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Sterilization

Sterilization is the freeing of an article from all living organ-isms, including bacteria and their spores.

STERILIZATION

Definition

Sterilization is the freeing of an article from all living organ-isms, 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 infec-tion.

Sterilization can be effected in a variety of ways, which can be conveniently categorized as follows:


I. PHYSICAL METHODS

1.           Heat :

 

1.           Dry heat

 

2.           Moist heat

B.         Radiations

 

1.           Ultraviolet radiations

 

2.           Ionizing radiations

 

C.   Filtration


II. CHEMICAL METHODS

STERILIZATION BY HEAT

Heat can be applied in two forms.

1.  The dry heat

2       Moist heat.

Mechanism of killing by dry heat

l          Dry heat kills the organisms by destructive oxidation of essen-tial cell constituents

 

l          Killing of the most resistant spores by dry heat requires a tem-perature of about 160C for 60 minutes

 

l          Dry heat is employed for glassware; syringes, metal instru-ments 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.

 

Mechanism of killing by moist heat

l          Moist heat kills the organisms by coagulating and denaturing their enzymes and structural protein.

 

l          Sterilization by moist heat of the most resistant spores gener-ally requires 121C for 15-30 minutes.

 

l          Moist heat is used for the sterilization of culture media, and all other materials through which steam can penetrate

 

l          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.

FACTORS INFLUENZING STERILIZATION BY HEAT

1.     The temperature and time: they are inversely related, shorter time is sufficient at high temperatures.

 

2.     Number of microorganisms and spores: The number of survi-vors diminished exponentially with the duration of heating

 

3.     Depends on the species, strains and spore forming ability of the microbes.

 

4.     Thermal death point is the lowest temperature to give com-plete killing in aqueous suspension within 10 minutes

 

5.     Depends on the nature of material: a high content of organic substances generally tends to protect spores and vegetative organisms against heat.

 

6.     Presenceganicdisinfectantsoforganic orfacilitatesinor kill-ing by heat

 

7.     pH also plays an important role in the killing of microorgan-isms

 

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 with-out 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 160C° 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 tempera-ture of 180C° can be obtained.

 

METHODS OF STERILIZATION BY MOIST HEAT Moist heat can be employed at

1.   CTemperature below 100

 

2.     TemperatureC of  100

 

3.   Temperature above 100C

MOIST HEAT BELOW 100 °C

EXAMPLES

1.    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.

 

MOIST HEAT ABOVE 100°C

 

1.    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

 

STERILIZATION BY 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 bac-terial growth such as toxins

 

l          Liquids that would be damaged by heat such as serum and antibiotic solutions can be sterilized by filtration

 

l          Efficient filters should be able to retain Serratia marcescens

 

TYPES OF FILTERS

 

There are different kinds of filter

 

1.     Earthenware candles - called Berkfield & Chamberland filters

 

2.     Asbestos and asbestos-paper discs filters - called Seitz fil-ters

 

3.     Sintered glass filters

 

4.     Cellulose membrane filters

 

5.   Fibre glass 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

l          Made from unglazed porcelain

 

l          Four grades are available

 

a.           L1- clarifying filters

 

b.           L1a-Big

 

c.            L2 - normal

 

d.           L3- Finest

 

Seitz filter

l          Made up of asbestos pads

 

l          Three grades are available

 

a.           K- clarifying filters

 

b.           Normal

 

c.            Special EK bacteria stopping filters

 

Sintered glass filters

l          Made from sintered glass

 

l          Different grades available

 

Grades 1 to 5

Grades 1-2 are for clarifying purpose

Grades3-5 is for sterilization purpose

 

Membrane filters

l          Made up of  nitro-cellulose membranes

 

l          Made with different grades of porosity by adjusting the con-centration of constituents

 

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 variouscontrols like pressure gauge, temperature meters etc

 

4.     Established quality control methods available

 

Disadvantages

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

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