model number and range
 Reference to calibration of instruments
 Manuals for operation and maintenance including
computer software, If the system is computer
controlled.
 List of spare parts.
 List of utility connections (e.g. clean steam, water
for injection)

9. FO VALUE:
 The F0 value is mentioned both in the USP and in the Current
Good Manufacturing Practices (CGMPs) for large volume
parentrals (LVPs).
 Both sources indicate that the steam sterilization process must
be sufficient to produce an F0 value of at least 8 min.
 This means that the coolest location in the sterilizer loading
configuration must be exposed to an equivalent time of at
least 8 min of exposure to a temperature of at least 121°C.

10. OPERATIONAL QUALIFICATION
 OQ of sterilizer is based on critical specifications
and variables of the sterilizer.
 These variables are measured under the operating
conditions and data are recorded.
 For range values both minimum and maximum
should be checked.
 There are two important devices which play a
pivotal role in validation of sterilizer :
THERMOCOUPLE and BIOLOGICAL
INDICATORS.

11. CHARACTERISTICS OF THERMOCOUPLE
 Thermocouple should be sufficiently durable for
repeated use.
 Accuracy of thermocouple should be (+ or –) 0.5o
C
 Thermocouple should be calibrated before and after
use in a validation program at two temperatures, O
and 125o C
 Temperature recorder should be capable of printing
temperature data in 0.1oC increment

12. CHARACTERISTICS OF BIOLOGOCAL
INDICATORS
 The most commonly used biological indicator is the
spores of B. Stearothermophilus in steam
sterilization validation.
 The Main Purpose of biological indicator is to
assure that a minimum Fo Value has been
achieved in the coolest location of the autoclave
load.

13. OPERATIONAL QUALIFICATION INVOLVES
1. Heat distribution study
 Heat-distribution studies include two phases:
 heat distribution in an empty autoclave chamber
 heat distribution in a loaded autoclave chamber.
 Between 10 and 20 thermocouples should be used per
cycle.
 Thermocouples should be secured inside the chamber
according to a definite arrangement
 Teflon tape can be used to secure thermocouples
 One thermocouple each should remain in an ice bath
and high-temperature oil bath during each cycle for
reference when the temperature monitoring equipment
has the capability for electronically compensating each
temperature measurement against an internal reference.

14.  Heat-distribution studies following the initial study
may employ fewer thermocouples as the cool spot in
the chamber and in the load is identified.
 The key is to identify on a reproducible basis 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 than ±2.5°C
 Greater temperature differences may be indicative
of equipment malfunction.

15. 2.HEAT-PENETRATION STUDIES
 The success of a validated cycle depends on
determining the F0 value of the cold spot inside the
commodity located at the cool spot previously
determined from heat-distribution studies.
 Thermocouple probes are inserted within a container
and repeat cycles are run to establish the point inside
the container that is coldest most of the time. It is this
exact point that is monitored during heat-penetration
studies.
 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.

16.  The F0 value will be calculated based on the
temperature recorded by the thermocouple inside the
container at the coolest area of the load.
 Upon completion of the cycle, the F0 value will
indicate whether the cycle is adequate or if
alterations must be made.
 Following the attainment of the desired time–
temperature cycle, cycles are repeated until the user
is satisfied with the repeatability aspects of the cycle
validation process

17. 3.BIOLOGICAL CHALLENGE STUDIES
 Keep ampoules containing spores suspension of
Bacillus stearothermophilus population at various
location of the autoclave along with probes and
maintain the sterilization temperature at 15psi and
121°C during the heat penetration studies, once on
the maximum load.
 Autoclaved ampoules containing Bacillus
stearothermophilus spores suspension ampoules
should not show any colour change after five days of
incubation.

19. REFRENCES:
 P.P. SHARMA, Validation in pharmaceutical industry ,
Vandana publications pvt ltd, first edition 2007,page
no.294-301
 Robert A. Nash, Pharmaceutical Process Validation
Second Edition 1993.