Sterilization and Disinfection

1. Sterilization and Disinfection Xiao-Kui Guo PhD

2. Content • Basic terms and general mechanism • Physical agents • Chemical agents • Affecting factors

3. Section ABasic terms and general mechanisms

4. Basic terms • Sterilization灭菌: • A physical or chemical process that completely destroys all microbial life, including spore • Disinfection消毒: • Only destroy the diseases-producing microorganisms and the spore are sometimes not included

5. Basic terms • Bacteriostasis抑菌 • The property that is able to inhibit bacterial multiplication and the multiplication will resume upon when the inhibition agent is removed. • Antisepsis防腐 • A biocide or production that destroys or inhibits the growth of microorganisms in or on living tissue • Asepsis无菌 • The absence of pathogen microbes

6. The general mechanisms Disruption of cell membrane or wall Removal of free sulfhydryl groups Damage to DNA -SH -SS- DNA Chemical Antagonism Protein denaturation

7. Section BPhysical Agents

8. The element of physical agents Dry heat Heat Moist heat Radiation 辐射 Ultraviolet Radiation Ionizing Radiation microwave Physical agents Filtration 过滤 Ultrasonic Freezing

9. Heat The growth state of the bacteria changes with the temperature Temperature

10. Heat • Dry heat • Mechanism: the denaturation and coagulation of protein • Methods: • Hot air sterilization干烤 • Incineration焚化 • red heat赤热 • Flaming赤热

11. Heat • Moist heat • Mechanism: as same as dry heat but more effective • Autoclaving: 121℃,103.4kPa,20min • cidal for both vegetative organisms and endospores • Boiling water • Pasteurization: to kill particular spoilage organisms or pathogens • flash method: 71.7°C, 15~30s • holding method: 61.1~62.8℃, 30 min The hot stream surrounding the bodies of bacteria makes the moist heat a more effective mean for killing

12. Heat • Moist heat • fractional sterilization间歇蒸气灭菌法 • Steam heating to 100 °C for 30 min Vegetative cells are destroyed but endospores survive • Incubate at 30 °C -37 °C overnight Most bacterial endospores germinate • Second heat treatment, 100 °C, 30 min Germinated endospores are killed. • Second incubation at 30°C-37 °C overnight Remaining endospores germinate • Third heat treatment, 100 °C, 60 minLast remaining germinated endospores are killed

13. Radiation • Ultraviolet • Mechanism: Ultraviolet leads to the formation of adjacent two thymine to form dimer. • The most effective wavelength is in the 240 to 280nm with the optimum at about 260nm Even if faces the glass or paper, the Ultraviolet can not penetrate The thymine-thymine dimer

14. Radiation • Ionizing Radiation • X-rays and gamma rays • more energy and penetrating power than UV • used to sterilize pharmaceuticals and disposable medical supplies such as syringes, surgical gloves, catheters, and sutures • used to retard spoilage in seafoods, meats, poultry, and fruits

15. Filtration • The heat destroys the bacteria and meanwhile the useful ingredient doesn’t survive. • Filtration set a barrier to keep out the particle at a low temperature • Diameter of the pores: 0.22~0.45μm Particle, including bacteria, keeping out Water and gas can pass the barrier

16. Ultrasonic To form cavities about 10 μm in diameter in liquid The cavities collapse of the pressure of 1000 atmosphere The cavitation produces a number of chemical and physical changes in medium. Hydrogen peroxide produced Ultrasonic Causing a depolymerization of macromolecules and intramolecular regrouping • Because there are numerous survivors after the ultrasonic treatment, so it is of no practical value in sterilization and disinfection Bacteria killed

17. Freezing • Although many bacteria are killed by exposure to cold, freezing is not a reliable method of sterilization. • The primary use is in the preservation of bacteria. • The best way to preserve the bacteria nowadays is lyophilization冷冻干燥.

18. Section CChemical Agents

19. Chemical agents I: • Phenol and phenol derivatives • alter membrane permeability and denature proteins • chlorhexidine • ineffective against endospores

20. Chemical agents II: • Soaps and detergents • Anionic (negatively charged) detergents: mechanically remove microorganisms and other materials but are not very microbicidal. • Cationic (positively charged) detergents: alter membrane permeability and denature proteins; ineffective against endospores, M. tuberculosis, and P. species.

21. Chemical agents III: • Alcohols • denature membranes • 70% solutions of ethyl or isopropyl alcohol • ineffective against endospores and non-enveloped viruses

22. Chemical agents IV: • Heavy metals denature proteins • Mercury compounds (mercurochrome; merthiolate) : bacteriostatic, ineffective against endospores • Silver nitrate (1%) : put in the eyes of newborns to prevent gonococcal ophthalmia.

23. Chemical agents V: • Chlorine reacts with water to form hypochlorite ions, which in turn denature microbial enzymes • Iodine and iodophores • denatures microbial proteins • effective against some endospores

24. Chemical agents VI: • Aldehydes denature microbial proteins • Formalin (37% aqueous solution of formaldehyde gas) • glutaraldehyde: kill vegetative bacteria in 10-30 minutes and endospores in about 4 hours

25. Section DAffecting factors

26. Factors Influencing Antimicrobial Activity • The concentration and kind of a chemical agent used; • The intensity and nature of a physical agent used; • The length of exposure to the agent; • The temperature at which the agent is used; • The number of microorganisms present; • The species or strain of microorganism; • The nature of the material bearing the microorganism; • The presence of organic or other interfering substances.

27. The End Thanks