Sterilization optimization involves improving the process of sterilization to ensure it is efficient, effective, and safe123. Sterilization itself is a process that destroys or eliminates all forms of microbial life and is carried out in healthcare facilities by physical or chemical methods.

1. Optimizing The
Sterilization Process
Tutor: Eamonn Hoxey

2. How can I ask questions?
Questions & Answers

3. Optimizing The Sterilization Process

4. Sterilization
• Sterilization processes need to be properly controlled by the
management of the responsible organization
• Procedures for validation and routine control of the sterilization
process and the sterile barrier system should be within the
organization's QMS
• Compliance with ISO 13485 provides a basis for exercising that
control

5. • EN 556 - Requirements for designating sterile medical devices
• EN ISO 11135 - Ethylene oxide
• EN ISO 11137 - Radiation
• EN ISO 14937 - General criteria
• EN ISO 17665 - Moist Heat
• EN ISO 20857 - Dry Heat
• EN ISO 14160 - Liquid chemical sterilizing agents
• ISO 22441 - Low temperature vaporized hydrogen peroxide
• EN ISO 25424 - Low Temperature Steam Formaldehyde
Standards focus

6. (102 - 10-6 = 8D Reduction)
Sterility Assurance
2
0
-2
-4
-6
Probability of there
being a survivor on an
individual unit
Number of survivors
on each unit
Time
10
10
10
10
10

7. Countable
number of
organisms
surviving on
each unit
Sterilizing treatment
106
True
bioburden
remaining
on
each
unit
Probability
of
a
surviving
organism
remaining
on
any
individual
unit
102
10-2
10-4
10-0
10-6
104
Bioburden estimate
EN ISO 11737-1
Test of
Sterility
EN ISO
11737-2
1 or more
organisms
surviving on some
units in sample of
practical size (up
to 100 units)
Impractical to
sample for
surviving
organisms
Impossible to sample
for surviving organisms
Sterility Assurance

8. Nordic Pharmacopeia Addendum 1970
“Sterile drugs must be
prepared and sterilised
under conditions that aim at
such a result that in one
million units there will be no
more than one living
microorganism”.
“The preparation procedure
shall be controlled regularly
in order to ensure a low
content of microorganisms
prior to sterilization.”

9. 4.1 For a terminally-sterilized medical device to be
designated "STERILE", the theoretical probability of there
being a viable micro-organism present on/in the device
shall be equal to or less than 1x10-6.
EN 556-1: Requirements for terminally
sterilized medical devices.

10. EN 556-1: Requirements
4.2: Compliance shall be shown by the manufacturer or
supplier through provision of documentation and records
which demonstrate that the devices have been subjected to a
validated sterilization process fulfilling 4.1

11. EN 556-1: Requirements contd
NOTE 1: Evidence that a medical device is sterile comes from: i) the
initial validation of the sterilization process and subsequent
revalidations that demonstrate the acceptability of the process,
and ii) information gathered during the routine control and
monitoring which demonstrates that the validated process has
been delivered in practice.
NOTE 2: The achievement of sterility is predicted from the
bioburden level on products, the resistance of the
microorganisms comprising the bioburden and the extent of
treatment imposed during sterilization

12. Format of sterilization standards
• Common format
• based on ISO 14937:2000
• Sterilization of health care products - General requirements for characterization of a
sterilizing agent and the development, validation and routine control of a sterilization
process for medical devices.
• ISO 14937 revision published 2009
• Consistent definitions
• based on ISO TS 11139 - Terminology
• Common Quality Management System Elements
• based on reference to ISO 13485

13.  Introduction
 Scope
 Normative references
 Definitions
 Quality management system elements
 Sterilizing agent characterization
 Process / equipment characterization
 Product definition
 Process definition
 Validation
 Routine control and monitoring
 Product release from sterilization
 Maintaining process effectiveness
High level structure of sterilization standards

14. Process definition
The purpose of this activity is to obtain a detailed specification
for the sterilization process to be applied to defined product
without compromising the safety, quality and performance of
that product.

