STERILIZATION AND DISINFECTION

1. Sterilization and disinfection
Presented By: Dr Md Mahbubul Hoda
Moderator: Dr Veerendra Kumar

2. CONTENT
Introduction
Definitions and basics of infection control
History
Most common infective agents in Maxillofacial regions.
Methods of sterilization and disinfection
a) By physical agents
B) By chemical agents
Design and disinfection of operation theater
Guidelines for hand scrub , gloves, masks, gowns and foot wear.
Waste management
References

3. Introduction
Significance-
Microorganism are ubiquitous. Since they cause
contamination, infection and decay, it becomes
necessary to remove or destroy them from operating
materials and from operating site.
Aims-
•In microbiology-
Prevents contamination by extraneous organisms.
•In surgery-
For maintaining asepsis.
•In food and drug manufacture
Ensuring safety from contaminating organisms

4. Definitions and basics of infection
control
Sterilization-
It’s a process by which an article, surface or medium
is made free of all microorganisms either in the
vegetative or spore form.
Disinfection –
The destruction or removal of all pathogenic
organisms, or organism capable of giving rise to
infection.( incapable of destroying spores)

5. Disinfectants are generally classified as follows-
High level disinfectants:
Chemicals that kill all microorganisms except bacterial
spores, when exposed for limited time.
Example - 2.4% glutaraldehyde,
7.5% hydrogen peroxide
Intermediate level disinfectants:
They kill mycobacteria, vegetative bacteria, most
viruses and most fungi but do not necessarily kill
bacterial spores.
Example- Phenol, alcohol .

6. Low level disinfectants:
Can kill most vegetative bacteria, some fungi and
some viruses. Commonly used detergents and soaps
act as low level disinfectants along with their
cleaning properties.

7. Instruments and different types of disinfection-
(Recommended by Spaulding)-
Critical items: These are instruments or objects that
are introduced directly into the bloodstream or into
other normally sterile areas of the body.
Absolute sterility at the time of use is required for
these items.
Example-
Surgical instruments, cardiac and urinary catheters,
implants, and needles

8. Semicritical items:
These instruments are introduced into body cavities
and therefore come into contact with intact mucous
membranes, but do not ordinarily penetrate body
surfaces.
Examples - non invasive flexible and rigid fiber-optic
endoscopes, endotracheal tubes, anesthesia
breathing circuits, and thermometer.
2% activated glutaraldehyde solution can be used for
about a fortnight. The equipment should be kept
immersed in glutaraldehyde solution for at least 45
minutes.

9. Non-critical items: These are items that do not
ordinarily penetrate, but touch only intact skin.
Example - crutches, bed boards, blood pressure
cuffs, and a variety of other medical accessories.
Simple washing with a detergent may be sufficient.

10. Antisepsis- The term used to indicate the prevention
of infection, usually by inhibiting the growth of the
bacteria in wounds or in tissue .
Antiseptics- chemical disinfectants which can be
safely applied to skin or mucous membrane and are
used to prevent infection by inhibiting the growth of
bacteria.
Decontamination- Refers to the process of rendering
an article or area free of danger from contaminants,,
including microbial, chemical, radioactive and other
hazards.

11. Infection control essentially consists of
-Cleaning
-Disinfection
-Sterilization of equipments and environment
-Hand hygiene practices
-Preoperative preparation of patient and antibiotic
prophylaxis

12. • Early civilization practised salting, smoking, pickling and
exposure to sunlight.
• Prior to the mid-19th century, gangrene set in before the
wound healed
• Repeated amputations performed in an effort to stop
the infection reaching the body, sometimes to no avail.
• The first reliable method of preventing infection during
an operation was developed by Dr. Joseph Lister
• Cleaning, sterilisation, disinfection are the cornerstones
of hospital infection prevention activities.
HISTORY

13. HISTORY
Louis Pasteur (1822-1895) a trained chemist of
France, known as father of microbiology introduce
sterilization techniques and development of steam
sterilizer, autoclave and hot air oven.
Joseph Lister(1827-1912) was a professor of surgery
in Glasgow Royal infirmary applied Pasture’s work
and introduce antiseptic techniques in surgery(1867)
effecting pronounced drop in mortality and
morbidity due to surgical sepsis, for this great work
he is known as father of antiseptic surgery.

