Overview of various methods of sterilisation and disinfection.

1. Sterilization
and
Disinfection
Dr. Bharat Paul

2. Contents
• Historical background
• Introduction
• Factors Affecting Disinfection
• Types of disinfection
• Methods Of Disinfection & Sterilization
– Natural Methods
– Physical Methods
– Chemical Methods
• Some practical aspects
• CSSD
• References

3. Historical background
• The scientific use of disinfection and
sterilization methods originated more than 100
years ago.
• Ignatz Semmelweis(1816-1865) and Joseph
Lister(1827-1912) -important pioneers for the
promotion of infection control.

4. Historical background
• More than 100 years ago, Semmelweis
demonstrated that routine hand washing can
prevent the spread of disease.
• He worked in a hospital in Vienna where
maternity patients were dying at an alarming
rate.
• He recognized that medical students worked
on cadavers during an anatomy class and
afterwards they went to the maternity ward.

5. Historical background
• Students did not wash their hands between
touching the dead and the living!!!
• After administrating the hand washing before
examining the maternity patients the mortality
rate decreased.

6. Historical background
• Lister, for the first time, used carbolic
acid in operating theatres that significantly
reduced mortality rates.
• In 1867, Lister introduced the use of phenol as
antimicrobial agent for surgical wound
dressings.
• His principles were gradually adopted in
Britain and later in US.
• This was the beginning of infection control.

7. Introduction
Sterilization-
• Destruction of all forms of life, including the
bacterial spores, viruses, prions
• No degrees of sterilization: an all-or-nothing
process.
• Physical or chemical methods.

8. Introduction
Disinfection -
• Less lethal than sterilization. Destroys most
recognized pathogenic organisms but not
necessarily all microbes (e.g. Spores)
• Purpose-prevent transmission of certain
microorganisms with objects, hands or skin
and prevent spreading the infection.
• Physical or chemical methods.

9. Introduction
• Disinfectant – usually a chemical agent that
destroys disease causing pathogens or other
harmful microorganisms, but might not kill
bacterial spores. Refers to substances applied
to inanimate objects.
• Antiseptic – Substance that prevents or arrests
the growth of microorganisms. Preparations
applied topically to living tissues.

10. Introduction
• Asepsis- prevention of contact with
microorganisms.
• Sanitizer – Agent that reduces the number of
bacterial contaminants to safe levels.
• Sterile – State of being free from all
microorganisms.

11. Introduction
• Detergent – Surface cleaning agent that makes
no antimicrobial claims. Acts by lowering
surface tension.
• Cleaning – Removal of adherent visible soil,
blood , and other debris from surfaces ,
crevices , serrations and lumens of
instruments.

12. Factors that influence the degree of
killing
• Types of organisms
• Number of organisms
• Concentration of disinfecting agent
• Presence of organic material (e.g., serum, blood)
• Nature (composition) of surface to be disinfected
• Contact time
• Temperature
• pH
• Biofilms
• Compatibility of disinfectants and sterilants

13. Factors that influence the degree of
killing
Type of organisms
• Organisms vary in their ability to withstand
chemical and physical treatment, e.g.,
• Spores–have coats rich in proteins, lipids and
carbohydrates
• Mycobacteria–cell walls are rich in lipids
• Prions–the most resistant known organisms to
the action of heat, chemicals, and radiation !!!

14. Factors that influence the degree of
killing

15. Factors that influence the degree of
killing
Number of organisms
• Microbial load-the total number of organisms
which determine the exposure time of killing
agent
• organisms have varying degrees of susceptibility
to killing agents
• not all organisms die at the same time
• the death curve is logarithmic!!!
• higher numbers of organisms require longer
exposure.

16. Factors that influence the degree of
killing

17. Factors that influence the degree of
killing
Concentration of disinfecting agent
• a proper concentration of disinfecting agents
ensure the killing of target organisms, e.g.,
• povidone-iodine should be diluted with water
before use because there is not enough free
iodine to kill microorganisms in concentrated
solution.

18. Factors that influence the degree of
killing
• Presence of organic material(such as blood,
mucus, pus) affects killing activity by
inactivating the disinfecting agent, e.g,
• by coating the surface to be treated, prevents
full contact between object and agent
(Glutaraldehyde)
• For optimal killing activity, instruments and
surfaces should be cleansed of excess organic
material before disinfection !!!

