This document discusses sterilization and disinfection methods. It defines sterilization as the process of removing all microorganisms, including spores, while disinfection reduces microorganisms to non-disease causing levels. Physical sterilization methods include heat (dry heat, moist heat like autoclaving), radiation, and filtration. Chemical methods use substances like alcohols, aldehydes, halogens, dyes, phenols, and biguanides. Proper sterilization requires management systems, facilities, equipment, trained staff, and record keeping to be effective.

1. STERILIZATION AND
DISINFECTION
UPAKAR PAUDEL (B.Sc. MIT)
UCMSTH BHAIRAHAWA

2. INTRODUCTION
• Microorganisms are the agents of
contamination, infection, and decay.
• Hence it becomes necessary to remove
them from materials and areas.
• Early civilization practiced salting, smoking,
pickling and exposure to sunlight .

3. In mid 1800s Lister developed
Aseptic techniquesAseptic techniques to prevent contamination
of surgical wounds. Prior to this
development:
•Nosocomial infections caused death in
10% of surgeries.
•Up to 25% mothers delivering in hospitals
died due to infection.

4. Requirements for decontamination practice
The basic requirements for good decontamination
practice are:
• A Management control system;
• Appropriate facilities;
• Appropriate equipment;
• Properly trained and supervised staff;
• Ensuring that single use medical devices are not
reused;
• Records of decontamination are kept.

6. DEFINITIONDEFINITION
STERILIZATIONSTERILIZATION
The process of freeing an article from microorganisms including
their spores.
DISINFECTION:DISINFECTION:
Reducing the number of pathogenic microorganisms to
the point where they no longer cause diseases.

7. SepsisSepsis: Comes from Greek for decay or putrid.
Indicates bacterial contamination.
AsepsisAsepsis: Absence of significant contamination.
• Aseptic techniques are used to prevent
contamination of surgical instruments, medical
personnel, and the patient during surgery.
• Aseptic techniques are also used to prevent
bacterial contamination in food industry.

8. Bacteriostatic Agent:
An agent that inhibitsinhibits the growth of bacteria, but
does not necessarily kill them.
Bactericide:
An agent that kills bacteria. Most do not kill Endospores.
Sporicide:
An agent that kills spores.

9. Methods of Sterilisation
Microbial ControlMicrobial Control
MethodsMethods
Physical AgentsPhysical Agents Chemical AgentsChemical Agents
Mechanical RemovalMechanical Removal
MethodsMethods

10. Physical AgentsPhysical Agents
Heat Radiation
Dry Moist
Incineration
Dry Oven
Steam Under
Pressure
Boiling Water/Hot Water
Pasteurization
Sterilization
Ionizing Non Ionizing
X Ray, Cathode,
Gamma
Disinfection
UV
Sterilization Disinfection

11. Chemical AgentChemical Agent
Gas Liquids
Sterilization Disinfection Animate Inanimate
Chemotherapy Antiseptics Sterilization Disinfection

12. Mechanical RemovalMechanical Removal
MethodsMethods
Filtration
Air Liquids
Disinfection Sterilization

13. Physical Methods of Sterilisation
Sterilisation By Dry Heat:
Hot Air Oven
• Kills by oxidation effects
•The oven utilizes dry heat to sterilize articles
• Operated between 50o
C to 250/300o
C.
•A holding period of 160o
C for 1 hr is desirable.
• There is a thermostat controlling the
temperature.
•Double walled insulation keeps the heat in and
conserves energy,
Uses:Uses:
• To sterilise Forceps, Scissors, Scalpels, Swabs.
• Pharmaceuticals products like Liquid paraffin, dusting powder, fats
and grease.

14. FLAMING
• Inoculation loop or Wire, the tip of
Forceps and spatulas are held in a
bunsen flame till they are red hot.
INCINERATION
• This is an excellent method of destroying materials such
as contaminated cloth, animal carcasses and
pathological materials.