15. Process Definition
- development activities
- mechanical and biological safety
- process residuals
- product compatibility
- limits on resterilization
Process Definition

16. General approaches to process definition
1. Develop a specific process for a particular product
2. Demonstrate effectiveness of process already in use
For each of these approaches, the microbicidal performance can be
determined using:
– Natural bioburden
– Reference microorganisms
– Conservative approach
• 12 D
• Half cycle
• Reference process

17. Process optimization
• Opportunity to tailor the sterilization process to
– Reduce undesirable effects of the sterilization process
• Physical or chemical deterioration
• Residuals
– Reduce processing time
– Reduce costs

18. Sterilization - Process Definition
Sterilization processes therefore must achieve a maximal SAL of 10-6
Seen that achievement of a maximal SAL of 10-6 by a process is dependant
on the numbers and type of organisms present
Approaches to demonstrating a maximal SAL of 10-6
• Conservative or overkill approach
• Reference process
• Use of a reference organism and a knowledge of the bioburden
• Inactivation of the bioburden in its natural state

19. Conservative approach
Based on inactivation kinetics of a
highly resistant spore present in
high numbers
Sterilizing treatment
106
102
10-2
10-4
10-0
10-6
104
Product bioburden
Product bioburden
inactivation kinetics
Sterilization - Process Definition
Combined bioburden and biological
indicator method.
Based on inactivation of an
organism of know resistance
greater than the natural bioburden
and the numbers of organisms on
the product
SAL 10-6

20. Biological indicators

21. Control of Sterilization
Control of sterilization processes requires integration of process
variables.
A biological indicator integrates process variables
Variables
Process
Dose received
Irradiation
Time, Temperature
Dry heat
Time, Temperature, Moisture
Moist heat
Time, Temperature, EO concentration, Humidity
Ethylene Oxide

22. • Physical/chemical monitoring – mandatory
• Chemical indicators – not mandatory
• Biological indicators – not mandatory
Control of Sterilization

23. Biological Indicators (BIs)
Definition
A test system containing viable
microorganisms providing a specified
resistance to a specified sterilization
process
ISO 11139 3.29

24. • Selected microorganisms with known, usually high,
resistance to specific sterilization processes
• Prepared on a carrier, e.g. paper, sand, cotton yarn or metal
coupon = inoculated carrier
• Inoculated carrier in primary packaging = biological indicator
(BI)
• Ready for use and with a defined resistance to specified
sterilization process
Description of a Biological Indicator

25. Examples of process development
approaches
• Conservative approach – Ethylene oxide
• Reference process – Moist heat
• Reference organism and knowledge of bioburden –
Ethylene oxide
• Natural bioburden – Ionizing radiation

26. Ethylene oxide – Process definition

27. Annex A - D value determination
• Survivor curve construction
• Fraction negative (Most Probable Numbers)
• All BIs at each time increment must be exposed to same conditions
Annex B - Conservative process definition - (Half-cycle)
• All BIs at each time increment must be exposed to same conditions
• Inactivation of 12 log cycles
‒ Demonstration of 6 ‘spore log reduction’ from 106 to 100 –then double
exposure time
‒ Demonstration of 8 ‘spore log reduction’ from 106 to 10-2 -then add 50%
to exposure time
• Complete inactivation of 106 biological indicator – then double exposure time
Ethylene Oxide Process Definition -
Microbiological PQ

28. Half cycle approach
106
100
10-2
10-6
x x

29. Demonstration of 12 log cycles of reduction
106
100
10-2
10-6
x x

30. Demonstration of 12 log cycles of reduction
106
100
10-2
10-6
x x/2

31. Conservative approach
Based on inactivation kinetics of a
highly resistant spore present in high
numbers
Sterilizing treatment
106
102
10-2
10-4
10-0
10-6
104
Product bioburden
Product bioburden
inactivation kinetics
Ethylene oxide Sterilization – Combined BI and
bioburden process definition
Combined bioburden and biological
indicator method.
Based on inactivation of an organism
of know resistance greater than the
natural bioburden and the numbers of
organisms on the product
SAL 10-6

32. Moist heat sterilization – Reference processes

33. Moist heat reference processes
Perkins (1956)
Temperature Time
1320C 2 min
1250C 8 min
1210C 12 min
1180C 18 min
1160C 30 min
MRC (1959)
Temperature Time
1340C 3 min
1260C 10 min
1210C 15 min
Added additional (safety) period to allow to for
Deviations in steam quality

34. Moist heat - Sterility assurance level attained by different “accepted”
cycles
34
10-6
106
100
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23
G stearothermophilus
spores D121 1.5min
Natural bioburden D121
often a few seconds
SAL v’s BI with a
count of 106
SAL at F0 = 8
SAL at 1210
for 15 mins
10-12
Cycle time in
mins at 1210

35. Radiation Sterilization - Dose setting methods

36. Dose Setting Methods – Principles ..cont..
 The underlying concept for establishing a ‘product specific
dose’ is the use of naturally contaminated product items and a
measurement of the radiation resistance of micro-organisms
as they occur on the product.