19. Factors that influence the degree of
killing
• Nature (composition) of surface to be
disinfected
• some medical instruments are manufactured of
biomaterials that exclude the use of certain
disinfection and sterilization methods because
of possible damage, e.g.,
• endoscopic instruments cannot be sterilized by
the heat in an autoclave.

20. Factors that influence the degree of
killing
Contact time
• the amount of time a disinfectant or sterilant is in
contact with the object is critical e.g.,
• Betadine must be in contact with object for at
least 1 to 2 min to kill microbes.
• the spores of bacteria and fungi need a much
longer time
• determine whether it is disinfecting or sterilizing
the object.

21. Factors that influence the degree of
killing
Biofilms
• communities of microorganisms.
• can be on a surface of either inanimate or
animate objects, e.g, catheters , pipes that carry
water.
• make disinfection more difficult.
• the concentration of the disinfectant and the
contact time need to be increased.

22. Factors that influence the degree of
killing
Compatibility of disinfectants
• a common mistake is to believe that two
disinfectants are better than one !
• some of them may inactivate other, e.g.,
• the bleach and quaternary ammonium
compound together negate the activity of both.

23. Ideal disinfectant
• Highly efficacious
• Fast
• Good penetration
• Compatible with most materials
• Non-toxic
• Effective despite presence of organic material.
• Residual effect
• Economical
• Odourless

24. Types of disinfection
• Concurrent disinfection –
– Application of disinfective measures as soon as
possible after the discharge of infectious material
from the body of an infectious person.
– Consists of disinfection of urine ,faeces , vomit,
clothes ,hands ,dressings, apron ,gloves.
• Terminal disinfection -
– Application of disinfective measures after the
patient has died or has ceased to be a source of
infection.

25. Types of disinfection
• Precurrent (prophylactic) disinfection –
Disinfection of water by chlorine ,
pasteurization of milk ,hand washing.

26. Types of disinfectants
High-level disinfectants
• activity against bacterial spores
Intermediate-level disinfectants
• tuberculocidal activity but not sporocidal
Low-level disinfectants
• a wide range of activity against
microorganisms but no sporocidal or
tuberculocidal activity

27. Types of materials in healthcare
Critical materials
• invade sterile tissues or enter the vascular
system.
• most likely to produce infection if
contaminated and therefore require
sterilization.
• E.g. Surgical instruments , cardiac catheters

28. Types of materials in healthcare
Semicriticial materials
• have contact with mucous membranes
• require high-level disinfection agents
• E.g.cystoscope ,laryngoscope blade.
Noncriticial materials
• have contact with intact skin
• require intermediate-level to low-level
disinfection
• E.g. bedpans , BP Cuffs

29. Methods of disinfection
• Natural methods
• Physical methods
• Chemical methods

30. Natural agents
• Sunlight –
– UV rays of sunlight lethal to bacteria and some
viruses.
– Articles such as linen, bedding and furniture may
be disinfected.
• Air
– Acts by drying or evaporation of moisture which is
lethal to most bacteria.

31. Physical agents
• Burning
– Inexpensive articles such as contaminated
dressings , rags and swabs – disposed off by
burning.
– Best done in an incinerator.
– Addition of sawdust, paper ,kerosene or other
combustible material aid in burning.

32. Physical agents
• Hot air
– Articles such as glassware
,syringes , swabs , dressings
and sharp instruments.
– Drawback – no penetrating
power: not suitable for bulky
materials.
– Hot air oven – temp 160-180
deg C for at least 1hour to kill
spores.Hot air oven

33. Physical agents
• Thermal Death Point (TDP): Lowest
temperature at which all of the microbes in a
liquid suspension will be killed in ten minutes.
• Thermal Death Time (TDT): Minimal length of
time in which all bacteria will be killed at a given
temperature.
• Decimal Reduction Time (DRT): Time in
minutes at which 90% of bacteria at a given
temperature will be killed. Used in canning
industry.

34. Physical agents
MOIST HEAT
• Kills by denaturing proteins and destroying
cytoplasmic membranes.
• More effective than dry heat; water better
conductor of heat than air.
• Methods of microbial control using moist heat
– Boiling
– Autoclaving
– Pasteurization

35. Physical agents
• Boiling
– for 5-10 minutes :kill bacteria but not spores or
viruses.
– Temp. above 100 deg C required for destruction of
spores.
– Disinfection of small instruments , tools not used
for subcutaneous insertion , linen and rubber
goods such as gloves.
– Slow process , unsuitable for thick beddings and it
fixes albuminous stains.

