Transcript Lecture 11_PPT - DrMinkovskyScienceWiki

Physical and
Chemical
Control of
Microbes
Chapter 11
Copyright © The McGraw-Hill Companies, Inc) Permission required for reproduction or display.
Learning Objectives
• Distinguish between sterilization, disinfection, antisepsis and
decontamination
• Define “-static” and “-cidal”
• Identify factors affecting microbial death rate
• Name four categories of cellular targets for physical and
chemical agents
• Name and describe six methods of physical control of
microorganisms
• Give examples, describe modes of action and application,
and discuss advantages and disadvantages of the following
groups of chemical agents: halogens, phenolics, alcohols,
hydrogen peroxide, detergents, heavy metals, and aldehydes
Some Important Terms
•
Sterilization - kill all vegetative cells and
spores
•
Disinfection - reduces number of pathogens
on an inanimate surface (sanitation)
•
Decontamination - makes contaminated
surfaces safe to handle by reducing the
number of microbes present
•
Antisepsis - killing microbes on living tissue
Antimicrobial Effects
• Bacteriostatic
Inhibits growth
• Bactericidal
Kills cells
• Bateriolytic
Kills and lyses cells
Microbial Death
• Begins when a certain
threshold of
concentration and time
is reached
• Proceeds in a
logarithmic manner
• Young cells die faster
than old cells
Microbial Death Rate
• Affected by many
factors:
• Number of
microorganisms
• Type of microorganism
• Temperature and pH
• Concentration of the
agent
• physiological state
• other substances
(solvents, organic
matter, inhibitors)
Cellular Targets
• The cell wall
• The cell
membrane
• Cellular
synthetic
processes
(DNA, RNA)
• Proteins
Physical (Dry Heat)
• Incineration - flaming your
loops
• Baking - requires long periods
• 150° - 180C for 2-4 hours
• Advantages: cheap and easy
• Disadvantages: materials must
withstand high temperatures
and be dry (not aqueous)
Physical (Moist Heat)
• Boiling - will not kill endospores, used for
disinfecting drinking water, food, to sanitize
materials for babies
• Tyndallization – non-pressurized steam,
intermittent sterilization, used for heat-sensitive
materials.
• Pasteurization - High heat, short time
• Autoclaving – Steam under pressure
Pasteurization
• Commonly used with juice, beer, milk, and other
dairy products to prevent spoilage
Batch - 63°C for 30 min
High Temperature Short Time - 72°C for 15-20 sec
Ultra-High Temperature - 134°C for 1-2 sec
• Does not kill spores and thermodurant bacteria
• Target: Salmonella, Brucella, Campylobacter
jejuni, Listeria monocytogenes, Coxiella burnetii,
Mycobacterium bovis.
Autoclaving
• Commonly use in the
laboratory
Recorder
• Temperatures higher
than boiling
Control
handle
• Use steam pressure 15 p.s.i. above normal
(2 atm)
Door
gasket
Discharge
• 121°C for 20 min
Steam
supply
valve
• Kills all endospores
• Home pressure
cookers do the same
thing
Pressure regulator
Safety valve
Exhaust to atmosphere
Steam from
jacket to chambe
or exhaust from
chamber
(a)
Temperaturesensing
bulb
(b)
Steam from
Steam
to jacket Jacket to chamber
Strainer
Steam
jacket
Steam
trap
Condensate
to waste
Thermal Death Measurements
Both time and temperature have to be considered
• Thermal death point the lowest possible temperature that will achieve
complete killing within ten minutes
• Thermal death time the minimum time to achieve complete killing in a
liquid solution at a given temperature
Physical (Cold)
• Freezing - kills some cells due to ice crystal
formation
• Refrigeration – does not kill bacteria, only slows
down growth. Ignorance of this fact results in
food poisoning
• Lyophilization preservation of microbes by
freezing and drying.
• Advantages: many products tolerate cold better
• Disadvantages: very little killing and is expensive
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Physical (Filtration)
Liquid
• Pass liquid or gas
through a filter with
sufficiently small pore
size
Filter Pore
• HEPA - filter out > 0.3
µm particles
• Advantages: No thermal
damage
Filter
Sterilized
fluid
(a)
Vacuum
Pump suction
• Disadvantages: viruses
not eliminated and must
be either liquid or gas
(b)
b: © Fred Hossler/Visuals Unlimited
Ionizing vs Non-ionizing radiation)
• Ionizing Radiation:
ejects electrons from an
atom, causes ions to form
• Gamma ray - very good
penetration
• X-ray - less penetration
• Non-ionizing radiation
raises electrons in an
atom to a higher energy
state
• Ultraviolet - damages DNA,
with poor penetration
Ultraviolet Damage
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Normal
segment of
DNA
• Wavelenth: 200-300 nm
A
C
A
A
C
T
G
T
T
G
• Poor penetrating power
• Pyrimidine dimers
Thymine
dimer
• Mutations induced
UV
A
C
A
A
C
T
G
T
T
G
O
Details
of bonding
O
CH3
T
O
CH3
T
O
Radiation
• Advantages: very
effective
with little product
damage (“cold”
sterilization)
• Disadvantages:
dangerous
materials need
shielding,
and lack of public
trust.
Chemical Treatments
• Chemotherapeutics - disease treatment
• Disinfectants - cleaning surfaces
• Many are available today (hundreds)
• Choice is based upon:
 Nature of the object
 Kinds of microbes targeted
 Desired effect
Chemical (Oxidizers)
• Damage proteins and membranes
• Halogens
• Chlorine - disinfectant (added to water)
• Iodine - antiseptic (tincture of iodine, betadine)
• Hydrogen peroxide (H2O2)
• 3% is a weak antiseptic
• Your body and many bacteria can break this
down enzymatically
Chemical (Phenolics)
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
• Denature proteins
OH
OH
OH
CH3
• Disrupt membranes
• Joseph Lister
Examples: phenol,
lysol, chlorhexidine
• Effective on surfaces
• Many of these are
too toxic to apply to
tissue
o-cresol
Phenol
(basic aromatic
ring structure)
OH
CH3
p-cresol
OH
OH
Cl
Cl
Cl
Chlorophene
(a chlorinated phenol)
Cl
CH2
Cl
Cl
Cl
Hexachlorophene
(a bisphenol)
Chemical (Alcohols)
• Denature proteins
• Disrupt membranes
• Examples: ethanol, and
isopropanol
• Most effective at 50-70%
• Increased plasmolysis after damage
• These are commonly used for antisepsis
Chemical (Surfactants)
• Amphiphilic compounds
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
R1
• Disrupt membranes
• Quaternary ammonium
compounds (quads)
N
R2
• Example - cepacol,
and roccal
Hydro carbon chain (C number from 8
to18)
R4
R3
(a)
+
• Charged nitrogen
• Four hydrophobic
groups
+
CH3
CNH2N+
N+
CH2
CH2
(b) Benzalkonium chloride
Cl–
Oligodynamic Effect
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
• Inhibition by
heavy metals
• Silver
• Copper
• Mercury
• Gold
Silver amalgam
Gold foil
© Kathy Park Talaro/Visuals Unlimited
Chemical (Alkylators)
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
• Damage proteins or
DNA by adding
Glutaraldehyde
carbon adducts
O
O
• Examples: formalin,
glutaraldehyde, and
ethylene oxide Polyglutaraldehyde
Polymerization
O
O
O
• Highly noxious
• Ethylene oxide is
used to sterilize
products via gas
O
Cross-linking with
microbial protein
N
N
N
N
G–
G+
Amino groups in
peptidoglycan