14. • Cytomegalovirus
• Hepatitis B
• Hepatitis C
• Herpes simplex virus
• Human immunodeficiency virus
• Mycobacterium tuberculosis
• Staphylococci and streptococci
Organisms that are transmitted in a
dental setup

15. • PHYSICAL METHOD:
• Sunlight, heat (dry / moist)
• Filtration.
• Radiation.
• CHEMICAL:
• Alcohols.
• Aldehyde.
• Phenols.
• Halogens
• Salts.
• Surface active agents.
• Dyes.
• Gases.
METHODS

16. Physical agents
Sunlight:
-Sunlight having antimicrobial properties because of
ultraviolet radiation.
-It’s a natural source of sterilization .

17. Heat- Heat is considered to be most reliable method
of sterilization of articles that can withstand heat.
Heat acts by oxidative effects as well as denaturation
and coagulation of proteins.
Mainly two types of heats are used for sterilization
1) Dry heat
2) Moist heat

18. Dry heat- Acts by protein denaturation, oxidative damage
and toxic effects of elevated levels of electrolytes.
1) Red heat:
Articles such as bacteriological loops, straight wires, tips of
forceps and searing spatulas are sterilized by holding them
in Bunsen flame till they become red hot.
2) Flaming:
This is a method of passing the article over a Bunsen flame,
but not heating it to redness. Articles such as scalpels,
mouth of test tubes, flasks, glass slides and cover slips are
passed through the flame a few times. Even though most
vegetative cells are killed, there is no guarantee that spores
too would die on such short exposure.

19. Incineration:
This is a method of destroying contaminated
material by burning them in incinerator. Articles
such as soiled dressings; animal carcasses,
pathological material and bedding etc should be
subjected to incineration. This technique results in
the loss of the article, hence is suitable only for
those articles that have to be disposed.

20. 3) Hot air oven-
Introduced by Pasteur.
Holding period of 1600 C for one hour is used to
sterile
Glassware, forceps, scalpel, scissors, forceps, all glass
syringes, swabs, some pharmaceutical products.
It consists of a metallic chamber, walls contain
heating elements with fan for equal distribution of
hot air on objects

21. Holding time
60 minutes at 1600 C
40 minutes at 1700C
20 minutes at 1800C
Sterilization control-
Spores of non toxic strains clostridium tetani are
used as a microbiological test of dry heat efficiency
Brown tube
Thermocouples

22. Moist heat-
Moist heat acts by coagulation and denaturation of
proteins.
At temperature below 100 0C –
For pasturisation-
Holders method – 63 0 C for 30 minutes
Flash method- 720 C for 20 seconds
followed by rapid cooling to 13 0 c
By this method all non sporing pathogen such as
mycobacterium, brucellae and salmonellae are
destroyed.

23. At temperature 100 0c-
Boiling- Vegetative bacteria are killed almost
immediately at 90-100 0c for 10-30 minutes , but
sporing bacteria requires prolonged period of boiling.
Note –
Boiling is not recommended for sterilization, because
nothing short of autoclaving under pressure can
destroy spores and ensure sterilization

24. Steam at atmospheric pressure( 100 0c) –
An atmosphere of free steam is used to sterilize
culture media which may decompose if subjected to
higher temperature.
Koch or Arnold steamer is usually used.

25. Steam under pressure ( autoclave)-
Principle-
Water boils when its vapour pressure is equals to
surrounding atmospheric pressure, Hence when
pressure inside a closed chamber increases, the
temperature at which water boils also increases.
Saturated steam has penetration power, and when
this saturated steam comes into contact with cooler
surface it condenses to water and gives up its latent
heat to that surface.

