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The process that effectively kills or eliminates transmissible agents (such as fungi, bacteria, viruses, spore forms, etc.) from the surface of equipment, article, food, medication, or biological culture medium is called sterilization. The Sterilization is carried out by the methods according to requirement. The methods are: 1. Moist Heat Sterilization 2. Dry Heat Sterilization 3. Gas Sterilization and Others.
Method # 1. Moist Heat Sterilization:
Moderate pressure is used in steam sterilization. Steam is used under pressure as a means of achieving an elevated temperature. It is important to ensure the correct quality of steam is used in order to avoid the problems which follow, superheating of the steam, failure of steam penetration into porous loads, incorrect removal of air, etc.
For aqueous preparations and for surgical dressing, heating in saturated steam under pressure is carried out. A number of time-temperature combinations have been proposed.
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The British and European Pharmacopoeia (2001) recommended 121°C temperature and 15 Ib/inch2 maintained throughout the load for 15 minutes as the preferred combination for this method of terminal sterilization. It is used to sterilize aqueous parenteral solutions and suspensions, surgical dressing and fabrics, plastic and rubbers closures, metal instruments, glass apparatus etc.
Method # 2. Dry Heat Sterilization:
Dry heat sterilization is used for heat-stable non-aqueous preparations, powders and certain impregnated dressings. It may also be used for sterilization of some types of container. Sterilization by dry heat is usually carried out in a hot-air oven. Heat is transferred from its source to load by radiation, convention and to a small extent by conduction.
Temperature-time exposures necessary to kill pathogen by dry heat indicates that a period of 90 minutes at 100°C destroys all vegetative bacteria but a period of 3 h at 140″C kills the spores.
Mould spores are of intermediate resistance and are killed in 90 minutes at 115°C. Most viruses have resistance similar to vegetative bacteria but some viruses are as resistant as bacterial spores e.g. virus that causes homologous serum jaundice.
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The British Pharmacopoeia (2001) states a minimum temperature of 160°C for at least 2 h for dry heat sterilization. Other combinations of temperature and time are permissible subject to first demonstrating a reproducible level of lethality in routine operation. Dry heat treatment, greater than 220°C, provides a useful method for sterilization and dehydrogenation of glassware in a particular container intended for a large volume of parenteral dosage.
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This process can remove heat-resistant endotoxin. In each cycle it is important to ensure that the whole content of each container is maintained for an effective combination of time and temperature especially to allow temperature variations in hot-air ovens, which may be considerable. Dry heat is used to sterilize glassware, porcelain and metal equipment, oils and fats and powders i.e. talc, etc.
Method # 3. Gas Sterilization:
Gaseous sterilizing agents are of two main types, oxidizing and alkylating agents. Vapour phase hydrogen peroxide is an example of the former. Ethylene oxide and formaldehyde are examples of the alkylating agents. However, the BP states that gaseous sterilization is used when there is no suitable alternative. The main advantage of ethylene oxide is that many types of materials, including thermolabile materials, can be sterilized without damage.
The gas can diffuse through packaging materials and rubber, and diffuse out after sterilization. It has the disadvantages of being toxic and combustible and also requires the correct humidity. In practice, the relative humidity in the chamber atmosphere is usually between 40 and 50% with temperatures up to 6°C.
Low temperature steam with formaldehyde has been used as an option for sterilizing thermolabile substances. Both ethylene oxide and formaldehyde have health risks and strict monitoring of personnel exposed to the gases required to ensure protection from harmful effects.
Method # 4. Sterilization by Radiation:
Radiations can be divided into two groups: electromagnetic waves and streams of particulate matter. The former group includes infrared radiation, ultraviolet light, X-rays and gamma rays. The latter group includes alpha and beta radiations. Most commonly infrared radiation, ultraviolet light, gamma radiation and high-velocity electrons are used for sterilization.
(i) Ultraviolet Light:
A narrow range of UV wavelength (220-280 nm) is effective in killing the microorganism. The wavelength close to 265 nm and adjacent wavelengths are strongly absorbed by the nucleoproteins. The most serious disadvantage of UV radiation as a sterilizing agent is its poor penetrating power. This is the result of strong absorption by many substances. The application of UV radiation is limited.
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(ii) Ionizing Radiations:
Ionizing radiations are suitable for commercial sterilization processes. It must have good penetrating power, high sterilizing efficiency, little or no damage effect on irradiated materials and are capable of being produced efficiently. The radiations that fulfill these four criteria are best high-speed electrons from machines and gamma rays from radioactive isotopes.
Sterilization by gamma rays is carried out using the radioactive isotope of [CO-60]. Articles for sterilization by radiations are packed in boxes of standard size which are sterilized by a series of slow passages around the gamma ray source. The absorbed dose for sterilization is 25 kGy.
The plastic syringes and catheters, hypodermic needles and scalped blades, adhesive dressings, single-application capsules of eye ointment, containers made of polythene and packaging materials using aluminium foil and plastic films are sterilized by gamma radiation.
Method # 5. Sterilization by Filtration:
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Membrane filters are made from cellulose derivates or other polymers. There are no loose fibres or particles in membrane filters. They retain particles larger than the pore size on the filter surface such filters particularly useful in detection of small numbers of bacteria.
Sterilization by filtration is a method permitted by the British pharmacopoeia (2001) for heat sensitive solutions or liquids that are not sufficiently stable to withstand the process of heating in an autoclave.
Passage through a filter of appropriate pore size (e.g. 0.22 pm) can remove bacteria and moulds. Viruses and mycoplasma may not be retained. After filtration the liquid is aseptically distributed into previously sterilized containers which are later sealed. This method has disadvantages that specialized facilities and skilled operations are required.
The final preparations cannot be released until the manufacturing batch has passed the appropriate test for sterility. Validation of autoclave has been described in detail in Practical Microbiology by Dubey and Maheshwari (2007).
