Chapter 5
Microbial growth control
Physical methods
Chemical agents
Food Preservation
Control of microorganisms by
physical and chemical agents
frequently used terms
• Sterilization: the process by which all living cells, viable
spores, virus, and viroids are either destroyed or removed
from an object or habitat.
• Disinfection: the killing, inhibition, or removal of
microorganisms that may cause disease.
• Sanitization: the microbial population is reduced to levels
that are considered safe by public health standards.
• Antisepsis: the prevention of infection or sepsis
• -cide: kill; -static: stop
germicide: kill pathogens but not necessarily endospores.
Bactericide; fungicide; algicide; viricide; bacteriostatic;
fungistatic
Conditions influencing the effectiveness of
antimicrobial agent activity
• 1. Population size.
• 2. Population composition. Spore, young/mature
• 3. Concentration or intensity of an antimicrobial agent.
• 4. Duration of exposure.
• 5. Temperature. higher
• 6. local environment.
eg. 1. Heat kill more readily at an acid pH.
2. Organic matter protect microorganisms.
3. Biofilm protection.
Microbial Growth Control
• Physical ways
– Heat sterilization (including autoclave
and pasteurization)
– Radiation (microwaves, UV, X-rays,
g-rays and electrons)
– Filtration (depth filter, membrane
filter and nucleation track
(nucleopore) filter)
Heat(terms)
• TDP: the lowest temperature at which a microbial
suspension is killed in 10 minutes.
• TDT: the shortest time needed to kill all organisms
in a microbial suspension at a specific temperature
and under defined conditions.
• Decimal reduction time (D) or D value: the time
required to kill 90% of the microorganisms or
spores in a sample at a specified temperature. D121
Moist heat sterilization
• Autoclave: a device somewhat like a fancy
pressure cooker.
• The air initially present is forced out the chamber
is filled with saturated stream
• 121oC or 15 pounds
15-30min
1. All air must be flushed out of the chamber, or it
will not reach 121oC even though it may reach
pressure of 15 pounds.
2. The chamber should not be packed too tightly.
Pasteurization
• Milk, beer, and many other beverages are treated
with controlled heating at temperatures well
below boiling. Not sterilize, just pasteurized.
• Past : 63oC, 30min
• HTST: 72oC, 15sec
• UHT: 140-150oC, 1-3sec
Dry heat sterilization
• 160-170oC
2-3h
• Suitable for glass petri dishes and pipettes
• Not suitable for heat-sensitive materials like
many plastic and rubber items.
Filtration
• Heat-sensitive solution
• Depth filters: diatomaceous; unglazed porcelain.
• Membrane filters: membranes with pores about
0.2m in diameter are used to remove most
vegetative cells, but not viruses.
• Air: 1. Surgical masks and cotton plugs on culture
vessels.
2. Biological safety cabinets.
Radiation
• 1. Ultraviolet radiation: 260nm is quite lethal but
does not penetrate glass, dirt films, water and
other substances.
uv lamps: the ceilings of rooms
biological safety cabinets
attention! uv lamp must be off
• 2 . Ionizing radiation: not always as effective
against viruses.
Co60: cold sterilization for antibiotics, hormones,
sutures.
Microbial Growth Control
• Chemical ways (germicides)
– Agents that kill organisms are often called cidal agents
(bactericidal, fungicidal and viricidal agents)
– Agents that do not kill but only inhibit growth are called
static agents (bacteriostatic, fungistatic, and viristatic
agents)
– Disinfectants are chemicals that kill microorganisms and
are used on inanimate objects
– Antiseptics are chemical agents that kill or inhibit growth
of microorganisms and that are sufficiently nontoxic to be
applied to living tissues.
Ideal disinfectant
• 1. Must be effective against a wide variety of
infectious agents at high dilutions and in the
presence of organic matter.
• 2. Toxic for infectious agents
no toxic to people or corrosive for common
materials.
