sterilization basics

1. STERILIZATION AND
DISINFECTION

2. CONTENTS
• DEFINITIONS
• STERILIZING AGENTS
PHYSICAL AGENTS
CHEMICAL AGENTS
• CONCLUSION
• REFERENCES

3. DEFINITIONS
 STERILIZATION - The process by which an article, surface or
medium is freed of all micro-organisms either in the vegetative
or spore state.
 DISINFECTION - Destruction of all the pathogenic organisms or
organisms capable of giving rise to infection but not necessarily
bacterial spores.
 ANTISEPTICS - Chemical disinfectants which can safely applied
to living tissues and are used to prevent infection by inhibiting
the growth of microorganisms.

4.  ASEPSIS - Technique by which the occurrence of infection into
an uninfected tissue is prevented.
 DECONTAMINATION – Process of rendering an area or article
free of danger from contaminants, including microbial,
chemical, radioactive hazard.
 BACTERICIDAL - Agents which are able to kill the micro-
organisms.
 BACTERIOSTATIC - Agents which only prevent the multiplication
of bacteria.
 CLEANING – The process which removes visible contamination,
but does not necessarily destroy microorganisms.

5. STERILIZING
METHODS
• Physical agents
• Chemical agents

6. 1. Sunlight
2. Heat
a. Dry heat
b. Moist heat
4. Filtration
5. Radiation
6. Sonic & Ultrasonic
vibration
PHYSICAL
AGENTS
CHEMICAL
AGENTS
1. Alcohols
2. Aldehydes
3. Dyes
4. Halogens
5. Phenols
6. Surface active agents
7. Metallic salts
8. Gases

7. SUNLIGHT
 Appreciable BACTERICIDAL activity.
 Action by combined effect of UV rays and HEAT rays.
 Recognised pathways :
 Direct - UVB rays damage nucleic acid prevents
microbial
replication.
 Natural method of disinfection of water in tanks, rivers and lakes.

8. HEAT
• Most RELIABLE method of sterilization of articles.
• Factors influencing sterilization by heat:
1. Nature of heat: Moist heat/Dry heat.
2. Temperature and time: Inversely proportional.
3. Number of microorganisms.
4. Nature of microorganism: Spores are highly resistant to
heat.
5. Type of material.
6. Presence of organic material.

9. MOA
• Protein denaturation
• Oxidative damage
• High electrolyte level toxicity
1. DRY HEAT
Good penetrability and non-corrosive nature of dry
heat makes it applicable for sterilizing glass wares and
metal surgical instruments.
Destroys bacterial endotoxins (or pyrogens) which
makes it applicable for sterilizing glass bottles.

10. a. Red Hot
• Held in a Bunsen flame until red
hot along their whole length,
almost up to the tip of their
metal holder.
• Items sterilized by red heat –
 Inoculating wire loops.
 Points of forceps.
 Needles.

11. Only exposed to the
flame for a few seconds
& not heating to red hot.
Scalpel blades
glass slides
mouth of culture
tubes, flasks.
b. Flaming

12. • Excellent & safe method.
• Material destroyed completely –
reduced to ashes.
• Used for – Pathological wastes,
surgical dressing, animal carcasses,
plastics like PVC/ polythene.
• Avoid – Materials made of polystyrene
as they emit black dense smoke.
c. Incineration

13. • Most widely used method by dry
heat
• Glass-wares - glass syringes, petri
dishes, flasks, pipettes and test
tubes.
• Surgical instruments like Scalpels,
scissors, forceps.
• Chemicals - liquid paraffin, fats,
sulphonamides powders, glycerol
d. Hot air oven

14. • Heating elements fitted in chamber
– fans for even distribution of heat.
• Thermostat – maintains the chamber
air at a chosen temperature.
• TEMPERATURE:
160˚C – 2 hours.
170˚C – 60 minutes.
180˚C – 30 minutes.
• Temperature to fall up to 40°C
(approx. 2 hours).

15. 1. Should not be overloaded.
2. Arranged in a manner which allows free circulation
of air.
3. Material to be sterilized should be perfectly dry.
4. Test tubes, flasks etc. should be fitted with cotton
plugs.
5. Rubber materials and inflammable materials should
not be kept inside.
Precautions:

16. • Easy to install and
has relatively low
operating costs.
• It penetrates
materials.
• It is nontoxic.
• noncorrosive for
metal and sharp
• Less reliable than
autoclaving.
• Large temp
difference may arise
within device.
• Sharp instruments
get dulled.
• Many materials do
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17. STERILIZATION CONTROL
A)Biological controls: Spores of Bacillus atrophaeus or
Clostridium tetani on paper strips inoculated in
thioglycolate or cooked meat medium & incubated for 3
to 5 days at 37°C.
B) Chemical control: Browne’s tube No. 3 which shows a
green color after sterilization at 160°C for 60 minutes
(color changes from red to green).
C) Physical control: Thermocouples and temperature chart
recorder used.

