Sterilisation and disinfection


  • It is defined as the process by which an article, surface or medium is freed of all living microorganisms either in the vegitative or spore state.

Sterilising agents-

1. Physical agents

(a) Heat

  • It is the most reliable method of sterilisation.

Mechanism of action-

  • Killing effect of dry heat is due to protein denaturation, damaging by oxidising molecules, destroying the cell constituents and the toxic effect of elevated levels of electrolytes.
  • The lethal effect of moist heat is due to denaturation and coagulation of proteins.
  • Thermal death time is inversely proportional to the temperature of exposure
    • The sterilisation time is related to the number of organisms in the suspension, presence or absence of spores and the strain and the characteristics of the organism.
    • It does not include time taken to reach the specified temperature.
    • The presence of disinfectants and high acidic or alkaline pH hastens bacterial killing.

    Dry heat-

    1. Flaming-

    • Inoculating loop or wire, the tip of forceps and searing spatulas can be sterilised by holding them over a Bunsen flame till they become red hot.

    2. Incineration –

    • It is the excellent method for terminal sterilisation for destroying biomedical waste.
    • Plastics such as PVC can be dealt with it.

    3. Hot air oven-

    • Sterilisation is achieved by conduction.
    • A holding period of 160℃ for two hours is necessary to sterilise glassware, forceps, scissors, scalpels, all glass syringes, swabs etc.
    • At 180℃ cotton plugs may get charred.
    • At 150℃ sharp instruments, such as those used in ophthalmic surgery should be sterilised.

    Moist heat-

    Temperature below 100℃-

    1. Pasteurisation of milk-

    • The milk is heated at either 63℃ for 30 minutes (the holder method) or 72℃ for 15-20 seconds (the flash process), followed by cooling quickly to 13℃ or lower.

    2. Inspissation-

    • Media such as Lowenstien-jensen and Loefller’s serum are rendered sterile by heating at 80-85℃ for half an hour on three successive days in an inspissator.
    • Bacterial vaccines are heat inactivated in the special vaccine baths at 60℃ for one hour.
    • Serum or body fluids containing the coagulation proteins can be sterilised by heating for one hour at 56℃ in a water bath on several successive days.

    Temperature at 100℃-

    1. Boiling-

    • Vegetative bacteria are killed almost immediately at this temperature.
    • Boiling is not recommended for sterilisation of instruments used for surgical procedures.
    • Hard water should not be used for boiling.
    • Addition of 2% sodium bicarbonate in the water make it soft and make it suitable for sterilisation.

    2. Steam at atmospheric pressure-

    • It is used for culture media.
    • A Koch and Arnold steamer is usually used.

    Tyndallisation or intermittent sterilisation-

    • It is used for media containing sugars or gelatin.
    • An exposure to 100℃ for 20 minutes on three successive days is used.
    • The principal is that the first exposure kill all the spores, since they are in favourable medium, will germinate and killed on the subsequent exposure.
    • However this method may fail to kill spore of certain anaerobes and thermophiles.

    Steam under pressure-

    • The equipment used is autoclave.
    • Sterilisation by steam under pressure is carried out at the temperatures between 108℃ and 147℃.
    • Aqueous solutions are sterilised between 108℃ and 126℃.
    • Autoclaves ( steam seteriliser ) are used in healthcare setup: laboratory autoclaves, hospital dressing steriliser, instrument steriliser, rapid cooling steriliser.
    • Two types of autoclaves are available: gravity displacement type and high vacuum sterilisers.

    (b) Filtration-

    • It helps in removing bacteria from heat labile liquids such as sera and solutions of sugars and antibiotics.
    • Viruses can pass through ordinary filters.
    • Bacterial toxins can be obtained by passing cultures through filters.
    • Asbestos filters are no longer used due to its carcinogenic property.

    Types of filters-

    1. Candle filters – have been widely used for purification of water for industrial and drinking purposes.

    2. Sintered glass filters

    3. Membrane filters- are routinely used in water purification and analysis, sterilisation and sterility testing, and for the preparation of solutions of parenteral use.

    (c) Radiation-

    • Two types of radiation are used for sterilisation: non ionising and ionising.
    • Non ionising radiation- infrared and ultraviolet rays.
    • Ionising radiation- gamma rays and high energy electrons.