26. Equation says –
1600ml steam at 100 0 c condenses into 1ml of
water and releases 518 calories of heat.
This large reduction of volume sucks in more steam
to the area and the process continues till the
temperature of the surface is raised to that of
steam.
Temperature(0C) Holding time (minutes)
121 15 (15 lbs)
126 10 (20 lbs)
134 03 (30 lbs)

27. Materials can be sterilized by autoclave –
Dressing, Lab ware, glassware, pharmaceutical
products, metal instruments, surgical instruments,
cultural media, disposable and non disposable
syringes.
Sterilization control-
Bacillus steriothermophillus are used
Chemical indicators
Autoclave tapes
Thermocouples

28. Filtration-
Filtration helps to remove bacteria from heat labile
liquids such as sera and solutions of sugars or
antibiotics used for culture media.
The following types of filters have been used-
1) Candle filter- this type of filter used widely for
purification of water for industrial and drinking
purpose.
its of two types
a) Unglazed ceramic filters
b) Diatomaceous earth filters

29. 2) Asbestos filters-
Are disposable, single use discs. They have high
adsorbing capacity and tend to alkalinize filtered
liquids, the carcinogenic potential of asbestos has
discouraged their use
3) Sintered glass filters-
Prepared by heat fusing finely powdered glass
particles of graded sizes. They have low absorptive
property and can be cleaned easily but are brittle
and expensive.

30. 4) Membrane filter-
Made of cellulose esters or other polymers have
largely replaced other types of filters.
They are routinely used in water purification and
sterilization and sterility testing and for the
preparation of solution of parenteral use.

31. Radiation-
Two types of radiation are used for sterilization
1) Nonionising- Infrared and Ultraviolet rays
( low energy type)
2) Ionising – gamma rays and high energy electron
( high energy type)

32. 1) Nonionising radiation –
Here electromagnetic rays with wavelength
longer than those of visible light are used. These are
to a larger extent absorbed as heat, hence infrared
radiation can be considered as a form of hot air
sterilization
Used for –
Rapid mass sterilization of prepacked items such as
syringes and catheters.

33. While ultraviolet radiation is used for disinfecting
enclosed areas such as entry ways, operation
theaters and laboratories.
2) Ionization radiation-
X-ray, gamma rays and cosmic rays are highly lethal
to DNA and other vital constituents. They have very
high penetrative power, since there is no appreciable
increase in temperature in this method, this is
referred to as Cold sterilization
Uses- items like plastic syringes, swabs, catheters,
oils and metal foils sterilization .

34. Chemical agents
The ideal properties of chemical agent should have –
-Effective against all microorganisms and have wide spectrum of
activity.
-Have speedy action.
-Effective in the presence of organic matter.
-Be stable.
-Be effective in varying pH.
-Compatible with other antiseptics.
-Have high penetrating power.
-Not corrode metals.
-No cause irritation.
-Not interfere with healing.
-Not to be toxic if absorbed into circulation.
-Be cheap and safe.
-Easy available.

35. Chemical agents acts in various ways to micro-
organisms-
1) Protein coagulation
2) Disruption of cell membrane resulting in
exposure, damage or loss of the contents
3) Removal of the free sulphydryl groups essential
for the functioning of the enzymes.
4) Interruption of cellular metabolism by acting as
substrate competitors.

36. Alcohol-
Ethyl alcohol and isopropyl alcohol are the most frequently
used .
-Denaturating bacterial proteins (against vegetative bacteria),
disrupt cytoplasmic membrane, dissolves lipids.
-Rapidly kill T.B bacilli.
-No sporicidal / virucidal.
-Skin antiseptic (swabbing of skin prior to injection)
Methyl alcohol –
Against fungal spores, toxic and inflammable.

37. Uses-
Rubber stoppers , thermometers, stethoscope,
scissors, ventilation bags, external surface of
ventilators, electrical equipment's.

