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Sterilization Methods:

Sterilization Methods Physical Means of Sterilization Chemical means of Sterilization

Physical Means of Sterilization:

Physical Means of Sterilization Sterilization by heat Moist heat Sterilization Dry Heat Sterilization Radiation Filtration

Sterilization by heat:

Sterilization by heat Heat is the most reliable and widely used means of sterilization. Affording its antimicrobial activity through destruction of enzymes and other essential cell constituents This method of sterilization is limited to thermostable products , but can be applied to both moisture-sensitive and moisture resistant products by dry ( 160-180°C ) and moist (1 21-134°C ) heat sterilization, respectively.

Steam sterilization (moist heat):

Steam sterilization (moist heat) Boiling Autoclaving Pasteurization

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Widespread application in the processing of many thermostable products and devices . Used in the sterilization of dressings, surgical and diagnostic equipment , containers and closures , and aqueous injections , ophthalmic preparations and irrigation fluids Involves the use of steam at temperatures in the range 121-134°C. Achieved by the generation of steam under pressure

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The elevated temperatures generally associated with moist heat sterilization methods can only be achieved by the generation of steam under pressure. By far the most commonly employed standard temperature/time cycles for bottled fluids and porous loads (e.g. surgical dressings) are 121 °C for 15 minutes and 134°C for 3 minutes, respectively.

Table 1: Pressure-temperature relationships and antimicrobial efficacies of alternative steam sterilization cycles :

Table 1: Pressure-temperature relationships and antimicrobial efficacies of alternative steam sterilization cycles Temperature ( o C ) Holding time (minutes) Steam pressure ( kPa ) 115 30 69 121 15 103 126 10 138 134 3 207

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Moist heat sterilization may be achieved effectively by the following widely accepted known methods, such as : Boiling, Autoclaving Pasteurization Boiling Boiling at 100°C at 760 mm atmospheric pressure is found to kill particularly several varieties of vegetative states of microbial strains, a good number of viruses and fungi ; besides their ‘spores’ within a span of 10 minutes only .

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It is quite obvious that the ‘unpressurized’ i.e., free-flowing steam is practically equivalent to the prevalent temperature of boiling water (i.e., 100°C). It has been revealed that the endospores plus certain viruses are evidently not destroyed in such a short duration of 10 minutes. Examples. - a typical hepatitis virus - certain microbial endospores

Autoclaving(moist heat):

Autoclaving(moist heat) The most reliable sterilization methods Requires such ranges of temperature that are critically above the boiling water i.e ., above 100°C These high temperatures [120 ± 2°C] are most conveniently accomplished by moist steam under positive pressure (350-400kPa ) usually in an ‘autoclave’. One may make use of ‘autoclaving’ as a means of sterilization unless the drug substance or material to be sterilized can suffer serious type of damage either by heat or by moisture .

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In fact, higher the pressure inside the autoclave, the higher will be the temperature inside the autoclave.

Applications of an Autoclave:

Applications of an Autoclave To sterilize culture media To sterilize various surgical stainless steel instruments that are required for most of the surgical procedures , dental procedures , obstretrics etc . To sterilize various types of surgical dressings, gauzes, sutures etc . To sterilize a host of IV applicators, equipments, solutions, and syringes as well Aqueous parentral solution and suspension

Dry-Heat Sterilization:

Dry-Heat Sterilization It is a well known fact that microorgansims get killed by dry heat due to the oxidation effects . Direct Flaming . Direct flaming designates one of the most simple method of dry-heat sterilization. Used in a ‘ microbiology laboratory ’ for the sterilization of the ‘ inoculating loops ’, which is duly accomplished by heating the loop wire to a ‘ redglow ’, This is 100% effective in actual practice . ‘ Inceneration ’ to sterilize as well as dispose of heavily contaminated paper bags, cups, and used dressings.

