Chemical Methods for Controlling Microbial Growth

Controlling Microbial Growth
• Bactericidal: Bacteria-killing
• Bacteriostasis: Bacterial growth inhibiting
• Disinfection: Destruction of vegetative pathogens
• Antisepsis: Destruction of vegetative pathogens
on living tissue
• Aseptic: Absence of a pathogen from an object or
an area
Vocabulary Cont.
• Sanitation: Reduction of pathogens on eating
utensils, etc. to public health standards
• Antiseptic: chemical used for disinfection of
skin or mucous membranes
• Disinfectant: Chemical used on inanimate
objects to kill or inhibit bacterial growth
• Sterile: Without life, free of microorganisms
Measuring the effectiveness of
bacterial control methods
• Not all genera of bacteria are equally
susceptible to a given method of control.
– Example:
• E. coli bacteria are highly susceptible to heat and die
quickly when they reach boiling temperatures.
• Pathogens that create endospores on the other hand,
can survive long periods of boiling.
• How do we determine how hot and how long
we need to keep a microbe at to kill it?
Three acceptable ways to measure
microbial tolerance to heat
1. Decimal reduction time (DRT): time in which
90% of the population of bacteria at a given
temperature will die.
2. Thermal death time (TDT): Time in which all
cells in a suspension are killed at a given
3. Thermal death point (TDP): The lowest
temperature in which all microorganisms will
be killed in 10 minutes.
Death Curve
Microbes die at a constant, logarithmic rate when treated.
Death Curve
What is the DRT for the above graph?
What is the TDT?
Physical Methods of Controlling
Microbial Growth
Moist Heat Methods
• More effective than dry heat
• Kills by coagulation of proteins
• Three methods: boiling, autoclaving,
• 100 C for 10 min: kills vegetative bacteria,
most viruses, and fungal spores.
– Endospores and some viruses (hepatitis) can
survive for long boiling periods.
• Good for making drinking water and food safe
for consumption.
– Unreliable for sterilization of surgical equipment
(endospores survive without stomach acid to kill
them). Free flowing unpressurized steam is
equivalent to boiling.
• Pressurizing of steam to make it hotter. 1 atm
temp = 121 C kills all organisms and their
endospores. Most effective and most
preferred method of sterilization. Steam must
contact all solid surfaces since solids don’t
have convection currents like liquids do.
• Mildly heating food products to kill particular
spoilage microorganisms or pathogens. 72 C
for 15 seconds with refrigeration needed
afterwards. Many heat resistant bacteria
survive pasteurization but these are unlikely
to cause disease.
• Ultra high temperature pasteurization 140 C
for 3 seconds sterilizes milk.
Dry Heat
• Kills and sterilizes by oxidation effect.
• Direct flaming or incineration
• Hot air sterilization (170 C for 2 hours)
• Filters are made of cellulose or plastic
polymers with pores of about .2 micrometers.
Viruses can still get through.
• There are filters with pore size as small as .01
micrometers for viruses but filtration is very
• Filters are important when you can’t heat
something up because you’ll destroy what
you’re trying to keep.
• Cold is bacteriostatic
• Slow freezing is more harmful as crystals can
set up better.
• Deep freezing is quick freezing at temp
between -50 and -95 degrees. This is usually
done to preserve the specimen.
• Desiccation is the absence of water. Microbes
need water in order to grow and reproduce.
However, microbes in a desiccated state can
be viable for years. When water is added, they
can resume growth.
• Desiccation is bacteriostatic, though the ability
to survive varies by species.
Lyophilization (freeze drying)
• A suspension of microbes is quickly frozen at
temp between -54 and -72 C and water is
removed by high vacuum. Microbes can be
stored for years in this state.
Chemical Methods for
Controlling Microbial Growth
Test Guide to Chemotherapy
• Different strains of microbes have different
levels of susceptibility to different
chemotherapeutic agents.
• Susceptibility of a microorganism can change
over the course of a treatment.
• Physicians need to know the sensitivities of
the pathogen before treatment can be
started. Various tests are employed to give
Physicians the knowledge they need.
Methods for Testing Disinfectants
• The American Official Analytical Chemist’s
Use-dilution method (Use-Dilution Method)
• The Disk-diffusion Method
• Three bacteria, Salmonella, Staphylococcus,
and Pseudomonas, are used as standards to
examine the effectiveness of a given
• Small stainless steel cylinders are dipped and
coated with these bacteria and than allowed to
• The cylinder is than dipped into one of several
dilutions of the chemical agent to be tested for 10
minutes, removed, rinsed with water, and placed
into a tube of nutrient broth.
• Chemicals that prevent growth at the greatest
dilutions are considered to be the most effective.
Example Results
Disinfecting Agents
Dilution Ratio Agent 1
Agent 2
+ indicates growth
- indicates no growth
Agent 3
Disk-diffusion (Filter Paper) Method
• Most common testing method.
• Small disc of filter paper is soaked with
chemical and placed on surface of agar plate
that has been previously inoculated with the
test organism.
Disk-diffusion Cont.
• The chemical is concentrated in one area, and
as it diffuses out, it is less concentrated. The
effectiveness is based on how big of a circle
“kill-zone” or zone of inhibition is around
where the chemical is introduced.
• Multiple chemicals can be tested against one
bacteria at a time with this method.
Example Results
Chemical Disinfectants
• Can kill in three ways:
– Damage of Plasma Membrane
– Inactivate Enzymes
– Denature Proteins
Phenol and Phenolic
• Phenol: First used by Joseph Lister in an
operating room. It proved too toxic for skin
and had a terrible odor.
• Phenolic: Adapted version of phenol that is
less potent and more useful.
• Kills in all three ways.
Iodine (a halogen)
• Kills by inactivating enzymes by attaching to the
amino acid tyrosine.
• Effective against bacteria, endospore, fungi, and
some viruses.
• Iodine is available as tincture solution (in solution
with alcohol). Tincture solutions often stain and
patients may develop hypersensitivity to it.
• Iodine can come in Iodophor (combined with an
organic molecule for slower releasing).
Chlorine (a halogen)
• Chlorine is mostly just a disinfectant. Chlorine
turns to hyochlorous acid, its active form,
when it’s added to water.
• Kills by deactivating enzymes. The acid
oxidizes cellular molecules into inactive forms.
Alcohol (ethanol and isopropyl)
• Alcohols work in two ways: by denaturing
proteins and dissolving lipids (plasma
• Effective against bacteria and fungi but not
endospores and non-enveloped viruses.
Alcohol is most effective when it is diluted by
water. (ethanol 70%, isopropyl 90%). Nonwatered alcohol doesn’t carry into cells very
Don’t drink isopropyl alcohol.
That is all.
Heavy Metals
• Gold, silver, copper and zinc are germicidal.
Silver nitrate is used in infants’ eyes.
• Selenium and zinc in shampoos control fungi
that cause dandruff.
• Heavy metals combing with –SH groups to
denature proteins.

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