18. 2. MOIST HEAT
Temp below 100˚C Temp at 100˚C Temp above 100˚C
• Pasteurization
• Inspissation
• Water bath
• Boiling
• Koch & Arnold
steamer/
Tyndallization
• Autoclave
MOA
• Denaturation of proteins
• Coagulation of proteins

19. a. Pasteurization
• Used in dairy industry for milk.
• All non sporing pathogens like
Mycobacteria, Brucella & Salmonella are
destroyed.
• Coxiella burentii causative agent of Q
fever survives holder method but killed by
flash method
METHOD TEMPERATURE TIME COOLING
Holder method
(LTLT)
63⁰C 30 mins 4˚C
Flash process (HTST) 72⁰C 15-20
sec
13˚C

20. • Process of heating an article on 3
successive days at 80-85⁰C for 30 minutes
by a special instrument called Inspissator.
• Egg based media – e.g. Lowenstein-Jenson
medium and Dorset’s egg medium.
• Serum based media- e.g. Loeffler’s serum
slope.
b. Inspissation

21. • It is used to incubate samples in water.
• Disinfection of serum, body fluids and
vaccine.
• Bacterial vaccine are disinfected at 60⁰C
for 1 hour.
• Serum or heat labile body fluids can be
disinfected at 56 ⁰C for 1 hour.
c. Water bath

22. • 100˚C for 10-30 min kills all vegetative
bacteria but not the spores.
• Addition of 2% sodium bicarbonate may
promote sterilization.
• Uses - For the disinfection of medical and
surgical equipment – when sterility is not
essential in emergency or under field
conditions.
d. Boiling

23. • An exposure of steam100˚C for 20 min
on three consecutive days is known as
Tyndallization.
• Koch or Arnold steam sterilizer.
• Similar to that of inspissation.
Uses
• Used for sterilization of gelation, egg,
serum or sugar containing media.
e. Tyndallization

24. • Principle – water boils when its vapor
pressure equals that of surrounding
atmosphere.
• When pressure inside a closed vessel
increases, the temperature at which
boils also increases.
• Steam condenses to water & gives up its
latent heat to the cooler surface it comes
in contact with.
AUTOCLAVE

26. USES:
 Sterilizing culture media containing water, aqueous solutions,
rubber materials, dressing materials, linen, gloves, instruments &
pharmaceutical products.
PRESSURE (Psi) Temperature (˚C) Time (mins)
15 121 15
20 126 10
30 134 5

27. • Nontoxic to patient, staff,
environment
• Cycle easy to control and
monitor
• Rapidly microbicidal
• Penetrates medical
packing, device lumens
• Deleterious for heat-
sensitive instruments
• Microsurgical
instruments damaged by
repeated exposure
• May leave instruments
wet, causing them to rust
• Potential for burns
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28. STERILISATION
CONTROLS
 Thermocouples
 Bacterial spores-
Bacillus
stearothermophilus
 Browne’s tube
 Autoclave tapes

29.  132 -134˚C at 28-30 lb/inch for 4 min
 Air is mechanically removed from the
chamber and load through a series of
vacuum and pressure pulses.
 This allows the steam to penetrate
porous areas of the load that couldn’t
otherwise be reached with simple
gravity displacement.
RECENT DEVELOPMENT:
PREVACUUM
AUTOCLAVE

30. FILTRATION
• Removal of microbes by passage of a liquid
or gas through a screen like material with
small pores.
• Heat sensitive materials.
• Separation of bacteriophages & bacterial
toxins from bacteria.
• Isolating viruses or organisms which are
scanty in fluids.

31. Types of Filters
Earthenware filters
(Candle filters)
Asbestos disc
(Seitz) filters
• Used widely for
purification of
water for industrial
and drinking
purposes
• Made up of a disc
of asbestos
(magnesium
trisilicate)
Membrane filters
 Water purification &
analysis
 Sterilization & sterility
testing
 Bacterial count of water

32. AIR FILTERS
 Large volumes of air may be rapidly freed from
infection by passage through high efficiency
particulate air (HEPA) filters.
 Remove fine particles of size 0.3mcm or larger.
 Widely used in laminar air
flow (LAF)
cabinets/chambers.