Hot-Air Sterilization:

Hot-Air Sterilization In this particular process, the various items need to be sterilized are duly kept in an electric oven, preferably with a stainless-steel chamber inside, and duly maintained at 170°C for a duration of approximately 2 hours (to ensure complete sterilization ). It has been adequately observed that the longer the period plus higher temperature are needed profusely due to the fact that the heat in water is more rapidly passed onto a ‘cool body’ in comparison to the heat in air.

Application of hot air oven:

Application of hot air oven Restricted to : glassware and metal surgical instruments non-aqueous thermostable liquids thermostable powders . impregnated dressing Oils, fats and waxes Ophthalmic ointments


Pasteurization Pasteurization refers to ‘ the process of heating of a fluid at a moderate temperature for a definite period of time to destroy undesirable microorganisms without changing to any extent the chemical composition .’ Example. In pasteurization of milk, pathogenic organisms are invariably destroyed by heating at 62 ° C for a duration of 30 minutes, or by ‘flash’ heating to higher temperatures for shorter period of time which is otherwise known as high-temperature short time (HTST) pasteurization.

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In a pasteurization of milk, effectively lowers the total bacterial count of the milk by almost 97 to 99 %. This is due to the fact that the most prevalent milk-borne pathogens viz., Tubercle bacillus , and Samonella , Streptococcus , and Brucella organisms, fail to form ‘spores’, and are quite sensitive to heat . It may, however, be observed that several relatively heat-resistant ( thermoluric ) microorganisms do survive pasteurization

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Ultra-High-Temperature (UHT) Treatments. Sterilization of milk is absolutely different from pasteurization . It may be duly accomplished by UHT treatments in order that it can be most easily and conveniently stored even without any sort of refrigeration Summary Pasteurization : At 63°C for 30 minutes ; HTST-Treatment : At 72°C for 15 seconds ; UHT-Treatment : At 140°C for < 1 second ;


Filtration Filtration may be defined as ‘the process of removing particles from a solution by allowing the liquid position to pass through a membrane or other particle barrier ’ Contains tiny spaces or holes which exclusively allow the liquid to pass but are too small to permit the passage of the small particles . the process of a liquid or gaseous substance via a screen-like material having suitable pores small enough to retain the microorganisms (bacteria).

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Employed to sterilize the specific heat sensitive substances, namely : culture media ; vaccines ; enzymes ; and several antibiotic solutions . High-Efficiency Particulate Air (HEPA) Filters . HEPA-Filters are mostly used to get rid of practically all microbes that happen to be larger than 0.3 μm in diameter . Examples . HEPA-Filters are largely used in : Intensive-Care Units [ICUs] in specialized hospitals treating severe burn cases. In Sterile Zones of High-Value Antibiotic Preparations, Packaging, IV-injections, and other such sensitive sterile preparations.

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Membrane Filters advanced membrane filters made up of either Cellulose Esters or Plastic Polymers have been employed profusely for the laboratory and industrial applications. Membrane filters usually have a thickness of 0.1 μm , and having almost uniform pores . The pores of membrane filters usually range between 0.22 to 0.45 μm , intended for microorganisms.


Radiation Several types of radiation find a sterilizing application in the manufacture of pharmaceutical and medical products. They are generally categorized into two groups: electromagnetic waves such as X-rays, gamma rays, ultraviolet (UV) light, and Infrared accelerated electrons (particulate radiation) such as alpha and beta radiation

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The major target for these radiation is microbial DNA . Gamma rays and electrons cause ionization and free radical production while UV light causes excitation . UV irradiation is not as efficient a sterilization method as electron or gamma-irradiation.

Applications of radiation sterilization:

Applications of radiation sterilization useful method for the industrial sterilization of heat-sensitive products Applied to articles in the dry state ; including surgical instruments, sutures, prostheses, unit dose ointments, plastic syringes and dry pharmaceutical products.

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UV light , with its much lower energy, and poor penetrability finds uses in the sterilization of air , for surface sterilization of aseptic work areas , for treatment of manufacturing grade water, but is not suitable for sterilization of pharmaceutical dosage forms.

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