Sterilization

• Heating


  Most common method of sterilization
• Mechanisms of heating


  1. Formation of single strand breaks in the DNA

  2. Coagulation and Denaturation of proteins

  3. Accumulation of toxic levels of electrolytes

  4. Alteration of cell membrane structure
• Affecting factors of heating

  • Nature of the heat- moist heat is preferred
  • Temperature and Time- these are inversely proportional
  • Number of microorganisms- these are proportional
  • Type of material- Heat-sensitive materials will require a lower temperature compared to heat-resistant materials
  • Presence of organic material- Fats, proteins and sugars may need higher temperatures
• Moist Heat


  Has more rapid killing actions due to it speeding up coagulation and denaturation of proteins
• Pasteurization


  1. Destroys pathogens present in dairy products

  2. Conventional method: milk is heated at 60-65 C followed by rapid cooling

  3. Flash method: heating at 72 C for 15 second follow by quick cooling to 13 C

  4. UHT (Ultra-High Temperature): heating is done at 140 C for 15s and 149 C for half a second
• Vaccine Bath


  Destroys pathogens in vaccine preparations by heating to 60 C for 1 hour
• Serum Bath


  Inactivates bacteria using serum preparations by heating it in a water bath to 56 C for several days
• Inspissation


  Solidifies and disinfects egg-containing and serum-containing media by heating at 80-85 C for 30 minutes for 3 days
• Boiling


  Utilizes boiling water to destroy vegetative forms but not spores
• Fractional Sterilization


  Intermittent sterilization by exposing material to live steam at 100 C for 30-90 minutes for three days
• Autoclave


  Destroys all microbial forms at 15 psi + 121 C for 15-30 mins
• Red Flame


  Used to sterilize lab equipment by heating the object until it glows red
• Open Flame


  The object is passed several times on a flame and not heated until red, only destroys vegetative forms
• Incineration


  Burns organism to ashes, used for soiled beddings, carcasses and pathological material
• Hot Air Oven


  Articles are placed in an oven at 160 C for one hour to sterilize metallic instruments and some glasswares
• Infrared Rays


  Articles are placed on a conveyor belt and passed through a tunnel with infrared radiators at 180 C for seven and a half minutes
• Dessication


  Deprives the microorganism of moisture, used mainly for food preservation
• Freezing


  Not considered a reliable method of sterilization, used for preservation of microorganisms (lyophilization)
• Filtration


  Mechanical sieving that does not kill microorganisms but merely separates them from the fluid, using a cellulose ester filter
• Ultraviolet Light (UVL)/Non-ionizing Radiation


  Induces formation of thymine-thymine dimers, can inactivate microorganisms within seconds but is not sporicidal and has low penetrance
• Ionizing Radiation


  Have greater penetrance than UVL, causes formation of free-radicals that chemically interact with proteins and nucleic acids, not used much as they can damage human tissue
• Sonic and Ultrasonic Vibration

  Can kill bacteria after exposure to certain frequencies of around 20,000 cycles/second for one hour, by disrupting cells
• Factors affecting chemical sterilization

  • Concentration and potency of the chemical agent
  • Duration of exposure
  • Temperature
  • Nature of Surrounding Medium
  • Nature of the organism
  • Number of organisms
• Classification of Chemical Disinfectants

  • Damage to the Cell Membrane
  • Denaturation of Cellular Proteins
  • Modification of the Functional Groups of Proteins and Nucleic Acids
• Surface Active Agents


  Contain long chain hydrocarbons that are fat-soluble that cause leakage of cell components
• Cationic Agents


  Fat soluble portion is positively charged
• Anionic Agents


  Negatively charged chains of long hydrocarbons
• Phenol Compounds


  Disrupt cell membranes as well as precipitate proteins, bactericidal and fungicidal with good activity against Mycobacterium but poor activity against spores and most viruses
• Alcohols


  Disorganize lipid structure of the cell membrane
• Heavy Metals


  Cause damage to enzyme activity of bacteria
• Heavy Metals


  • Mercurials
  • Silver Compounds
• Halogens


  Oxidation of sulfhydryl groups
• Halogens


  • Iodine
  • Chlorine
  • Hydrogen peroxide
• Alkylating Agents


  Damage nucleic acids, kill everything including spores
• Alkylating Agents


  • Formaldehyde
  • Glutaraldehyde
  • Ethylene oxide