Sterilisation & disinfection-PROCEDURE, DIFFERNCE ETC

2. DISINFECTION:
• PROCEDURE OF DESTRUCTION & REMOVAL OF ANY MICROORGANISM OUTSIDE BODY , CAUSING DISEASE OR INFECTION , BY
DIRECT EXPOSURE TO PHYSICAL & CHEMICAL AGENTS.
STERILISATION:
• PROCEDURE BY WHICH ANY OBJECT , SURFACE OR MEDIUM IS MADE FREE OF ANY KIND OF LIVING ORGANISMS (INCL.
SPORES).
SEPSIS:
• THE CONDITION BY WHICH A MICROORGANISM PROLIFERATES TO GIVE RISE TO ANY SYMPTOMS.
ANTISEPTICS:
• THE CHEMICALS , WHICH PREVENT OR INHIBIT THE PROLIFERATION OR GROWTH OF MICROORGANISM INSIDE HUMAN BLOOD.
• ANTISEPTICS ARE SUITABLE FOR APPLICATION TO LIVING TISSUES.
DISINFECTANTS (GERMICIDES):
• SUBSTANCES THAT DESTROY HARMFUL MICROBES (NOT USUALLY SPORES) WITH THE OBJECT TO PREVENT TRANSMISSION
OF DISEASES.
• DISINFECTANTS ARE SUITABLE FOR APPLICATION ONLY TO INANIMATE OBJECTS.

6.  SUNLIGHT- DRYING+UV RAY.
 DRYING- EVAPORATION.
 HEAT:-
 DRY HEAT: OXIDATION & DAMAGE TO DNA.
1. INCINERATION- BURNING TO ASHES.
2. RED HOT.
3. FLAMING- AT ABOUT 80˚C
4. HOT AIR OVEN- USUALLY 160˚C FOR 2 hrs.
5. INFRA-RED RADIATION- FROM A SOURCE HEATED AT ≥180˚C.
 MOIST HEAT:
 TEMPERATURE<100˚C
1. PASTEURISATION.
2. VACCINE BATH.
 TEMPERATURE AROUND 100˚C
1. BOILING AT NEUTRAL OR ALKALINE pH.
2. STEAM STERILISER (KOCH OR ARNOLD).
3. TYNDALISATION OR FRACTIONAL STERILISATION OR INSPISSATION.
 TEMPERATURE>100˚C
DRY SATURATED STEAM- AUTOCLAVE (USUALLY 15 ppsi PRESSURE AT 121˚C FOR 15 mins.).
 LOW TEMPERATURE STEAM WITH FORMALDEHYDE.

7.  COLD- FREEZE DRYING OR LYOPHILISATION.
 COLD & OSMOTIC SHOCK.
 IONISING RADIATION- BY DNA BREAKDOWN ABOVE 25 Grays (X RAY,β RAY,γ RAY).
 NON-IONISING RADIATION-
 BY FORMATION OF PURINE-PYRIMIDINE DIMERS BETWEEN ADJACENT MOLECULES IN THE SAME
DNA STRAND RESULTING IN NON-CODING LESIONS IN DNA & BACTERIAL DEATH OCCURS.
 UV RADIATION- WAVELENGTH OF 240-480 nm FOUND TO BE MOST EFFECTIVE.
 FILTRATION:-
 CANDLE FILTERS-
1. UNGLAZED CERAMIC FILTERS (CHAMBERLAND & DOULTON).
2. DIATOMACEOUS EARTH FILTERS (BERKEFELD & MANDLER).
 ASBESTOS FILTERS-
1. SEITZ.
2. STERIMAT.
3. CARLSON.
 SINTERED GLASS FILTERS.
 MEMBRANE FILTERS- MILLIPORE (PORE SIZE – 0.005-12 μ).
 HIGH-EFFICIENCY PARTICLE ABSORBENT FILTER (HEPA).

8.  APPLICATION OF FILTRATION:
I. THERMOLABILE,PARENTERAL & OPHTHALMIC SOLUTIONS.
II. STERILITY TESTING OF PHARMACEUTICAL PRODUCTS.
III. WATER QUALITY SURVEILLANCE.
IV. VIABLE COUNTING.
V. AIR STERILISATION.
VI. SERA STERILISATION.
1. BERKFIELD FILTERS
 MADE FROM KIESELGUHR, A FOSSIL DIATOMACEOUS EARTH.
 THREE GRADES OF POROSITY ARE AVAILABLE-
A. VEIL - COARSEST ONE.
B. N - NORMAL ONE.
C. W- WENIG THE FINEST ONE.
2. CHAMBERLAND FILTERS
 MADE FROM UNGLAZED PORCELAIN.
 FOUR GRADES ARE AVAILABLE.
A. L1- CLARIFYING FILTERS.
B. L1A-BIG.
C. L2 – NORMAL.
D. L3- FINEST.

