LECTURE 6 Sterilization
LECTURE 6 Sterilization
LECTURE 6 Sterilization
STERILIZATION
1
Introduction
2
Methods of Sterilization
Ethylene oxide
Saturated heat autoclaves Batch sterilizers
Vaporized hydrogen peroxide
Superheated water autoclaves Continuous tunnel sterilizers Electromagnetic particulate Membranes
Hydrogen peroxide/steam
Air oven steam autoclaves
Other gases
3
Introduction … cont.
4
Introduction … cont.
5
Hardest to Kill Prions (Creutzfeldt-Jakob Disease (CJD), mad
cow disease)
Spores (C. difficile)
Mycobacteria (Tb)
Non-enveloped viruses (norovirus)
Fungi (Candida)
Vegetative bacteria (MRSA, VRE)
Enveloped viruses (HIV, HBV)
Easiest to Kill
General Facts
7
General Facts… (cont.)
Heat stable reusable medical devices that enter the blood stream or enter
normally sterile tissue should always be reprocessed using heat-based methods
of sterilization (e.g., steam autoclave or dry heat oven).
8
General Facts … (cont.)
9
STEAM STERILIZATION
Terminology
11
Terminology … cont.
12
Terminology … cont.
13
Terminology … cont.
14
Action (mechanism) of Steam
15
Advantages of Steam Sterilization
16
Disadvantages of Steam Sterilization
• Preparing, packaging, loading and operating the sterilizer. Also, drying time
• Instruments need to be free from grease and oil and not be sensitive with heat
• All items must have direct contact with steam and must be able to penetrate
packaging material
• Timing is adjusted for the differences in load and material and subject to
human error
• The steam may not be pure. Refers to the amount of solid, vapor, or liquid
contamination in the steam
17
Autoclave
Autoclave:
An apparatus for
sterilization by steam under
pressure, usually at
temperatures of 250 degrees
to 270 degrees F
18
19
Principles
1- Application of pressure:
Because it is not possible to raise the temperature of the steam above
100 °C under atmospheric conditions, pressure is employed to achieve
higher temperature (it should be recognized that the temperature, not the
pressure is destructive to the microorganism and that the application of
pressure solely for the purpose of increasing the temperature of the
system.
20
Principles … cont.
2- Application of time:
Time is another important factor in the destruction of microorganisms by
heat. Most modern autoclaves have gauges to indicate to the operator
the internal conditions of temperature and pressure and timing device to
permit the desired exposure time for the load. The usual conditions
(pressure/temperature/time), are as follows:
22
How Long to Autoclave
Instruments, wrapped 30
Utensils, wrapped 30
23
How Long to Autoclave … cont.
24
How Long to Autoclave … cont.
25
Autoclave applicable:
26
Autoclave not applicable:
27
Types of steam sterilizers
1. Gravity displacement
28
Gravity Displacement Uses
1. Metalware
2. Glassware
3. Thermoplastics
4. Linens
5. Rubber
29
Types of steam sterilizers … cont.
30
Pre-vacuum Sterilizer Uses:
Metalware
Rubber
Thermoplastics
31
Types of steam sterilizers … cont.
32
High Pressure Flash Uses
33
Types of steam sterilizers … cont.
4. Washer-sterilizer
Definition: Wash and sterilize instruments. The
water floods the chamber with cold water to
prevent proteins from setting on the instrument.
The water is then heated with steam to agitate and
clean the instruments.
34
Washer-sterilizer Uses:
35
Biological Indicator Controls/Spore
Tests
Bacillus stearothermophilus: is used strictly as biological
indicator of effective heat sterilization (steam and dry heat
sterilizers) by including filter paper strips carrying
a saturated number of spores into the autoclave cycle.
The strips are then incubated to attempt to recover viable
organism. The usual autoclave cycle of 121 °C for
15 minutes is adequate to kill Bacillus stearothermophilus
with a high margin of safety.
36
Biological Indicator Control/Spore
Tests…cont.
When Used:
Upon installation of new system
After major repairs
Routine quality assurance.
With all implants
37
Biological Indicator Controls/Spore
Tests
• Record results:
• Negative- no color change from original.
38
STERILIZATION by FILTRATION
Filtration
- It depends upon the physical removal of
microorganisms by adsorption on the
filter medium or by sieving mechanisms.
- It is used for sterilization of heat-
sensitive solutions.
- Medicinal preparations sterilized by this
method are required to undergo severe
validation and monitoring since the
effectiveness of the filtered product can
be greatly influenced by the microbial
load in the solution being filtered.
- It removes, but does not destroy
M.O.
- Commercially available filters are
produced with a variety of pore-size
specifications (e.g. Millipore filters).
40
Filtration …cont.
Most filters consist of:
1- diatomaceous earth
2- cellulose acetate or
3- nitrocellulose
41
Filtration …cont.
Factors affecting removal of M.O.:
1- Pore size of filter
2- Electrical charge of the filter
and that of the M.O.
3- pH of the solution
4- T, P, and Vacuum applied
Advantages:
1- Speed
2- Good for thermolabile
materials
3- Inexpensive
4- The complete removal of living
and dead M.O. as well as
other particulate matter from
the solution
42
Filtration …cont.
Disadvantages:
43
OTHER TECHNIQUES
Ultrasonic vibrations
and membranes. 45
PYROGENS AND PYROGEN
TESTING
PYROGEN
47
PYROGEN… cont.
Two main procedures are used for the detection of pyrogens.
48
PYROGEN… cont.
Steps:
1- Render the syringes, needles, and glasswares free form pyrogens by
heating at 250 °C for not less than 30 minutes.
2- Warm the product to be tested to 37 °C ± 2 °C
3- Inject into an ear vein of each of three rabbits 10 ml of the product per
kg of body weight.
4- Record the temperature at 30-minute intervals between 1 and 3 hours
subsequent to the injection
49
PYROGEN… cont.
Steps:
5- If no rabbit shows an individual rise in temperature of 0.5 °C or more
above its respective control temperature, the product meets the
requirements for the absence of pyrogens.
6- If any rabbit shows an individual temperature rise of 0.5 °C or more,
continue the test using five other rabbits.
50
PYROGEN… cont.
Steps:
7- If no more than three of the eight rabbits show individual rise in
temperature of 0.5 °C or more and if the sum of the eight individual
maximum temperatures rises does not exceed 3.3°, the material under
examination meets the requirements for the absence of pyrogens.
51
PYROGEN… cont.
In recent years, it has been shown that an extract from the blood cells of
the horseshoe crab (Limulus polyphemus) contains an enzyme and
protein system that coagulates in the presence of low levels of
lipopolysaccharides.
52
PYROGEN… cont.
Proclotting enzyme
Endotoxin
In the test procedure, the lysate is mixed with equal volume of the test
solution in a depyrogenated container, such as a glass tube. The tube is
then incubated undisturbed at 37 °C for a period of about 60 minutes.
The test is a pass or fail test. The end point is identified by gently inverting
the glass tube. A positive result is indicated by the formation of a solid
clot. The clot doesn’t disintegrate when the tube is inverted. A negative
result is indicated if no gel clot has been formed.
54