33. Standard Filtration Rate of MPPS
ISO / European
(ISO29463 / EN 1822) ≥ 99.95%
US Standard (MIL-STD-
282) ≥ 99.97%

34. RADIATION
rapid mass
sterilization of
syringes and
catheters
bacteriological
laboratory,
inoculation
hoods, laminar
flow and
sterilization of
disposable
items such as
plastic
syringes,
swabs, culture
plates,
cannulas,
catheters

35. NON IONISIZING RADIATION:
Types Produced by Mode of action Uses Comments
Infrared rays:
Sterilization of
Low energy items such as syringes Use of UV light
(Infrared and UV lamps DNA damage and catheters. is limited by
ultraviolet rays) Ultraviolet rays: penetration and
Used for disinfecting hazardous.
operation threatres and
laboratories.
IONISIZING RADIATION:
High energy
For the sterilization of They are
ionizing type antibiotics, hormones expensive.
(Gamma rays
and Cobalt-60 DNA damage and other prepacked They are very
X-rays and based disposable items such effective due to
cosmic rays) instruments as catheters, gloves, high penetrative
syringes, infusion sets, power.
oils, animal feeds, etc.

37. IDEAL REQUIREMENTS
 Broad & wide spectrum of activity
 Be active in the presence of organic
matter
 Be effective in acidic or alkaline media
 Have high speedy action
 High penetration power
 Be stable
 Be compatible with other agents
 Not corrode metals
 Not interfere with healing
 Not cause local irritation or sensitivity

38. Protein
coagulatio
n
Disruption of
cell
membrane
Removal of free
sulfhydryl groups
Substrate
competition

39. SPAULDING CLASSIFICATION

41. ALCOHOLS
 Used as skin antiseptics and act by
denaturing bacterial proteins.
 ETHYL (70%) & ISOPROPYL alcohol
(70%)- widely used
 ISOPROPYL ALCOHOL - better fat
solvent
more bactericidal
action
less volatile
 Methyl alcohol is effective against fungal
spores but weak bactericidal action.
DISADVANTAGES
• Relatively inefficient
in the presence of
blood and saliva.
• Causes corrosion of
metals.

42. ALDEHYDES
Two aldehydes are currently of considerable
importance:
Formaldehyde
 Both in liquid and gaseous
states.
 Aqueous solution is a
bactericide, tuberculocide,
fungicide, virucide and
sporicide.
 A 10% aqueous solution is
routinely used.
 However, penetration
is poor and the
surface must be
exposed to the gas
for upto 12hrs. for
effective sterilization.
GASEOU
S STATE

43. USES:
• Anatomic specimen
preservation
• 0% Formalin + 0.5% sodium
tetraborate – sterilised clean
instruments.
• Fumigating sick rooms,
laboratories.
• Disinfecting - clothing, bedding,
furniture (under controlled
• Formaldehyde leaves a
residue and instruments
must be rinsed before
use.
• Can cause
hypersensitivity
reactions

44. Glutaraldehyde
 High-level disinfectant and chemical
sterilant.
 It is used as 2% buffered solution.
 Destroys vegetative cells in 10 – 30 mins &
spores in 10hrs.
 It is recommended for use on surgical
instruments where residual blood may be
present.
 Kills by denaturation of proteins.

45. ADVANTAGES:
1.High biocidal activity.
2.Tuberculocidal,
sporicidal.
3.Used only as
immersion
sterilization/
disinfectant.
4.Active in organic
debris.
5.Doesn’t degrade
rubber & plastic
items.
Use:
• corrugated rubber
anesthetic tubes
and face masks.
• plastic
endotracheal
tubes.
• Dental
instruments.
• Optic fiber
endoscopes.
DISADVANTAGES:
1. It can damage
many metal items
e.g. nickel coated
impression trays &
carbon steel burs
2. It can cause
hypersensitivity on
repeated exposure
3. Should be
thoroughly rinsed
in sterile water
prior to use.

46. HALOGENS
 Iodine in aqueous and alcoholic solution
used as skin disinfectant.
 Active bactericidal agent and moderate
action on spores.
 Tincture of iodine: 2% iodine solution in
50% ethyl alcohol and sterile water.
 Used as an antiseptic for application
over skin and mucous membrane.
IODINE
DISADVANTAGES:
• Allergy
• Irritation
• Stains skin &
clothes.

