1. Norazli Ghadin

2. Objectives Understand and utilize correct sterilization and disinfection techniques Distinguish between sterilization and disinfection List the characteristics of an ideal antiseptic Describe sterilizing agents and rank their effectiveness Discuss the time/temperature relationship in destroying microorganisms

3. Disinfection Disinfection is the elimination of pathogens, except spores, from inanimate objects  Disinfectants are chemical solutions used to clean inanimate objects (physical processes, e.g., UV radiation, may also be employed to effect disinfection)  Germicides are chemicals that can be applied to both animate (living) and inanimate objects for the purpose of eliminating pathogens  Antiseptics are formulated for application to living tissue

4. The Ideal Disinfectant  Resistant to inactviation  Broadly active (killing pathogens)  Not poisonous (or otherwise harmful)  Penetrating (to pathogens)  Not damaging to non-living materials  Stable  Easy to work with  Otherwise not unpleasant

5. Disinfectant Performance…  Is dependent on Disinfectant concentrations  Is dependent on length (time) of administration  Is dependent on temperature during administration (usual chemical reaction 2x increase in rate with each 10°C increase in temperature)  Microbe type (e.g., mycobacteria, spores, and certain viruses can be very resistant to disinfection—in general vegetative cells in log phase are easiest to kill)  Substrate effects (e.g., high organic content interferes with disinfection—stainless steel bench easier to disinfect than turd)  It is easier (and faster) to kill fewer microbes than many microbes

6. Other Terms  Sanitization: Lowering of microbial counts to prevent transmission in public setting (e.g., restaurants & public rest rooms)  Degerming: Mechanical removal of microbes, e.g., from hands with washing  Sepsis: Bacterial contamination  Antisepsis: Reduction of or Inhibition of microbes found on living tissue  Germincides, Fungicides, Virucides  Physical versus Chemical disinfectants  Static (stasis) versus Cidal (e.g., bacteriostatic versus bacteriocidal)

7. Resistance to Killing  Gram-negative bacteria (with their outer membrane) are generally more resistant than gram-positive bacteria to disinfectants and antiseptics  Stationary-phase (I.e., non-growing) bacteria generally are more resistant than log-phase (I.e., growing) bacteria  Mycobacteria, endospores, and protozoan cysts and oocysts are very resistant to disinfectants and antiseptics  Nonenveloped viruses are generally more resistant than enveloped viruses to disinfectants and antiseptics  Organic matter (such as vomit and feces) frequently affects the actions of chemical control agent  Disinfectant activity is inhibited by cold temperatures  Longer application times are preferable to shorter  Higher concentrations, though, are not always preferable to lower concentration (e.g., alcohols)

8. Chemical Antimicrobi als AgentMechanisms of ActionComments Surfactants Membrane Disruption; increased penetration Soaps; detergents Quats (cationic detergent) Denature proteins; Disrupts lipids Antiseptic - benzalconium chloride, Cepacol ; Disinfectant Organic acids and bases High/low pHMold and Fungi inhibitors; e.g., benzoate of soda Heavy MetalsDenature proteinAntiseptic & Disinfectant; Silver Nitrate HalogensOxidizing agent Disrupts cell membrane Antiseptic - Iodine (Betadine) Disinfectant - Chlorine (Chlorox) Alcohol s Denatures proteins ; Disrupts lipids Antiseptic & Disinfectant Ethanol and isopropyl PhenolicsDisrupts cell membraneDisinfectant Irritating odor AldehydesDenature proteins Gluteraldehyde - disinfectant (Cidex) ; Formaldehyde - disinfectant Ethylene OxideDenaturing proteinsUsed in a closed chamber to sterilize Oxidizing agents Denature proteinsHydrogen peroxide – antiseptic ; Hydrogen peroxide – disinfectan; Benzoyl peroxide – antiseptic

9. Physical Antimicrobials AgentMechanisms of ActionComments Moist Heat, boiling Denatures proteinsKills vegetative bacterial cells and viruses Endospores survive Moist Heat, Autoclaving Denatures proteins121°C at 15 p.s.i. for 30 min k ills everything Moist Heat, Pasteurization Denatures proteinsKills pathogens in food products Dry Heat, Flaming Incineration of contaminants Used for inoculating loop Dry Heat, Hot air oven Oxidation & Denatures proteins 170°C for 2 hours ; Used for glassware & instrument sterilization Filtration Separation of bacteria from liquid (HEPA: from air) Used for heat sensitive liquids Cold, Lyophilization (also desiccation) Desiccation and low temperature Used for food & drug preservation; Does not necessarily kill so used for Long-term storage of bacterial cultures Cold, Refrigeration Decreased chemical reaction rate Bacteriostatic Osmotic Pressure, Addition of salt or sugar Plasmolysis of contaminantsUsed in food preservation (less effective against fungi) Radiation, UV DNA damage (thymine dimers) Limited penetration Radiation, X-rays DNA damageUsed for sterilizing medical supplies Strong vis. LightLine-drying laundry

10. Moist Heat  Moist heat kills microbes by denaturing enzymes (coagulation of proteins)  Boiling (at 100°C, I.e., at sea level) kills many vegetative cells and viruses within 10 minutes  Autoclaving: steam applied under pressure (121°C for 15 min) is the most effective method of moist heat sterilization—the steam must directly contact the material to be sterilized  Pasteurization: destroys pathogens (Mycobacterium tuberculosis, Salmonella typhi, etc.) without altering the flavor of the food—does not sterilize (63°C for 30 seconds)

11. Sterilization Times  121 o C, 15 minutes, moist heat (but don’t start the clock until entire item is up to temp—e.g., large volumes fluid)

12. Filtration: Air & Fluids

13. Filtration Membrane filters These are porous membrane about 0.1mm thick, made of cellulose acetate, cellulose nitrate, polycarbonate, and polyvinylidene fluoride, or some other synthetic material. The membranes are supported on a frame and held in special holders. Fluids are made to transverse membranes by positive or negative pressure or by centrifugation.

15. USES Remove toxin from bacterial cells Sterilize drugs (antibiotic) sterilize protein, glucose or other sugars Sterilize media with serum/heat sensitive substance Study virus size separate virus from cells

16. RADIATION Energy transmision through a space ruangan Basicly all radiation can cause a death/mutation on microorganisme

17. EFFECT OF RADIATION Changes Cells membrane inhibit DNA replication

18. Types of Radition 1. Ultraviolet (UV) 2. Ionizing radiation

19. UV Naturally from the sun. MoA; Induction of dimer thymine on DNA chain (mutation):  inhibit DNA replication  cell death. not all organisme can be kill. Wavelength used (260 nm)

20. UV: MERCURY LAMP 240 – 280 nm To decrease bacterial volume in atmosphere, operation theatre, nursery, restaurant & food factory. weakness : Low penetration power. Stop by glass, paper, dust or stain Can only penetrate a few milimeter of solution.

21. UV Adjacent bases bond with each other, instead of across the “ladder.” makes a bulge, and the distorted DNA molecule. Cant replicate MEDA 1012:MEDICAL MIKROBIOLOGI

22. IONIZING RADIATION X-RAY and gamma radiation (  ). More effective than UV. MoA: destroying by mutation or oxidation

23. Cont… Efective against SPORA. Source : isotope Cobalt 60. Dos 2.5 mrad. sterilizing disposable: · Syringe and needles · Chateter · heat sensitive items EXPENSIVE