15. MOIST HEAT STERILIZATION
Kills microorganisms by coagulatingcoagulating their
proteins.

16. MOIST HEAT STERILISATION IS CARRIED OUT WITH FOLLOWING METHODSMOIST HEAT STERILISATION IS CARRIED OUT WITH FOLLOWING METHODS
• Temp below 100o
C: “Pasteurisation”, Inspissator.“Pasteurisation”, Inspissator.
• Temperature at 100o
C:.. BoilingBoiling
• Steam at atmospheric pressure: Koch/Arnold’s steamerKoch/Arnold’s steamer..
• Steam under pressure: Autoclave.Autoclave.

17. HOT WATER BATH
• To inactivate non sporing
bacteria for the preparation of
vaccines - Special vaccineSpecial vaccine
bathbath at 60o
Cfor one hour is
used
• Serum or body fluids
containing coagulable
proteins can be sterilized by
heating for 1 hr at 56o
C in a
water bath for several
successive days.
Water BathWater Bath

18. INSPISSATOR
• Sterilizes by heating at 80-85o
C for half an hour for 3
successive days
• Used to sterilize media such as Lowenstein-Jensen &
Loefller’s serum

19. TEMPERATURE AT 100O
C
Boiling:
Kills vegetative forms of bacterial pathogens.
• Hepatitis virus: Can survive up to 30 minutes
of boiling.
Endospores: Can survive up to 20 hours or
more of boiling

20. Steam Sterilizer

21. STEAM AT ATMOSPHERIC PRESSURE
• Steam is generated using a steamer (Koch/
Arnold)
• Consists of a Tin cabinet
• Has a conical lid to enable the drainage of
condensed steam
• Perforated tray above ensures materials are
surrounded by steam.
• For routine sterilization exposure of 90 mins is
used

22. • For media containing sugar and gelatin
exposure of 100o
C for 20 min for 3 successive
days is used
• The process is termed as
Tyndallisation /Intermittent SterilizationTyndallisation /Intermittent Sterilization

23. AUTOCLAVE

24. STEAM UNDER PRESSURE - AUTOCLAVE
• Works on the principle of Steam under pressureSteam under pressure
• Invented by Charles Chamberland in 1879.

25. • Autoclave consists of a vertical or a horizontal
cylinder.
• One end has an opening which is meant for
keeping materials to be sterilised.
• The lid is provided with a Pressure gauge, to
measure the pressure
• A safety valve is present to permit the escape
of steam from the chamber

26. • Articles to be sterilised are placed in the
basket provided
• Sterilisation is carried out under pressure at
121º for 15 mnts.

27. Methods of Sterilization
The air in the chamber is evacuated and filled with saturated steam.
The chamber is closed tightly the steam keeps on filling into it and the
pressure gradually increases.
The items to be sterilized get completely surrounded by saturated
steam (moist heat) which on contact with the surface of material to
be sterilized condenses to release its latent heat of condensation
which adds to already raised temperature of steam so that eventually
all the microorganisms in what ever form –are killed.
The usual temperature achieved is 121 °C at a pressure of 15 pps.i. at
exposure time of only 15-20 mins. By increasing the temperature, the
time for sterilizing is further reduced.

29. Methods of Sterilization
Advantages of Autoclave
• Temperature is > 100°C therefore spores are killed.
• Condensation of steam generates extra heat (latent heat of
condensation).
• The condensation also allows the steam to penetrate rapidly into
porous materials.
• Note: that autoclavable items must be steam permeable. Can not be
used for items that are lacking water.

30. Monitoring of autoclaves
Physical- use of thermocouple to measure
accurately the temperature.
Chemical- it consists of heat sensitive chemical
that changes color at the right temperature and
exposure time.
Autoclave tape
Browne’s tube.
Biological – where a spore-bearing organism is
added during the sterilization process and then
cultured later to ensure that it has been killed.