• 3. Odorless or with a pleasant odor, soluble in
water and low surface tension.
• 4. Relatively inexpensive
Phenolics
• Denaturing proteins and disrupting cell
membranes. Phenol, orthocresol(邻甲酚)
• Advantages:
1. Tuberculocidal
2. Effective in the presence of organic material.
3. Remain active on surfaces long after application.
• Disadvantages:
1. Disagreeable odor
2. Cause skin irritation
alcohols
• Bactericidal and fungicidal, not sporicidal
• 70-80% ethanol and isopropanol
• Denature proteins and dissolve membrane
lipids.
• 10-15min soaking
Halogens
• Iodine and chlorine
• Iodine: oxidizing cell constituents
iodinating cell proteins.
2% or more iodine in a water-ethanol solution of
potassium iodide.
• Iodophor(碘载体): iodine is complexed with an
organic carries to form. water soluble, nonstaining,
release iodine slowly to minimize skin burns and
irritation.
• Chlorine: municipal water supplies
swimming pools. Not spores
Cl2, HClO, Ca(OCl)2,
oxidation
Heavy metals
• Mercury (Hg), silver (Ag), arsenic(As), zinc
(Zn), copper (Cu)
• 1% AgNO3: the eyes of infants to prevent
ophthalmic gonorrhea
• Silver sulfadizine is used on burns.
• CuSO4: algicide in lakes and swimming
pools.
Aldehydes
• Combine with nucleic acids and proteins
and inactivate them, probably by crosslinking and alkylating molecules.
• Sporicidal
• Formaldehyde; 2% glutaraldehyde(戊二醛)
Sterilizing gases
• Ethylene oxide gas
• 10-20% mixed with CO2 or
dichlorodifluoromethane(二氯二荧光甲烷)
• Relative humidity: 40-50%
38oC 5-8h or
54oC 3-4h
EtO con: 700mg/liter
• Toxic! Removed before use.
Evaluation of antimicrobial agent
effectiveness
• The phenol coefficient test:
• The test bacteria: Salmonella typhi
Staphylococcus aureus
a series of dilutions of phenol and the experimental
disinfectant.
bacteria + agent; 5min, 10min, 15min; subculture to
fresh medium, respectively; incubate 2-3 days.
The highest dilutions that kill the bacteria after a 10mins
exposure, but not after 5mins, are used to calculate the
phenol coefficient.
phenol coefficient =disinfect max dilution/phenol max
dilution
If >1: more effective than phenol
Microbial Growth Control:
Chemotherapeutic agents
• Chemotherapeutic agents: to be used
internally for control of infectious disease,
they must have selective toxicity, these
include:
– Growth factor analog:
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Sulfa drugs (磺胺药)
Amino acid analogs
Vitamin analogs
DNA base, RNA base analogs
Chemotherapeutic agents: Antibiotics
• Antibiotics are chemical substances produced by certain
microorganisms that inhibit and kill other microorganisms, they are
natural products rather than synthetic chemicals
– Gram-positive bacteria are usually more sensitive to Gramnegative bacteria
– In bacteria, the important targets of antibiotic action are the cell
wall, the cytoplasmic membrane and the biosynthetic processes
of protein and nucleic acid synthesis
 b-lactam (内酰胺) group, which includes the penicillins and
related compounds, has major clinical significance
Measuring antimicrobial activity
Minimum inhibitory concentration (MIC)
Agar diffusion method
Tube dilution technique
Range of actions of antibiotics and other agents
Mode of action of major antibiotics
Food Preservation
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Sterilization
Low temperature (best -20oC or 80oC)
pH or acidity
Low water availability
Canning
Chemical food preservation
Questions
• several concepts
• Conditions influencing the effectiveness of
antimicrobial agent activity?
• What are the physical- and chemical ways to
halt microbial growth?
• On what sites do antibiotics act on cells?
• How to preserve food?