38. Aldehydes –
There are mainly two types of aldehyde
1) Formaldehyde-
-Active against the amino group in the protein,
inactivates nucleic acid.
- Bactericidal, sporicidal, and virucidal.
- Used both in aqueous form and gaseous form.
- liquid Formaldehyde used to preserve
anatomical specimens, and destroying.
- Gas is used for sterilizing instruments and heat
sensitive catheters and for fumigating wards

39. FORMALDEHYDE (40% = formalin): to preserve
anatomical specimen.
10% +0.5% sodium tetra borate is used to sterilize
metal instruments.

40. Glutaraldehyde:-
-Similar action like formaldehyde
-Effective against pseudomonas, tubercle bacillus,
fungi and viruses.
-Activated gluteraldehyde 2% is rapid acting, non-
staining, and rust inhibiting.
-For complete disinfection: immersed for 10 minutes
to destroy vegetative pathogens.

41. For sterilization: Immersed for 10 hours at ph 7.5 to
8.5 to destroy pathogenic spores.
-Plastic tubes, metal, polythene tubes, platic
endotracheal tube, bronchoscope.

42. Dyes –
Aniline and Acridine dyes used extensively as skin
and wound antiseptics.
These dyes are more active against gram positive
than gram negative organism .
No activity seen against tubercle bacilli and hence
used in Lowenstein Jensen medium.
They shows lethal effect on bacteria by acting on
acidic group.

43. More advanced dyes are Proflavine, Acriflavine,
Evaflavine.
They slowly release and shows prolonged action on
organisms.
Acting by impairing the DNA complexes of organism
and thus inhibit the multiplication or micro-
organisms.

44. Chlorine is used in water supplies, swimming, food /
dairy. Bleaching powder, sodium hypochlorite and
chloramines are also used (release of free chlorine
 react with water  hypochlorous acid). (oxidation
/ denature).
Per-acetic acid – sporicidal used to sterilize
equipment.
Disinfectant for HIV infected material.
Bactericidal, fungicidal, virucidal, sporicidal

45. IODINE: ( 5-10%)
Iodine molecules rapidly penetrate the cell wall of
microorganisms and inactivate cells by forming
complexes with amino acids and unsaturated fatty
acids, resulting in impaired protein synthesis and
alteration of cell membranes.
2% iodine in 50% alcohol (Tincture)
Iodine with surface active agents called iodophores
= BETADINE.

46. They have bactericidal activity against gram
positive, gram negative, and certain spore forming
bacteria (clostridia and bacillus spp) and also active
against mycobacteria, viruses and fungi.

47. Hydrogen peroxide:
Damages proteins and DNA of micro-organsim
Used at 6%, to decontaminate the instruments,
equipments.
3% is used for skin disinfection and deodorising
wounds and ulcer.
Strong solutions are sporicidal.

48. Phenols-
These are obtained by distillisation of coal tar
between temperature of 170 0C and 270 0C.
Lister the father of antiseptic surgery, 1st introduced
their use in surgery 1865.
Mechanism of action is damage cell membrane and
releasing cell contents and causing lysis of cell.
Derivatives of phenol are cresol, lysol, chlorhexidine
, chloroxylenol, and hexachlorophane.

49. CHLORHEXIDINE: (savlon) chlorhexidine gluconate is a
commonly used agent and has been incorporated into
number of hand hygiene preparation also.
It is a cationic bisguanide and acts by disruption of
cytoplasmic membrane, resulting in precipitation of
celllular contents.
It has good activity against gram positve bacteria, some
what gram negative bacteria and fungi, and only limited
activity against tubercle bacilli.
The activity of chlorhexidine minimally affected in
presence of organic material such as blood

50. HEXACHLOROPHANE:
More against gram positive. Applied over the skin as
prophylaxis against staphylococcal. Potentially toxic.
CRESOLS –
used for infected glass wares, cleaning floors. Not
inactivated by presence of organic matter.

