STERILIZATION & DISINFECTION

DEFINITIONS
Sterilization: total inactivation of all forms of

microbial life ability to reproduce Bactericide: agent to destroy bacteria Bacteristatic: substance that prevent bacterial growth, Disinfectant (germicide): agent that kills microorganisms capable of producing an infection. Antiseptic: opposes sepsis or putrefaction by killing bacteria or preventing their growth. (on living tissue) Decontamination: get rid of contamination (some of the microorganisms.

STERILIZATION
Physical Agents
1. Heat - Moist heat - Dry heat 2. Freezing 3. Radiation - ultra violet radiation - ionizing radiation 4. Ultrasonic & Sonic vibration 5. Filtration

Chemical Agents Mechanisms of action: 1. Damage the Cell Membrane 2. Denature Protein 3. Modify Functional Groups of Proteins and Nucleic Acids.

CHEMICAL AGENTS
Factors Affecting Disinfectant Potency 1. Concentration (Cnt = K) 2. Time of Exposure 3. pH 4. Temperature 5. Nature of the Organisms 6. Presence of Extraneous materials organic matter: serum, blood, pus, etc.

DEATH RATE OF MICROORGANISMS

6 5 4 3 g10 jml kteri ng bertahan dup setelah manasan

2 1 0 -1 1 2 3 4 5 6 o Lama pemanasan pada 121 C 7 D121

1. Agents that Damage the Cell membrane

Compounds Surface disinfectants CATIONIC AGENTS Quaternary ANIONIC AGENTS NON-IONIC Name Effects Practical Applic

Zephiran Ceepryn chloride Duponal LS Triton W-30 Tween-80

Gram-positive (mostly) Gram-positive (Only)

Antiseptic

Phenolic Cresol Lysol Creolin Diphenyl compound Hexachlorophene Alcohols Ethanol 70% Bactericidal Gram-positive Bactericidal spores Disinfectant Germicidal soap Disinfectant Germicidal soap + Disinfectant

2. Agents that denature Proteins

Free H+ & OH- ions undissociated molecule or altering pH of organisms environment
Compounds Acids Name Effects Practical Applic.

Hydrochloric acid Antibacterial Disinfectant Benzoic acid Food preservative Lactic, citric, acetic, Food preservative propionic acids Hydroxide (metallic cation) Ethanol 70% Acetone Direct toxic Bactericidal + spores Disinfectant Disinfectant Disinfectant

Alkalies Alcohol Acetone

3. Agents that Modify Functional Groups of Proteins & Nucleic acids

Compounds Name Effects Practical Applic . Heavy metals Mercurials Mercuric chloride Metaphen, Merthiolate, mercurochrome Phenylmercury acid Silver compound Silver nitrate 1%

Very toxic Less toxic

Gram-pos & neg, fungi, yeast, algae Bactericidal Agents

Limited use Antiseptics

Control contamination of Pseudomonas & others Prophylaxis of ophthalmia neonatorum Skin Disinfectant Disinfectant Food preservative Wound cleaning

Oxidizing HALOGENS

Betadine, Jodium Germcide Iodine ticture Germcide Chlorine (Sunklin) Damage DNA HYDROGEN PERH2O2 3% OXIDE

3. Agents that Modify Functional Groups of Proteins & Nucleic acids

Compounds Name Effects Practical Applic . Dyes Triphenylmethae Malachite green dyes Cristal violet Acridine dyes Proflavine Gram-positive Staining, medium Gram-positive Staining Synthesis of Wound antiseptics Nucleic acid

Alkylating agents Formadehyde Formalin (sol. 37%) Paraformaldehyde (solid 91-99%) Glutaraldehyde Gas Ethylene oxide Gas

All organisms (Irreversible)

Disinfetant Latex gloves

10 x formal- For rooms, dehyde fabrics, instr. Disinfectant for All bacteria Endoscopic instr.

Sterilisasi fisik
Efek mematikan dari panas basah; karena
denaturasi dan koagulasi protein Efek mematikan dari panas kering; karena denaturasi protein, kerusakan oksidatif dan efek toksik dr peningkatan konsentrasi elektrolit.

1. H e a t
A. Dry heat a. Hot air oven: metal/glass, powders, oils 2 hr 180oC b. Flaming : loops/se, glass object c. Incineration incinerator: medical trash B. Moist Heat Boiling water : 100oC 1 hr, but hr --- DTT Steaming: 100oC Autoclaving: 125oC, 15 atmosphere Pasteurization: 72oC for 15 seconds milk Tyndalization- liquid & semisolid sterilized 80-100oC 30 min for 3 d continue

Autoclave

Mechanisms of Thermal Injury:

protein denaturation & coagulation
Moist Heat
Temperature Time (min ) Pressure

Dry Heat
Time (min)

121oC 126oC 134oC 140oC

150oC 160oC 170oC

15 10 3 -

15 20 30

180
150 120 60

2. Freezing
- Freezing : preserving - Repeated freezing & thawing bacterial destruction. In freezing: formation of ice crystals outside the cells withdraw water from the cell interior intracellular electrolyte concentration & protein denaturation damage of cell membrane. Lyophilization= freeze-drying preserving culture bacteria

3. Radiation
A. Ultraviolet radiation B. Ionizing radiation

Sunlight : spontaneous sterilization occur under
natural condition.

Effects of Radiation: inactivation of molecule by rays

energy absorbed by molecule.

UV: bactericide Energy content of Ionizing radiations >>> than UV radiation

A. Ultraviolet radiation
- lethal & mutagenic agent - Lethal dose: for non-spore-forming bacteria 1800
6500 W/cm2. Bacterial spores required 10 X more

- bactericidal effect: wavelength of 240-280 nm - wavelength of 260 nm lethal but can not

penetrate the glass or water. practical use: to control air-borne infection

operating rooms or hospital wards, biological safety cabinet (laminar flow)

for

Laminar Flow

B. Ionizing radiation
Lethal effects: most pathogenic non-spore-forming bacteria Products by :
a. radioactive decay: , , rays b. x-rays, c. particle bombardment d. Nuclear reactor

Practical applications:

- Sterilizing dose : 2.5 Mrad most resistance microorgn - radiation of cobalt 60 : pharmacy and medicine, catgut, nylon sutures, disposable medical and medical supplies. Note: 1 Mrad = 106 rad 1 rad = 100 ergs/gr air absorbs

4. Ultrasonic & sonic vibrations

Sound vibration (20 to 1000 kc) disrupting cells. Ultrasonic waves more effective as the frequency is increased. Effects: - disintegrating cells & H2O2 formation - Depolymerization of macromolecules The most susceptible: gram negative rods The most resistance Staphylococcus : require long period exposure. No practical value for sterilization & disinfection : numerous survivors.

5. Filtration
Sterilization: heat-labile materials (lab) Mechanisms:
- Electrostatic & absorption phenomena - Physical construction of filters

Types:
- Old types : Berkefeld, Chamberland, Seitz - Membrane filters : porous disks of cellulose

Membrane filters
Pore size: 14-0.023 m. Most widely used: 0.023 m. Practical used: serum, plasma, etc. Two-dimensional screen. Mechanisms: a. Particles > pore : retained on filter surface b. Particle < pore: retained by van der Waals forces.

Filter