Protection Against Infections – II

I. PHYSICAL AGENTS

There are different types of physical agents.

(1). Heat: The heat employed for removal of micro-organisms varied with the nature of object and also depend on the purpose. Based on these different processes are employed.

(a). Moist heat

  • It is the widely used effective means of sterilization process.
  • In this, steam under high pressure is employed which imparts high penetration power resulting in the hydration of cells and coagulation of protein leading to the death of the microorganism.
  • The autoclave is the apparatus used for sterilization by moist heat.
  • The autoclave is a double-jacketed steam chamber. The chamber is equipped with a device for generating saturated steam. It can be maintained at a particular temperature and pressure for any period of time.
  • During operation of autoclave the air in the chamber is evacuated by steam since presence of air will reduce the temperature in the chamber.
  • The time required for sterilization will depend upon the materials to be sterilized. Solid materials must be heated for a longer time (1-2 hours) while liquid media can be sterilized within 15-30 minutes.
  • Also, acidic materials require shorter period than alkali materials. A temperature of 121°C for 15 min at a pressure of 15 lbs/ sq.inch is the sterilizing condition in the autoclave.
  • Steam can penetrate through materials and sterilization is achieved by the coagulation or denaturation of proteins and other cell constituents.

Advantages

  • Liquid media, solid media, laboratory equipment (cloth, glassware, etc.,) can be sterilized.
  • The temperature and pressure is high enough to kill spores, vegetative cells and viruses.

Disadvantages

  • Temperature-sensitive media, animal tissue culture media, antibiotics, amino acids, cannot be sterilized.
  • Sometimes water may get inside in case of improper packing.
(b). Dry heat
  • This process is accomplished in a hot-air oven. Hot air or dry heat is employed for sterilization.
  • The dry heat penetrates substances more slowly than the moist heat. Hence, the time required for effective sterilization is long (2 to 3 hours), and also the temperature required is too high (160°C-180°C).
  • Microbial death results from the oxidation of cell constituents.

Advantages

  • Dry heat does not corrode glassware and metal instruments as moist heat does.
  • All glassware can be sterilized.

Disadvantages

  • The sterilization process is slow.
  • It is not suitable for heat-sensitive materials like many plastic and rubber items.
(c). Boiling
  • Boiling at 100°C for 30 minutes. Kills everything except some endospores (Actually, for the purposes of purifying drinking water 100o C for five minutes is probably adequate though there have been some reports that Giardia cysts can survive this process).
  • To kill endospores, and therefore sterilize the solution, very long or intermittent boiling is required.
(d). Pasteurization
  • Pasteurization is the use of mild heat to reduce the number of microorganisms in a product or food.
  • In the case of pasteurization of milk the time and temperature depend on killing potential pathogens that are transmitted in milk, i.e., staphylococci, streptococci, Brucella abortus and Mycobacterium tuberculosis.

For pasteurization of milk:

batch method (Low-temperature holding): 62.8°C for 30 minutes

flash method (High-temperature short time): 71.7°C for 15 seconds

(e). Intermittent sterilization or Tyndallization

  • Intermittent sterilization or Tyndallization is the process of boiling the materials at 100°C for 30 min. successively for three consecutive days.
  • Destroys vegetative cells and spores; germinated spores.

(b). Incineration

  • Incineration burns organisms and physically destroys them.
  • Incineration is the complete burning of the material into ashes.
  • Used for needles, inoculating wires, glassware, etc. and objects not destroyed in the incineration process.
  • This is the direct and ultimate method of destroying cells.
  • It is achieved by keeping the materials directly in contact with the flame of Bunsen burner as a result all the microorganisms in the surface are destroyed completely.
  • Inoculating loops, needles and spreading rods are sterilized by this method.

Advantages:

  • Immediate and quick.

Disadvantages:

  • Cannot be used to sterilize heat-labile material, material is lost by incineration.

 

(2). Radiation
  • Energy transmitted through space in a variety of forms is generally called radiation. It is also known as “cold sterilization” as only little heat is produced during the process.
  • The most significant of this is electromagnetic radiation. The energy content and radiation wavelength are inversely proportional to each other.
  • Radiation may be ionizing or non-ionizing.

Ionizing radiation

  • High-energy electron beams (Gamma, X-rays, alpha and beta particles) have sufficient energy to cause ionization of molecules.
  • They drive away electrons and split the molecules into ions.
  • Water molecules are split into hydroxyl radicals (OH-), electrons and hydrogen ions (H+). OH- ions are highly reactive and destructive to normal cellular compounds such as DNA and proteins.
  • Thus ionizing radiations are used in sterilization.
  • X-rays and Gamma rays have high penetrating power.
  • Packed food and medical equipment are sterilized by using x-rays and gamma rays.
  • Generating and controlling X-rays for sterilization is highly expensive.
  • This includes ultraviolet (UV) rays.
  • UV at a wavelength of 265 nm is most bactericidal.
  • e.g. 36Cs, 60Co

Advantages:

  • X-rays and Gamma rays have a high penetrating power.
  • Packed food and medical equipment are sterilized by using x-rays and gamma rays.

Disadvantage:

  • Generating and controlling X-rays for sterilization is highly expensive.

Non-ionizing radiation

  • This includes ultraviolet (UV) rays.
  • UV at a wavelength of 265 nm is most bactericidal
  • Absorption of UV radiation produces chemical modification of nucleoproteins i.e., thymine dimer formation that leads to misleading of genetic codes.
  • This mutation impairs the total functions of the organism, consequently causing its death.

Advantages:

  • It is used to maintain aseptic conditions in laminar air flow chamber, lab, hospitals, pharmaceuticals, industries etc., and also in the sterilization of water and air.

Disadvantage:

  • UV radiation has very little ability to penetrate matter and hence the microorganisms on the surface of an object are destroyed.

(3). Filtration

  • Filtration involves the passage of liquid or gas through a screen-like material that has spores small enough to retain the microorganism of a certain size.
  • It is used to sterilize heat-sensitive substances like enzyme solutions, bacterial toxins, certain biological media, cell extract and some sugars.
  • Various types of filters are available in different grades of porosity.
  • Vacuum or pressure is required to move the solutes through the filter.
  • Involves the physical removal of all cells in a liquid or gas, especially important to sterilize solutions which would be denatured by heat (eg: antibiotics, injectable drugs, amino acids, vitamins, etc.)

Advantages:

  • It is the best way to reduce microbial population in solutions of heat-sensitive materials and it is sued to sterilize liquid media, vitamin solutions, hormones, growth factors, enzymes.

Disadvantages

  • Pleomorphic structures like mycoplasma cannot be effectively filtered by this technique.
  • It is applicable to sterilize only small quantities.

(4). Low temperature

  • Most organisms grow very little or not at all at 0OC.

    Store perishable foods at low temperatures to slow rate of growth and consequent spoilage (eg: milk).

  • Low temperatures are not bactericidal.
  • Psychrotrophs, rather than true psychrophiles are the usual cause of food spoilage in refrigerated foods.

(5). Desiccation / Drying (removal of H2O)

  • Most microorganisms cannot grow at a reduced water activity (aw< 0.90).
  • Often used to preserve foods (eg: fruits, grains, etc).
  • Methods involve removal of water from product by heat, evaporation, freeze-drying, addition of salt or sugar.
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