36. Physical agents
• Autoclaving
–Operate at high temperature and pressure.
–Destroys all forms of life , including spores.
–Sterilization of linen , dressings , gloves ,
syringes , instruments and culture media.
–Not suitable for plastics and sharp
instruments.

37. Autoclave

38. Physical agents
Pasteurization
• A process in which fluids are heated at
temperatures below boiling point to kill
pathogenic microorganisms in the vegetative
state without altering the fluid’s palatability.
• Conditions: 62℃, 30min or 71.7℃, 15sec
• Significance: kills vegetative pathogens.

39. Pasteurisation

40. Physical agents
• Radiation
–Sterilization of bandages , dressings , catgut
and surgical instruments.
–Great penetrating power with little or no
heating effect.
–Most effective ,but very costly.
–Normally carried out by gamma radiation.

41. Physical agents
• Filtration: Removal of microbes by passage of a
liquid or gas through a screen like material with
small pores. Used to sterilize heat sensitive
materials like vaccines, enzymes, antibiotics, and
some culture media.
• 0.22 and 0.45um Pores: Used to filter most
bacteria. Don’t retain spirochetes, mycoplasma
and viruses.
• 0.01 um Pores: Retain all viruses and some large
proteins

42. Chemical agents
Phenol and related compounds
• Intermediate- to low-level disinfectants
• Denature proteins and disrupt cell membranes
• Effective in presence of organic matter and
remain active for prolonged time.
• Commonly used in health care settings, labs, and
homes (Lysol, triclosan)
• Have disagreeable odor and possible side effects.

43. Chemical agents
• Phenol(carbolic acid) – not an effective
disinfectant
• Crude phenol- phenol and cresol. effective
against gram +ve and –ve organisms.
• Cresol – used in 5 to10 % for disinfection of
urine and faeces.
• Cresol emulsions – very powerful
disinfectants

44. Chemical agents
• Chlorhexidine (hibitane ) – most useful skin
antiseptic. 0.5% solution used as
handlotion.1% creams used for burns
• Hexachlorphane – slow in action , but shows
cumulative effect on skin and is compatible
with soaps.
• Dettol (chloroxylenol) – relatively non toxic
antiseptic.5 % dettol for disinfection of
instruments and plastic equipments.

45. Chemical agents
Quaternary ammonium compounds
– Cetrimide(cetavlon)
– bactericidal against vegetative gram +ve
organisms , much less against gram -ve
– Used in 1-2 % strength
– Savlon
– Combination of cetavlon and hibitane .
– Plastic appliances may be disinfected in 20 min.
– Disinfection of clinical thermometers in 3 minutes.

46. Chemical agents
Halogens and their compounds
• Intermediate-level antimicrobial chemicals
• Believed that they damage enzymes via oxidation
or by denaturing them.
• Iodine tablets, iodophores (Betadine®), chlorine
treatment of drinking water, bleach, chloramines
in wound dressings, and bromine disinfection of
hot tubs.

47. Chemical agents
Alcohols
• Intermediate-level disinfectants.
• Denature proteins and disrupt cytoplasmic
membranes.
• Evaporate rapidly.
• Swabbing of skin with 70% ethanol prior to
injection.

48. Chemical agents
Isopropyl Alcohol (70%)
• Powerful disinfectant and antiseptic
• Mode of action: denatures proteins, dissolves
lipids and can lead to cell membrane
disintegration
• Effectively kills bacteria and fungi
• But does not inactivate spores!

49. Chemical agents
Aldehydes
• Denature proteins and inactivate nucleic
acids.
• Glutaraldehyde both disinfects (short
exposure) and sterilizes (long exposure)
• Formalin used in embalming and disinfection
of rooms and instruments.

50. Chemical agents
Formaldehyde
–Highly toxic and irritant gas
–Effective against vegetative bacteria ,fungi
and many viruses.
–Used as a 2-3 % solution for spraying rooms
, walls and furniture
–Most effective at a high temperature and
relative humidity of 80-90%

51. Chemical agents
Oxidizing agents
• Peroxides, ozone, and per acetic
acid kill by oxidation of microbial enzymes
• High-level disinfectants and antiseptics
• Hydrogen peroxide can disinfect and sterilize
surfaces of objects.
• Ozone -treatment of drinking water.
• Per acetic acid – effective sporocide used to
sterilize equipment.

52. Chemical agents
Surfactants
• Surface active chemicals that reduce surface
tension of solvents to make them more effective
at dissolving solutes
• Soaps and detergents
• Colorless, tasteless, harmless to humans, and
antimicrobial; ideal for many medical and
industrial uses.
• Low-level disinfectants.