51. Gases
Ethylene oxide-
This is a colourless liquid with a boiling point of
10.70C
At normal temperature and pressure is highly
penetrating gas with a sweet ethereal smell.
Highly inflammable and in concentration in air
greater than 3% highly explosive, by mixing it with
carbon dioxide or nitrogen , to a concentration of
10%, its explosive tendency is eliminated

52. Its action is due to alkylating the amiino, carboxyl,
hydroxyl and sulphydryl groups in protein
molecules, in addition it acts on DNA and RNA too.
Its active against all types of microorganisms
including viruses and spores.
ETO gas sterilization is more expensive and complex
process of sterilization , hence it is usually restricted
to objects that might be damaged by heat or
excessive moisture.
Bacillus subtilis niger used to check sterilization
completion .

53. It diffuses through many types of porous materials
and readily penetrates some plastics. It is specially
used for heart-lung machines, respirators, sutures
and dental equipments.
It is unsuitable for fumigating the wards or room
because of its explosive properties.

54. Formaldehyde-
Widely used for fumigating wards and rooms and
operation theaters.
After sealing the windows and other outlets,
formaldehyde gas is generated by adding 150g of
KMnO4 to 280ml formalin for every 1000cu ft of
room volume .
After starting generation of vapour the door should
be sealed for 48 hours.

55. Betapropiolactone (BPL)
This is a condensation product of ketane and
formaldehyde with boiling point 163 0C.
Though its having low penetrating power, It is said to
be more efficient for fumigating purpose than
formaldehyde.
Its having rapid biocidal action but unfortunately has
carcinogenic activity.
0.2% BPL is used for sterilization, is capable to kill all
microorganism and very active against viruses.

56. Surface active agents
Substances which alter energy relationship at
interfaces, producing a reduction of surface or
interfacial tension are referred to as surface active
agents
Used as wetting agents, detergents and emulsifiers.
They are classified into four main group -
Anionic, cationic, nonionic, amphoteric.

57. Cationic form shows most wide range of
antibacterial action.
Acts on the phosphate group of the cell membrane
and also enter the cell. The membrane loses its semi
permeability and the cell proteins are denatured.
These are markedly bactericidal being active against
gram positive and less active against gram negative
bacteria.
They have no action against spores, tubercle bacilli
and most of the viruses

58. The common compound are
Acetyl trimethyl ammonium bromide ( cetavlon and
cetrimide)
The anionic compound, example is common soap,
have moderate action, these are more active against
gram negative organisms than gram positive.
Amphoteric compounds known as Tego compound
are active against a wide range of gram positive and
gram negative organisms including viruses.

59. Metallic salts
Though all salts have some germicidal action
depending to their concentration.
Salts of silver, mercury, copper are used as
disinfectants.
They are protein coagulants and have the capacity to
combine with free sulphydryl groups of cell enzymes.
They shows bacteriostatic actions on topical use.

60. Design and disinfection of
operation theater
There are so many micro-organisms found in
operation theater environment including bacteria,
viruses and fungi which can cause significant
nosochomial infection.
To avoid this it is required to maintain operation
theater clean and disinfected at least.
Significant risk zone –
700-1800 bacteria carrying particle per cubic meter

61. The operation theater is merely a clean environment
but not sterile in which the concentration of
airborne particle is controlled and which is
constructed and used in a manner to minimize the
introduction, generation and retention of particle
inside the room and in which other relevant
parameters are under control.
Example –
Temperature, humidity, and pressure are under
controlled as necessary.

62. Parameter Desired range-
Temperature- 20–23°C
Relative humidity- 30–60%
Air movement- From clean to less clean areas
Air changes - Minimum 15 total air changes per
hour

63. Criteria -
The operating rooms are set away from hospital by
two sets of door.
People are required to remove their street cloths and
done scrub suits before entering the operation room
area.
Mask, gowns, gloves and special shoes worn during
the operation.
The patient and operating site are draped and
isolated.