9. 3. Seitz filter
 Made up of asbestos pads.
 Three grades are available-
A. K- clarifying filters.
B. Normal.
C. Special EK bacteria stopping filters.
4. Sintered glass filters
 Made from sintered glass.
 Different grades available-
 Grades 1 to 5-
A. Grades 1-2 are for clarifying purpose.
B. Grades3-5 is for sterilization purpose.
5. Membrane filters
 Made up of nitro-cellulose membranes.
 Made with different grades of porosity by adjusting the concentration of constituents.

10. MERITS AND DEMERITS OF
HEAT STERILIZATION
 Advantages
 Sterilization is very effective.
 Instruments are standardized to
deliver the required effective heat.
 Heat deliver system can be
monitored effectively with various
controls like pressure gauge,
temperature meters etc.
 Established quality control
methods available.
 Disadvantages
 Steam impermeable materials like fats, oils and
powders can not be sterilized by autoclaving.
 Heat sensitive materials can not be sterilized by heat.
 Examples:
1. Serum can not be sterilized
2. Antibiotics
3. Plastic materials
4. Vaccines
5. Rubbers
 Presence of organic matters interfere with effective
sterilization.
 Dangers of explosion when high pressure is used.

11. SPORES
 IMPAIRED GERMINATION.
 MEMBRANE DAMAGE.
 INCREASED SENSITIVITY TO
INHIBITORS.
 STRUCTURAL DAMAGE.
 CHROMOSOMAL DAMAGE.
VEGETATIVE FORMS
 RNA BREAKDOWN.
 COAGULATION OF PROTEINS.
 DAMAGE TO BACTERIAL
CHROMOSOMES.

12.  ACCEPTED METHOD WHICH CAN NOT BE RELIABLY PENETRATED BY STEAM AND CAN
WITHSTAND HIGH TEMPERATURES (160-180˚C).
 ELECTRICALLY OPERATED & FITTED WITH FANS FOR ENSURING ADEQUATE AIR CIRCULATION IN
THE CHAMBER.
 STERILISATION TEMPARATURE:
I. 160˚C FOR 1 hrs.
II. 170˚C FOR 40 mins.
III. 180˚C FOR 20 mins.
 USES: STERILISATION OF →
A. GLASSWARES- SYRINGES , PETRIDISHES , TEST TUBES , FLASKS ,PIPETTES etc.
B. SURGICAL INSTRUMENTS- FORCEPS , SCALPELS , SCISSORS.
C. OILS & FATS , IMPERMEABLE TO WATER.
D. CHEMICALS SUCH AS POWDERS WHICH FORM INTO CLUMP OR CAKE IN PRESENCE OF MOISTURE .
 STERILISATION CONTROL:-
 PHYSICAL- USE OF THERMOCOUPLE PERIODICALLY TO MEASURE ACCURATE TEMPERATURE .
 CHEMICAL- BROWNE’S TUBE WITH GREENISH SPOT.
 BIOLOGICAL- SPORES OF Bacillus subtilis niger . AFTER STERILISATION THE PAPER STRIPS CONTAINING THE
SPORES ARE REMOVED & CULTURED INTO THYOGLYCOLLATE/COOKED MEAT MEDIA FOR 5 DAYS AT 37˚C .

13.  STRUCTURE:
AN AUTOCLAVE ESSENTIALLY CONSISTS OF THE FOLLOWING →
o A CYLINDRICAL/RECTANGULAR CHAMBER , WITH CAPACITIES RANGING 400-800 LITRES.
o WATER-HEATING SYSTEM OR STEAM-GENERATING SYSTEM.
o STEAM OUTLET & INLET VALVES.
o SINGLE OR DOUBLE DOORS WITH LOCKING MECHANISMS.
o THERMOMETER OR TEMPARATURE GAUGE.
o PRESSURE GAUGE.
o SAFETY VALVE.
o STEAM JACKET.
o PERFORATED SHELF.
 INVENTED BY CHARLES CHAMBERLAND IN 1879.
 STERILISATION CONDITIONS: 15 ppsi PRESSURE FOR 15-20 mins. , WHICH YIELDS 121˚C OF
TEMPERATURE.
 USES: TO STERILISE CULTURE MEDIA( EXCEPT MEDIA CONTAINING EGG , SERUM & SUGAR ) , RUBBER
GOODS , SYRINGES , GOWNS , DRESSINGS etc. & IT IS THE SUREST METHOD OF DESTRUCTION OF BACTERIAL
SPORES.
 STERILISATION CONTROL :-
 PHYSICAL- USE OF THERMOCOUPLE PERIODICALLY TO MEASURE ACCURATE TEMPERATURE .
 CHEMICAL- BROWNE’S TUBE WITH GREENISH SPOT( MOST COMMON ) & AUTOCLAVE TAPE.
 BIOLOGICAL- SPORES OF Bacillus stearothermophillus , WHICH GROW IN AN OPTIMUM TEMPERATURE OF 55-60˚C & ARE