31. CHEMICAL AGENTS
Chemical agents act byChemical agents act by
• Protein coagulation
• Disruption of the cell membrane
• Removal of Sulphydryl groups
• Substrate competition

32. DISINFECTANTS
Alcohols
• Both ethanol and isopropanol (rubbing alcohol) are widely used,
normally at a concentration of about 70%.
• Concentrations of 60% to 95% are effective.
• They are bactericidal and fungicidal but are not effective against
endospores or non-enveloped viruses.
• Alcohols enhance the effectiveness of other chemical agents.
Uses
Disinfection of clinical thermometer.
Disinfection of the skin – Venupuncture

33. ALDEHYDES
• Formaldehyde & Glutaraldehyde are
frequently used
• Formaldehyde is bactericidal, sporicidal & has
a lethal effect on viruses.
• Glutaraldehyde is effective against Tubercle
bacilli, fungi and viruses

34. USES
FORMALDEHYDEFORMALDEHYDE
• To preserve anatomical specimens (dead body, etc)
Destroying Anthrax spores in hair and wool
• 10% Formalin+0.5% Sodium tetra borate is used to
sterilise metal instruments

35. USES
GLUTARALDEHYDEGLUTARALDEHYDE
• Used to treat corrugated rubber anesthetic
tubes, Face masks, Plastic endotracheal tubes,
Metal instruments and polythene tubing
Glutaraldehyde are more active against
hepatitis viruses than most other
disinfectants

36. DYES
Two groups of dyes are used
Aniline dyesAniline dyes
Acridine dyesAcridine dyes

37. ANILINE DYES
• Are Brilliant green, Malachite green & Crystal
violet
• Active against Gram positive bacteria
• No activity against tubercle bacilli

38. ACRIDINE DYES
• Acridine dyes in use are orange in colour
• Effective against Gram positive than Gram
negative
• Important dyes are Proflavine,
Acriflavine,Euflavine

39. HALOGENS
• IodineIodine in aqueous and alcoholic solution has
been used widely as a skin disinfectant
• Actively bactericidal with moderate against
spores
• ChlorineChlorine and its compounds have been used
as disinfectants in water supplies & swimming
pools

40. DISINFECTANTS
Biguanides
 Chlorhexidine, a member of the biguanide group, is not a phenol, but its structure
and applications resemble hexachlorophene. It is frequently used for surgical skin
preparation and surgical hand scrubs.
Halogens
 Iodine is effective against all kinds of bacteria, many endospores, fungi, and some
viruses. Its mechanism of activity may be its combination with the amino acid
tyrosine in enzyme and cellular proteins.
 An iodophore is a combination of iodine and an organic molecule. Iodophores do
not stain and are less irritating than iodine. Examples are Isodine and Betadine.
 Chlorine is used as a gas or in combination with other chemicals. Chlorine gas is
used for disinfecting municipal water supplies, swimming pools, and sewage.
Sodium hypochlorite – ordinary household bleach- is good disinfectant.
 Chloramines consist of chlorine and ammonia. They are more stable than most
chlorine. The U.S. military uses tablets for field disinfection of water.
 Chlorine dioxide in gaseous form is used for area disinfection, most notably to kill
endospores of anthrax bacteria.

41. PHENOLS
• Obtained by distillation of coal tar
• Phenols are powerful microbicidal substances
• Phenolic derivatives have been widely used as
disinfectants for various purposes in hospitals
• Eg: Lysol, cresol

42. USES
• Various combinations are used in the control
of pyogenic cocci in surgical & neonatal units
in hospitals.
• Aqueous solutions are used in treatment of
wounds

43. • Colorless gas, available as cartridges
• Toxic and flammable, Odor similar to ether
• Has an extremely well penetration, even
through plastics
• Microorganism destruction is caused by a
chemical reaction
• Effective sterilization is dependent on
concentration of gas, exposure time,
temperature, and relative humidity
• Powerful sterilizer: Kills all known viruses,
bacteria (including spores), and fungi
Gaseous Disinfection
Ethylene Oxide Sterilization: EO Gas

44. USES
• Specially used for sterilising heart-lung
machines,respirators,sutures,dental
equipments, books and clothing.
• Also used to sterilise Glass, metal and paper
surfaces ,plastics, oil,some foods and tobacco.