64. The surgical instruments and related instruments
should be sterile.
After entering the operation theater and before
gowning, personnel should take precaution to avoid
contamination the open packs of draping material.
Once the patient is prepared and draped, only those
who are scrubbed, gowned, and gloved may work in
surgical site.
The back of gowned are considered as unsterile as
below waist

65. Chemical agents used as disinfectant in OT
Size – 10×10 ft ( 1000 cubic ft)
Option 1 -Fogging by using agents like hydrogen
peroxide 8–10%, hydrogen peroxide 4–6% with
silver nitrate.
-Theatre can be used after 20–30 minutes
-No irritation, no toxicity issues.

66. Option 2 -
Formalin 200 ml + KMnO4 after adequate cleaning
and mopping.
Close the operation theatre for a period ofat least 8
hours. Then introduce liquor ammonia 200 ml and
allow a contact time of minimum 2 hours for
neutralization of formaldehyde vapors. Switch on
the
AC, ideally split AC, for a period of 1 hour.
Now the operation theatre is ready for use.

67. Guidelines for hand scrub , gloves,
masks, gowns and foot wear.
Hand washing guidelines
Hand washing removes contamination and decreases
the natural bacterial load.
1. It should be done before significant contact with
patient or activity likely to cause contamination,
such as in intensive care units, while caring for
immunocompromised patients.
2. For routine hand wash, neutral soap is adequate.
Preferably, liquid soaps in disposable dispensers with
dispensing nozzle should be used.

68. 3. Prior to surgical procedures, 4% chlorhexidine or
povidone iodine 0.75% is preferred. Manufacturer’s
instructions should be referred to.
Use a brush to thoroughly scrub up to elbows,
followed by rinsing under running water. Wash on all
sides of hands and above wrist,vigorously. Nails
should be specifically cleaned. Rinse under running
water.
4. The hands should be dried using a sterile towel.

69. 5. During procedures wherein multiple handwashes
are needed, alcohol-based hand rubs may be used
as an adjunct to conventional handwash.
6. Cuts/abrasions should be covered adequately by
water-resistant, occlusive dressings, which should
be changed if they become wet/soiled

70. Gloves
1. Gloves should be used in all procedures wherein
the tissue in contact should be maintained
sterile. They are not necessary while
adminstering injections.
2. For instrument cleaning, and decontamination
procedures, general purpose household gloves are
adequate.
3. Gloves should not be washed and reused.

71. 5. They should be changed if they are
torn,punctured, or after contact with nonsterile
objects.
6. Hands should be washed after removal of gloves

72. Masks, face shields, eye wear
These are needed in procedures where splashing,
spraying blood/body fluids are expected.
Manufacturer’s instructions should be followed. Filter
mask (for 0.3 mm particles) should be used for laser
plumes.

73. Gowns
A disposable or nondisposable gown/apron made of
impervious, fluid resistant material should be used
where there is a likelihood of contamination or
splashes with blood/body fluids

74. Foot wear
Enclosed foot wear, disposable/reusable, should be
used to protect from injury from sharps or contact
with body fluids.

75. Waste management
Yellow
Plastic Bag
Human and animal waste
Eg: Tooth, tissue
Solid waste
Eg: Cotton,plaster and gauze

76. Red
Disinfected container/ plastic bag
Microbiology and biotechnology waste
Solid waste (disposal items other than sharps and
tubing catheters, intravenous sets etc.)

77. Blue/ White translucent
Plastic bag/ Puncture proof container
Waste sharps
Eg: needles, blades.

78. Black
Plastic bags
Discarded medicines and cytotoxic drugs
Eg: outdated drugs and medicines
Inceneration ash
Chemical waste
Liquid waste
Eg: cleaning and disinfecting material

79. THANK YOU

80. References
Sterilization and disinfection – Annanthnarayan
Essentials of microbiology – Chakaraborthy
Hospital acquired infections – Purva Mathur
Sterilisation and disinfection – Laskins vol1
Narendra patwardhan, Uday Kelkar: Disinfection,
Sterilization and Operation theater guidelines for
dermatosurgical preactitinor in india; Indian journal of
Dermatology, Venereology 2011 (Vol 77)