14.  WORKS ON THE PRINCIPLE OF STEAM UNDER
PRESSURE LIKE PRESSURE COOKER WHERE WATER
BOILS AT INCREASED ATMOSPHERIC PRESSURE.
 THE WATER IN THE AUTOCLAVE BOILS WHEN ITS
VAPOUR PRESSURE EQUALS THAT OF THE
SURROUNDING ATMOSPHERE.FOLLOWING INCREASE
IN PRESSURE INSIDE THE CLOSED VESSEL , THE
TEMPERATURE AT WHICH THE WATER BOILS INSIDE
THE AUTOCLAVE ALSO INCREASES.
 THE SATURATED STEAM THAT HAS A HIGHER
PENETRATIVE POWER , ON COMING IN CONTACT
WITH A COOLER SURFACE , CONDENSES TO WATER
& RELEASES ITS LATENT HEAT TO THAT SURFACE.
 THE GROSS REDUCTION IN VOLUME OF STEAM
SUCKS IN MORE STEAM TO THE AREA & THIS
PROCESS CONTINUES TILL THE SURFACE
TEMPERATURE IS ELEVATED TO THAT OF STEAM.
 STERILISATION IS ACHIEVED WHEN THE STEAM
CONDENSES AGAINST THE OBJECTS IN THE
CHAMBER & GRADUALLY RAISES THEIR
TEMPERATURE.THE CONDENSED WATER
FACILITATES THE MOIST CONDITIONS THAT ENSURES

15. DRY HEAT STERILISATION MOIST HEAT STERILISATION
KILLS THE ORGANISMS BY
DESTRUCTIVE OXIDATION OF
ESSENTIAL CELL CONSTITUENTS.
KILLS THE ORGANISMS BY
COAGULATING & DENATURING
THEIR ENZYMES & STRUCTURAL
PROTEINS.
160˚C FOR 60 mins. 121˚C FOR 15-30 mins.
LESS EFFECTIVE. MORE EFFECTIVE.
STERILISATION OF GLASSWARES-
SYRINGES, PETRIDISHES, TEST
TUBES, FLASKS, PIPETTES;
SURGICAL INSTRUMENTS-FORCEPS,
SCALPELS, SCISSORS; OILS & FATS,
IMPERMEABLE TO WATER;
CHEMICALS SUCH AS POWDERS
WHICH FORM INTO CLUMP OR CAKE
IN PRESENCE OF MOISTURE .
TO STERILISE CULTURE MEDIA
(EXCEPT MEDIA CONTAINING EGG,
SERUM & SUGAR), RUBBER
GOODS, SYRINGES, GOWNS,
DRESSINGS etc. & IT IS THE
SUREST METHOD OF
DESTRUCTION OF BACTERIAL
SPORES.

17.  CONCENTRATION.
 FORMATION.
 TIME OF EXPOSURE.
 pH OF MEDIUM.
 TEMPERATURE OF THE MEDIUM.
 NATURE OF THE TARGET ORGANISMS.
 INACTIVATION IN THE PRESENCE OF EXTRANEOUS MATERIALS.

18. Chemical
Agent
Gas
Sterilization Disinfection
Liquids
Animate
Chemotherapy Antiseptics
Inanimate
Sterilization Disinfection

19.  AGENTS INTERFERING WITH MEMBRANE FUNCTION-
 SURFACE-ACTIVE AGENTS
• QATERNARY AMMONIUM COMPOUNDS(CATIONIC)- CETRIMIDE.
• BETWEEN-80 , TRITON X-100(NON-IONIC).
• SOAPS , FATTY ACIDS(ANIONIC).
 PHENOLS
• PHENOL(CARBOLIC ACID).
• CRESOL(METHYL PHENOL).
• LYSOL(CRESOL+SOAP).
• DETTOL(HEXYLRESORCINOL).
 BIGUANIDE- CHLORHEXIDINE.
 ORGANIC SOLVENTS
• CHLOROFORM.
• TOLUENE.
• ALCOHOL.(ETHYL ALCOHOL- HAND STERILISATION ; METHYL ALCOHOL- FUNGICIDE)
• ISOPROPANOL.
 AGENTS DENATURING PROTEINS-
 ACIDS- HCl , BENZOIC ACID , FORMIC ACID , H2SO4 .
 ALKALIES- KOH.
 AGENTS DESTROYING OR MODIFYING FUNCTIONAL GROUPS OF PROTEINS-
 HEAVY METALS- MERTHIOLATE , MECUROCHROME , AgNO3 .
 OXIDISING AGENTS- HALOGENS , H2O2 , KMnO4
 DYES- ANILINE & ACRIDINE DYES.