Sterilization is the freeing of an article from all living
organisms, including bacteria and their spores. Sterilization of culture media,
containers and instruments is essential in microbiological work for isolation
and maintenance of microbes. In surgery and medicine, the sterilization of
instruments, drugs and other supplies is important for the prevention of
infection.
Sterilization can be effected in a variety of ways, which can be
conveniently categorized as follows:
1.
Physical methods
a.
Heat
Dry heat
Mechanism of killing
by dry heat:
Dry heat kills the
organisms by destructive oxidation of essential cell constituents
Killing of the most
resistant spores by dry heat requires a temperature of about 160 °C for 60
minutes
Dry heat is employed
for glassware; syringes, metal instruments and paper wrapped goods, which are
not spoiled by high temperatures
l It is also used for anhydrous fats, oils and powders that are
impermeable to moisture
METHODS OF
STERILIZATION BY DRY HEAT:
1. RED HEAT
Inoculating wires, points of forceps and searing spatulas are sterilized
by holding them in the flame of Bunsen burner until they are seen to be
red-hot.
2. FLAMING
This method is used for sterilizing scalpel, mouth of culture tubes,
glass slides etc.
It involves passing of an article through Bunsen flame without allowing
it to become red-hot.
3. HOT AIR OVEN
This is the main means of sterilization by dry heat. Exposure at a
temperature of 160 °C for 1 hour is generally employed.
4. INFRARED RADIATIONS
Source employed is an electrically heated element, the infra red
rays are directed on to the object to be sterilized and temperature of 180 °C
can be obtained.
Moist heat
Mechanism of killing
by moist heat:
Moist heat kills the organisms by coagulating and denaturing their
enzymes and structural protein
Sterilization by moist
heat of the most resistant spores generally requires 121 °C for 15-30 minutes
Moist heat is used for
the sterilization of culture media, and all other materials through which steam
can penetrate
Moist heat is more
effective than dry heat
Sterilization can be
done at lower temperatures in a given time at a shorter duration at the same
temperature
METHODS OF
STERILIZATION BY MOIST HEAT
Moist heat can be employed at
1. Temperature below 100 °C
Examples : pasteurization
of milk
In Pasteurization of
milk the temperature employed is either 63 °C for 30 minutes or 72 °C for 20
seconds. All nonspore-forming pathogens in milk like Salmonellae,
M.tuberculosis are killed.
2. Temperature of 100
°C
3. Temperature above 100 °C
Examples : Sterilization
in an autoclave
Autoclaving is the
most reliable method
It is the method most widely used for sterilization of culture media
and surgical supplies
When water is boiled
within a closed vessel at an increased pressure, the temperature at which it
boils and the steam it forms will rise above 100 °C
This principle is used
in the autoclave
Normally autoclaving
is done at 15 lbs. (pounds per sq. Inch pressure) and 115 °C for 15 minutes
b.
Radiations
Ultraviolet radiations
Ionizing radiations
c.
Filtration
When fluids are passed
through bacteria stopping filters, they are made free from bacteria. l It is
useful for making preparations of soluble products of bacterial growth such as
toxins. Liquids that would be damaged by heat such as serum and antibiotic
solutions can be sterilized by filtration. Efficient filters should be able to
retain Serratia marcescens.
2.
Chemical methods
FACTORS INFLUENZING
STERILIZATION BY HEAT
a. The temperature and time: they are inversely related, shorter time
is sufficient at high temperatures.
b. Number of microorganisms and spores: The number of survivors diminished
exponentially with the duration of heating
c. Depends on the species, strains and spore forming ability of the
microbes.
d. Thermal death point is the lowest temperature to give complete killing
in aqueous suspension within 10 minutes
e. Depends on the nature of material: a high content of organic substances
generally tends to protect spores and vegetative organisms against heat.
f. Presence of organic or inorganic disinfectants facilitates killing
by heat
g. pH also plays an important role in the killing of microorganisms
TYPES OF FILTERS
There are different kinds of filters:
1. Earthenware candles - called Berkfield & Chamberland
filters
Berkfield Filters
Made from Kieselguhr, a fossil diatomaceous earth
Three grades of porosity are available:
a. Veil - coarsest one
b. N - normal one
c. W- wenig the finest one
Chamberland Filters
Made from unglazed porcelain
Four grades are available:
a. L1- clarifying filters
b. L1a-Big
c. L2 - normal
d. L3- Finest
2. Asbestos and asbestos-paper discs filters - called Seitz
filters
Made up of asbestos pads
Three grades are available:
a.
K- clarifying filters
b.
Normal
c.
Special EK bacteria stopping filters
3. Sintered glass filters
Made from sintered glass
Different grades available:
a.
Grades 1 to 5
b.
Grades 1-2 are for clarifying purpose
c.
Grades3-5 is for sterilization purpose
3.
Cellulose membrane filters
Made up of nitro-cellulose membranes. Made with
different grades of porosity by adjusting the concentration of constituents
4.
Fibre glass filters.
MERITS AND DEMERITS OF HEAT STERILIZATION
Advantages of heat sterilization:
1. Sterilization is very effective
2. Instruments are standardized to deliver the required effective
heat
3. Heat deliver system can be monitored effectively with various
controls like pressure gauge, temperature meters etc
4. Established quality control methods available
Disadvantages of heat sterilization:
1. Steam impermeable materials like fats, oils and powders can not
be sterilized by autoclaving.
2. Heat sensitive materials can not be sterilized by heat
Examples:
1. Serum can not be sterilized
2. Antibiotics
3. Plastic materials
4. Vaccines
5. Rubbers
3. Presence of organic matters interfere with effective
sterilization
4. Dangers of explosion when high pressure is used