53. Chemical agents
Metals-
• Ions are antimicrobial because they alter the 3-D
shape of proteins, inhibiting or eliminating their
function.
• Low-level bacteriostatic and fungistatic agents.
• 1% silver nitrate to prevent blindness caused by
N. gonorrhoeae .
• Thiomersal (mercury-containing compound) used
to preserve vaccines.

54. Chemical agents
Lime
–Cheapest disinfectant
–Used as fresh quick lime or 10-20 %
aqueous suspension known as milk of lime.
–Faeces and urine disinfected by 10-20%
aqueous solution (in 2 hrs).
–Used as a deodorant in public places where
toilets are located.

55. Chemical agents
Ethylene oxide
– for heat sensitive articles
– At 55- 60 deg C : kills bacteria ,spores and viruses.
– Mixed with 12 % carbon dioxide (as it is
explosive)
– Water vapor added to the mixture to increase
efficacy.
– To sterilize fabrics, plastic equipment , cardiac
catheters , etc.

56. Some Newer agents
• Bacillocid® rasant
– Formaldehyde-free disinfectant cleaner with low
use concentration
• Virkon
– gaining importance as non Aldehyde compound
– Disinfects medical devices and lab equipments.
– Decontaminate spillages with Blood and body
fluids.

57. Gas plasma sterilization
• Gas plasma generated in an enclosed chamber
under deep vacuum using Radio frequency or
Microwave.
• Can be used for hand sterilization.
• The mechanism of action of this device is the
production of free radicals within a plasma
field that are capable of interacting with
essential cell components

58. Sterilization Control
• To ensure that potentially infectious agents are
destroyed by adequate sterilization regimes
• Three levels:
• physical: measuring device control (temp.,
time, pressure)
• Biological
• Chemical

59. Sterilization Control
Biological:
• Bacillus stearothermophilus spores(104-106
organisms)
• survives steam heat at 121ºC for 5 min. and is
killed at 121ºC in 13 min.
• validate and determine the adequacy of steam or
chemical sterilization.
• Bacillus subtilis/B.atrophaeus spores
• validate and determine the adequacy of ethylene
oxide or dry heat sterilization.

60. CHEMICAL INDICATORS
• Browne's tubes are glass tubes that contain
heat sensitive dyes. These change color after
sufficient time at the desired temperature.
• Before heat exposure, the contents of the tube
appear red.
• As heating progresses, the color changes to
green.

61. CHEMICAL INDICATORS
•Bowie Dick tape
is applied to
articles being
autoclaved. Before
heat exposure, the
tape is uniformly
buff in color.
•After adequate
heating, the tape
develops dark
brown stripes.
The pack on the left has been properly
sterilized; that on the right has not.

62. SOME PRACTICAL ASPECTS
Disinfection of faeces & urine-
• Collected in impervious container.
• Add equal volume of disinfectant for 1-2 hrs.
Disinfectant Amount per litre Percent
Bleaching
powder
50 gms 5
Crude phenol 100ml 10
Cresol 50 ml 5
Formalin 100 ml 10

63. Disinfection of faeces & urine-
• Quick lime or milk of lime can be used if
above disinfectants not available.
• Bedpans and urinals – steam disinfected.

64. Disinfection of sputum
• Received in gauze or paper handkerchief .
• Destroyed by burning.
• Disinfected by boiling or autoclaving for 20
min at 20 lbs pressure.
• Or 5 % cresol can be used.

65. • Disinfection of blood/body fluids spills
– Clean the affected area with soap and water then
disinfect with a 1%Na hypochlorite solution for 15
minutes.
• HIV infected articles
– Cleaning and 2% glutaraldehyde for 30 minutes.
– 6% Hydrogen peroxide for 30 minutes.
– 1% hypochlorite solution for 30 minutes.

66. Fumigation
• To sterilize the operation theatre formaldehyde
gas (bactericidal & sporicidal, viricidal) is
widely employed as it is cheaper for
sterilization of huge areas like operation
theatres.
• Formaldehyde is irritant to eye & nose; and it
has been recognized as a potential carcinogen.

67. References
• K.Park Textbook of Preventive and Social
medicine 23rd edition.
• CDC – Sterilization and disinfection
guidelines
• Hospital hygiene and infection control –WHO.
• WHO Pharmacoepia Library- Methods of
Sterilization.
• Occupation Safety and Health administration-
Disinfection guidelines.