Sterilisation & Disinfection

 Sterilisation : A process by which an article, surface or medium is free of all living

organisms either in vegetative state or spore state.
 Disinfection: Removal or destruction of all pathogenic organisms
 Asepsis: Prevention of infection by inhibiting the growth of bacteria.
 Decontamination: The process of rendering an article or an area free of contamination
including microbial, chemical, radioactive, etc.
 Bactericidal agents: Agents that kill bacteria.
 Bacteriostatic agents: Agents that prevent multiplication of bacteria.

Uses of sterilisation: To prevent contamination by extraneous organisms, to prevent sepsis in

surgery, for safety in food and drugs

CLASSIFICATION

Physical methods Chemical methods

Sunlight, drying. Alcohols - Ethyl alcohol, isopropyl alcohol

HEAT : 1) dry heat – red hot heating, Aldehydes - Formaldehyde, glutaraldehyde
flaming, incineration, hot air oven
2) Moist heat- below 100C Pasteurisation, Dyes
water bath/ vaccine bath, inspissations Metallic salts

Steam under normal pressure at 100C: Halogens

Arnolds steam sterilizer, tyndallisation.
Boiling
Steam under pressure (Above 100C): Phenols
autoclave

Filters, radiation Surface active agents

Ultrasonic and sonic vibration Gases: ETO, formaldehyde, betapropiolactone

 • Physical methods

• Red hot : inoculating loops, wires, tip of forceps and needles are held in the flame of a
bunsen flame of a burner till they become red hot.

• Flaming: used to sterilize glass slides, scalpels & edges of test tubes by passing them
through Bunsen flame without allowing them to become red hot.

• Hot air oven:

• Most widely used method of sterilization by dry heat.

• The oven is electrically heated and fitted with a fan to ensure adequate and even
distribution of air. Also fitted with a thermostat to maintain the air at a chosen
temperature.

• Holding period – 160°C for 2 hour. Or 170°C for 1hr, or 180°C for 30 mins.

• Glassware like syringes, petridishes, flasks, pipettes and test tubes, surgical
instruments like scalpels scissors, forceps etc. liquid paraffin, dusting powder, fats,
grease, etc.

• Rubber, cotton avoided

• Incineration: Contaminated cloth, animal carcasses, pathological materials, infectious

hospital waste

MOIST HEAT

At Temp less than 100°C

 Pasteurisation- method of sterilization used to sterilise liquids such as vegetable/fruit

juices, milk etc.

2 methods – Holder method heating the liquid at 63°C for 30 min

– Flash method heating the liquid at 72°C for 20 sec

 Followed by quickly cooling to 13°C

 Vaccine bath: used to sterilise vaccines- Heat inactivation by heating at 60°C for 1 hr

 Inspissation: LJ medium, Loeffler’s serum slope can be sterilized by heating at 80–85°C

for 30 min on 3 days

At 100°C

 Boiling

 Koch & Arnold steam steriliser:

media –90 min

 Tyndalisation: sugars, gelatin

 100°C – 20 min - 3 successive days

At temperature higher than 100°C

Autoclave
o Most widely used method of sterilization by moist heat.
 o Uses steam under high pressure for sterilization

o Temperature used is 121°C for 15mins at 15 lbs pressure.

o Or 126°C with 20lbs pressure for 10mins
o Or 133°C at 30 lbs for 3 mins.
o Used to sterilize culture media, rubber materials, gowns, dressing, gloves etc.

Filtration

 Useful for sterilizing substances that get damaged by heating like sera, antibiotics,
enzymes, toxins.
 Also used for Separation of virus from bacteria

Types of filters

 Candle filters. ex: Berkfeld & Mandler: Made of kieselghur, graded porosity, sterilised
by steaming or autoclave
 Chamberland or Doulton: Made of unglazed ceramic

 Asbestos disc filters:

Seitz filter: - pad of asbestos - inserted into metal holder - discarded after use

 Sintered glass filters:

Used for air filtration, made of finely ground glass.

  Membrane filters:
o Cellulose nitrate
o Also called gradacol MEM
o To separate viruses, to know their size
o Cellulose acetate or millipore filters or oxoid MEM filters

Air filters: HEPA: High efficacy particulate air filters

Radiation

 Non-ionising radiation - Infrared and ultraviolet rays - low energy type

- Control of closed environment with UV lamp

- Bacteriological laboratory, inoculation hoods, laminar flow

 Ionising radiation - Gamma rays, X rays and cosmic rays lethal to penetrative
power

No appreciable increase in temperature – ‘cold sterilisation’

Use: Sterilising pharmaceuticals, medical, dental equipment, sterilising plastics,

syringes, swabs, catheters, oil grease

CHEMICAL METHODS

Alcohols

 Protein denaturation and lipid dissolution

 Bactericidal, fungicidal
  Not effective on spores and viruses

 Commonly used - Ethyl alcohol, isopropyl alcohol

 Concentration 60-90% in water

Aldehydes

Active against amino group in protein molecule

Formaldehyde

Used for sterilising instruments, heat-sensitive catheters, fumigation of wards, sick

rooms and laboratories

Glutaraldehyde

 Disinfection of medical equipment like cystoscopes and bronchoscope

 Effective against tubercle bacilli, fungi and viruses

Phenols

 Lister - antiseptic surgery

 Distillation of tar between 170°C and 270°C

 Damage to cell membrane, release cell contents and cell lysis

 Cresols, chlorhexidine, hexachloraphane

 Disinfection of floors, excreta, preoperative disinfection of skin

Halogens

Iodine – Skin disinfectant

 Bactericidal, sporicidal

 Iodophores more active

Chlorine – Used as hypochlorite

 Bactericidal

 Disinfect water supplies, swimming pools and used in diary industries

 Ethylene oxide

 - Kills microorganisms by altering their DNA by alkylation of amino carboxyl,

hydroxyl and sulphydryl groups in protein molecules

 - Used in sterilising packaged heat-sensitive devices

 - Respirators, heart–lung machines, dental equipment, sutures, books and

clothing

Formaldehyde

 Fumigation of theatres and rooms

 150 g of KMnO4 to 280 ml formalin for every 1000 cu ft.

 Doors sealed, opened after 48 hrs

Betapropiolactone (BPL)

 Condensation product of ketane and formaldehyde

 Hydrogen peroxide fogging

 Short cycle time, non-toxic