37. Dose Setting Methods – Product specific
 ISO 11137
– Method 1
– Method 2
 Use of Method 1 requires substantially less resource than that
required for Method 2. Most manufacturers prefer to use
Method 1 initially

38. Dose Setting Methods
 Method 1:
– Dose setting using bioburden information
 Method 2:
– Dose setting using fraction positive information from
incremental dosing to determine extrapolation factor –
measure of radiation resistance

39. Dose Setting Methods - Principles
• The extent of treatment required for a sterilization process (T) is a
function of:
– the number of micro-organisms present prior to processing (N)
– the distribution of resistances of these micro-organisms (R)
– the required Sterility Assurance Level (SAL)
• Or
T= f(N, R, SAL)

40. Radiation Sterilization Dose Setting
Method 1

41. Dose Setting Method 1
The choice of the sterilizing dose is based on experimental
verification that the resistance to radiation of the bioburden on
the device is less than that of a microbial population having a
standard distribution of resistances

42. Dose Setting Method 1 – ISO 11137-2

43. Dose Setting Method 1 – ISO 11137-2
 Using the Standard Distribution of Resistances (SDR), solutions
to the equation T= f(N, R, SAL) have been generated for a
range of levels of bioburden and SAL values.
 These are tabulated in ISO 11137-2

44. Radiation Sterilization Dose Setting -
Method 2

45. Dose Setting Method 2
 Requires considerably more experimentation than Method 1
 Provides a measurement of the radiation resistance of micro-
organisms as they occur on product units.

46. Dose Setting Method 2
 Two experiments performed to establish estimates of:
− the radiation dose giving an SAL of 10-2;
– the D10 value of micro-organisms surviving this dose
– Based on these data, the dose for an SAL of 10-2 can be
extrapolated to that required for an SAL of 10-6

47. Dose Setting Method 2
• Two experiments
– Incremental Dose Experiment;
– 20 items exposed to each of 9 incremental doses. Items are
tested individually for sterility. Data used to estimate 10-2
SAL dose and resistance of organisms surviving that dose.
– Verification Dose Experiment;
– 100 items irradiated at 10-2 SAL dose to confirm dose at
which 1 in 100 items non-sterile.

48. Log viable
microorganisms
Probability of
one surviving
microorganism
or Sterility
Assurance
Level (SAL)
6
5
4
3
2
1
0
-1
-2
-3
-4
-5
-6
10-1
10-2
10-3
10-4
10-5
10-6
Dose kGy
Incremental Dose experiment

49. Log viable
microorganisms
Probability of
one surviving
microorganism
or Sterility
Assurance
Level (SAL)
6
5
4
3
2
1
0
-1
-2
-3
-4
-5
-6
10-1
10-2
10-3
10-4
10-5
10-6
Dose kGy
Sterilization dose
Verification Dose experiment

50. Dose Setting Method 2
• Calculation
Sterilizing Dose is calculated by extrapolation to the
required SAL from the verified 10-2 SAL dose using the
estimate of the resistance of the micro-organisms
surviving that dose.

51. Radiation Sterilization - Dose Substantiation

52. Dose substantiation - Principles
T= f(N, R, SAL)
• In the case of substantiation of 25kGy:
– T = 25kGy
– N is determined in bioburden studies
– SAL = 10-6
• Solve the equation for the maximum resistance that can be tolerated to achieve SAL =
10-6 with 25 kGy
25kGy= f(Bioburden, R, 10-6)
or
R=f(Bioburden, 25kGY, 10-6)
• Verify that this resistance is not exceeded

53. ISO TS 13004: 2013 - ISO 13004:2022
• Sterilization of health care products – Radiation –
Substantiation of a selected sterilization dose: Method VDmax
SD
• Covers substantiation of sterilization doses from 17.5 to 35
kGy at 2.5 kGy increments
• Adopted as a European Technical Specification
• Not a harmonized European standard
• Intended to gain experience before considering for
inclusion in a future revision of ISO 11137-2

54. Process Optimization - Conclusion
• Different approaches to process definition provide different degrees
of process optimization
– Radiation Method 2
– Radiation Method 1
– Radiation Dose Substantiation (Vdmax
SD)
– Ethylene oxide combined BI/bioburden approach
– Moist heat reference process
– Ethylene oxide 12D approach
– Ethylene oxide half-cycle
Increasing
optimization

56. Ethylene Oxide Sterilization
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