STERILIZATION AND DISINFECTION

Important in:

1. Laboratory work with pure cultures requires the use of apparatus and culture
media that is sterile

2. Prevention of infection in patients requires the use of equipment,

instruments, dressings and parenteral drugs that are free from living micro-
organisms

General definitions:

Sterilization is the process of killing all of the microorganisms in all its forms in a
preparation/article such as lab media, surgical instruments and equipment. A sterile
environment is free of life of every kind.

Disinfection is the process of destroying pathogenic organisms on the surface, but

does not involve the tissues. It is a process which reduces the number of
contaminating micro-organisms, to a level which is no longer harmful to health
.This usually involves the application of chemicals. Handscrubbing prior to surgery is
an example.

Antisepsis is used to describe disinfection applied to living tissue such as a wound.

Septic is the presence of pathogenic organisms in living tissues, such as an infected

surgical site.

Aseptic is the absence of pathogenic microbes.

Bactericidal agents effectively kill bacteria, and no growth is seen after the removal
of the agent.

Bacteriostatic agents inhibit bacterial multiplication; growth is seen after the agent
is removed.

Pasteurization is the rapid heating and cooling of milk and other liquid products to
kill pathogens, such as Mycobacterium, Brucella and Listeria.

Tyndallisation is intermittent heating.

Mode of action of sterilization and disinfection:

Damage of DNA

Protein denaturation
 Disruption of cell membrane or wall

Chemical antagonism

Removal of free sulfa hydral group

Sterlization in practice, has a probability of failure. An article may be

regarded as sterile if it can be demonstrated that there is a probability of less than 1
in a million of there being viable micro-organisms on it.

Physical and chemical sterilization may be used.

5 main methods for sterilization:

1. Heat

2. Ionizing radiation

3. Filtration

4. Sterilant gases

5. Sterilant liquids

HEAT

Moist heat is more effective than dry heat because it kills micro-organisms by
coagulatin and denaturing their enzymes and structural proteins, a process in which
water participates. It is therefore necessary for all parts of the load to be in contact
with water molecules or steam. Sterilization requires moist heat at 121C for 15
minutes, usually using an autoclave. Steam is non-toxic and non-corrosive, but for it
to be effective, it must hold all the water that it can carry in the form of transparent
vapour.

There are sterilizers for porous load(dressings, wrapped instruments and wrapped
gowns and drapes); sterilizers for fluids in sealed containers;sterilizers for
unwrapped instruments; and laboratory sterilizers (culture media, glassware and lab
equipment).

Thermometers and pressure gauges are recorded for every load.

Indicator tapes may be used.

Before, Bacillus stearothermophilus was used to test autoclaves. It can withstand

121c for 12 mins.
Dry heat kills micro-organisms by the destructive oxidation of essential cell
constituents. Killing of the most resistant spores requires a temperature of 160C for
2 hours. This is an efficient method for sterilizationand disposal of contaminated
materials (pathologic waste,surgical dressings, sharp needles and other clinical
waste).

Red heat is a form of dry heat, such as sterilizing inoculating wires using the flame
of a Bunsen burner, avoiding splattering.

Flaming is used for scalpels and the necks of flasks and test tubes.

Hot air sterilizers are used to process materials which can withstand high
temperatures, but will be affected by steam. (powders, microsurgical instruments)

IONIZING RADIATION

Ionizing radiation, including x-rays and gamma rays are lethal to all cells. Bacterial
species difer intheir sensitivity to ionizing radiation, and spores are generally more
resistant. This is used in the large-scale sterilization of plastic syringes and
catheters. It uses a linear accelerator or cobalt-60.

GASEOUS PROCESSES

Ethylene oxide is a highly penetrative, non-corrosive, microbicidal gas which is used

for the sterilization of single-use, heat –sensitive medical devices like prosthetic
heart valves. Materials are exposed to a gas concentration of 700-1000mg/L at 45-
60C, 70% humidity, for 2 hours.

Formaldehyde and low-temperature steam may be used for the reprocessing of

heat-sensitive equipments.

FILTRATION

Fluids, including bacterial cultures,can be rendered free of bacteria by passage

through filters with a pore size of less than 0.45 m.Most viruses and mycoplasmas
can pass through fileters with a pore size as low as 0.22 m. This method is used in
:

1. Separation of toxins and other soluble products of bacterial growth

2. Preparation of thermolabile parenteral solutions (antibiotics)

3. Certain blood products

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DISINFECTION

Moist heat is the method of first choice. Washing of laundry or eating utensils in
water at 70-80C for a few minutes will kill most non-spore forming micro-organisms.
Steam at 73C for thermolabile reusable equipment is also used. Exposure to boiling
water for 20 minute achieves disinfection, and may be used for emergencies.

Ultraviolet radiation using mercury lamps at 240-280nm may be used for treatment
of air, water and lab cabinets.

Gases such as formaldehyde may be used for complex heat –sensitive equipment
like anaesthetic machines and incubators.

Filtration of air removes micro-organisms from critical sites such as the operating
room, and for the labs handling pathogenic organisms. A high-efficiency particulate
air (HEPA) filter achieves 9.997% arrestance to particles of 0.5 m, and can produce
a sterile air.

Disinfection by Chemicals

1. Alcohols – isoprophyl, ethanol; these have optimal bactericidal activity at 70-

90% concentration. They have limited activity against mycobacteria and
spores; effective against viruses. Due to their volatile nature, they are
recommened as rapidly drying disinfectants for skn and surfaces(trolley tops,
thermometers). They are less effective in the presence of blood, and other
protein products.

2. Aldehydes- glutaraldehydes; broad spectrum action against bacteria, fungi

and viruses, but acts slowly against spores; often used for endoscopes. It is
an irritant to the eyes, skin and mcosa, hence needs to be handled in a well-
ventilated environment.

3. Biguanides-chlorhexidine; good for the skin and mucous membranes; low

irritancy and toxicity; may be combined with alcohol or a detergent for
handwashing.

4. Hypochlorites-broad spectrum, chlorine releasing disinfectant of choice for

viruses, including Hepatitis B.

5. Iodine-may cause staining and hypersensitivity; the iodophors and povidone-

iodine are less irritant and less staining. Good for preoperative preparation of
the skin.

6. Phenolics-environment disinfectants for hospitals and labs; broad spectrum

activity.
7. Surface active agents-anionic, cationic and non ionic detergents have
antimicrobial activity except for mycobacteria and spores.