47. POVIDONE-IODINE
 Maintains extended duration of effect, while
reducing risk of irritation and cytotoxicity.
 Oral care applications is the 1% PVP-I gargle,
which contains 0.1% available iodine.
 Given to patients with chronic periodontitis,
irrigating extraction sockets

48. CHLORINE:
 In the form of:
Bleaching powder
Sodium hypochlorite
Chloramine
 Disinfection is due to release of free chlorine with
protein to kill micro-organism.
 Used in water supplies, swimming pools, food and
dairy industries.
 CLOROX & PUREX (5.25%) - household sanitation.
 Sodium hypochlorite – root canal irrigant (2.5%) &
Dakin’s sol (0.5%)

49. ADVANTAGES:
• It causes tissue
dissolution
• It has antibacterial
and bleaching
action
• It causes
lubrication of
canals
• Economical
• Easily available
DISADVANTAGES:
• Irritant to tissue
periapically
• If comes in contact, it
may cause gingival
inflammation
• Bleach the clothes
• Bad odour and taste
• Vapours can irritate
the eyes

50. PHENOLS
ACTION:
penetrating and disrupting cell wall
denaturation of intracellular proteins.
 Phenol is active against gram positive
bacteria.
 Bactericidal at 1% and fungicidal at 1.3%.
 Commonly used Phenol derivatives are:
1. Cresol
2. Chlorhexidine
3. Chloroxylenol
4. Hexachlorophenes

51.  Lysol is a solution of 50% cresols in soap.
 Active against a wide range of
organisms.
 uses:
1. Infected glass ware.
2. Cleaning of floors.
3. Disinfection of excreta.
 Formacresols used as R.C medicament.
Cresols :

52. • Non-corrosive and non-irritating to
intact skin.
• It is used as antiseptic and for
disinfecting surgical instruments.
• Skin cream and soap: 0.8%,
Commercial 4.8 % solution used for
surgical antisepsis.
Chloroxylenol (Dettol)

53. Chlorhexidine
 Active against gram positive and gram
negative bacteria.
 At low conc. - bacteriostatic effect.
 At high conc. - bactericidal effect.
 USES: RC irrigant
post dental surgeries
denture stomatitis
adjunct to oral hygiene
 Mouth rinses - 0.2% & 0.12%
 Sprays – 0.1% & 0.2%

54. GASES
a. Formaldehyde gas
 150 grams of KMnO4 + 280 ml Formalin
 Employed for fumigation of operation theatres,
wards and laboratories etc.
 Sterilization is achieved by condensation of
gas on exposed surfaces.
 The doors should be sealed and left
unopened for 48hours.

55. b. Ethylene oxide (ETO)
 Highly lethal to all kinds of microbes including
spores.
 Its alkylates the amino, carboxyl, hydroxyl and
sulfhydryl groups in protein molecules.
Uses:
• Sterilizing plastic and rubber articles,
respirators, heart-lung machines, sutures,
dental equipment and clothings.
• Disposable plastic syringes, Petri dishes.
• Its high penetrating power can sterilize
prepackaged materials.

56. c. Betapropiolactone (BPL)
• Condensation product of ketone and formaldehyde
• Has low penetrating power but has a rapid action
• For sterilization of biological products 0.2% BPL is used
• Capable of killing all microorganisms and is very active against
viruses
Uses:
In the liquid form it has been used to sterilize vaccines, tissue grafts,
surgical instruments and sera as a vapor-phase disinfectant in
enclosed spaces.

57. Recent vapour phase disinfectants
Hydrogen peroxide
• Used to decontaminate biological safety
cabinets
Peracetic acid
• It is an oxidising agent
• One of the high level disinfectants
• Used in plasma sterilizers

58. CONCLUSION
 “PREVENTION IS BETTER THAN CURE”- a proverb well suited
to sterilization.
Aim of sterilization is to break the chain of cross infection
between patients by killing micro-organisms including spores.
Effective instrument cleaning prior to sterilization should be
carried out – reduces the bacterial load and renders sterilization
very effective.
Sterilization by steam is considered as the most efficient, cost
effective and safe method of sterilizing dental instruments.
The sterilization process performed should not damage the
instruments and should be repeatable, standardizable, verifiable
and a documented method.

59. REFERENCES
 Textbook of microbiology 5th edition R.Ananthanarayan and
C.K.Jayaram Paniker.
 Microbiology for dental students T.H.MELVILLE and C RUSSELL.
 Textbook of microbiology for dental students 3rd edition. Prof. C P
Baveja.
 Linden, K. and Murphy J.R. 2017. Physical Agents. In: J.B. Rose and B.
Jiménez-Cisneros, (eds) Global Water Pathogen Project, Part 4
Management of Risk from Excreta and Waste.