45. FORMALDEHYDE GAS
• Widely employed for fumigation of operation
theatres, cath-lab, C.T, M.R.I and other rooms

46. BETA PROPIOLACTONE
• Used in fumigation
• For sterilisation 0.2% BPL is used
• Has a rapid biocidal activity
• Very effective against viruses

47. SURFACE ACTIVE AGENTS AND METALLIC SALTS
Substances which reduce the surface tension
–
Surface active agents

48. • Cations are widely used in the form of
quaternary ammonium compounds.
• Markedly bactericidal, active against Gram
positive organisms.
• No action on spores, tubercle bacilli, viruses

49. METALLIC SALTS
• The salts of silver, copper and mercury are
used as disinfectants.
• Act by coagulating proteins
• Marked bacteriostatic, weak bactericidal and
limited fungicidal activity

50. Heavy metals and their compounds
 Tiny amount of heavy metals (e.g. silver and copper) are effective antimicrobials. A
silver coin on an inoculated nutrient medium will inhibit growth for some distance.
 1% silver nitrate solution has been used to prevent gonorrheal ophthalmia
neonatorum, which the infants might have contracted as they passed through the
birth canal (recently been replaced by antibiotics).
 Silver-sulfadiazine is used in wound dressings. Available as topical cream for use on
burns.
 Mercuric chloride is highly bactericidal, but is toxic and corrosive and is inactivated
by organic matter. Organic mercury compounds such as Mercurochrome are less
irritating and less toxic than inorganic mercury.
 Copper sulfate is often used to destroy green algae in reservoirs or other water.
 Zinc chloride is used in mouthwashes, and zinc oxide is used in paints as antifungal.

51. Sterilisation by filtration
Filtration helps to remove bacteria from heat
labile liquids such as sera and solutions of
sugar, Antibiotics.
The following filters are used
Candle filtersCandle filters
Asbestos filtersAsbestos filters
Sintered glass filterSintered glass filter
Membrane filtersMembrane filters

52. CANDLE FILTER

53. CANDLE FILTERS
• Widely used for purification of water
Two types
(a) Unglazed ceramic filter – Chamberland
filter
(b) Diatomaceous earth filters – Berkefeld
filter

54. SEITZ FILTER
ASBESTOS DISCS

55. ASBESTOS FILTER
• Disposable single use discs
• High adsorbing tendency
• Carcinogenic
Eg: Seitz filter

56. SINTERED GLASS FILTER

57. SINTERED GLASS FILTER
• Prepared by heat fusing powdered glass
particles of graded size
• Cleaned easily, brittle, expensive.

58. MEMBRANE FILTER

59. MEMBRANE FILTERS
• Made of cellulose esters or other polymers
Uses
• Water purification & analysis
• Sterilization & sterility testing
• Preparation of solutions for parenteral use

61. RADIATION
Two types of radiations are used
NON –IONISINGNON –IONISING
IONISINGIONISING

63. Non- Ionising radiation:
• Electromagnetic rays with longer wavelength
• Absorbed as heat
• Can be considered as hot air sterilisation
• Used in rapid mass sterilisation of prepacked
Syringes and catheters
Eg: UV rays

64. IONISING RADIATIONS
• X- rays, gamma rays & cosmic rays.
• High penetrative power
• No appreciable increase in the temperature –
COLD STERILISATIONCOLD STERILISATION
• Sterilise plastics Syringes, catheters, grease
fabrics metal foils

66. MRI/CT SCAN/USG Suite
• m/c MRSA and VRE
• Infected wound & Spilling blood while using IV
Contrast media
• MRSA- 5%, survival-10-11days after contact
• VRE- 6-8%, 50% survival 7days

70. THANK YOUTHANK YOU