1. Control of microbial growth: Chemical and Physical methods

2. Vocabulary
Sterile: devoid of life. Something is either sterile or not.
Disinfect: kill most microbes, especially harmful ones, but probably not necessarily spores which are resistant.
Disinfectant: chemical used on inanimate objects to kill microorganisms.
Antiseptic: used to disinfect living tissue; must be gentler.
Bacteriostatic: keeps bacteria from growing.
Bactericidal: kills them. Sometimes subtle differences between bacteriostatic and bactericidal.
Sanitize: no specific meaning; clean thoroughly so that harmful bacteria are probably dead.

3. Kinetics of bacterial death
Bacteria not only grow exponentially, but die that way too.
Factors that affect the rate of death include: temperature, pH, concentration of disinfectant, type of microbe, and presence of organic material.
The longer the treatment is applied, the more that are killed.
The more microbes there are, the longer it will take.

4. How do antimicrobial agents work?
Attack proteins
Oxidize, hydrolyze, or bind to proteins.
Change 3-D structure, usually irreversibly; ruin protein.
Dissolve membranes or damages cell walls
Leaky membranes means vitamins, metabolites, escape
Proton gradient across membrane gone, little ATP made.
Wall destroyed, loss of osmotic protection
Damage to nucleic acids
DNA denatured or broken, cell can’t replicate
RNA molecules in ribosomes affected

5. Microbes vary in susceptibility
Bacterial endospores: hardest of all to kill
Resist high heat, dessiccation, chemicals, radiation
Cysts of protozoa
Resting cells of eukaryotes, similar to endospores
Mycobacteria: waxy cell wall provides resistance
Gram negative bacteria: more resistant than G+
Viruses: some hard to kill, other easier
Some fungi hard to kill, others easier
Parallels levels of disinfectants:
High level, intermediate level, low level

6. The Ideal Disinfectant
Fast and effective, even in the presence of organic material (like blood, vomit, feces..)
Effective, but non-toxic to humans.
Penetrate materials without damaging them.
Easy to prepare and stable over time.
Inexpensive and easy to apply.
Not stink!

7. Disinfectants vary in effectiveness Microbes vary in susceptibility
Difficult to develop standard tests for disinfectant effectiveness suitable for all situations
Use-dilution test
Metal cylinders coated with microbes are placed in different dilutions of disinfectant
Number of surviving microbes determined.
In-Use test
Swab surface of object to be disinfected, disinfect, then swab again.
Determine number of bacteria before and after.

8. Disinfectants -1 Halogens: Cl, I, Br, Fl Phenols and phenolics
Chlorine: Cl2 gas, hypochlorites (bleach), and chloramines (NH2Cl)
I as tincture or as iodophors such as betadine.
Phenols and phenolics
O-phenyl phenol (Lysol), hexachlorophene, triclosan, chlorhexidine
Alcohols: isopropanol, ethanol; best at 70-95%
Good at removing lipids w/ attached bacteria from skin
Weakly attacks proteins, cell membranes
phenol

9. Disinfectants -2 Hydrogen peroxide and ozone
H2O2, stored at 30%, used at 3%
new plasma gas sterilizers; read article
Soaps and detergents: cationic (Quats), anionic
Soaps are alkaline salts of fatty acids, weak
Quaternary ammonium compounds weak but useful.
Mostly wash away microbes or damage membranes.
Heavy metals: Hg, Ag, Se, Cu
Used less; toxic and corrosive. Hg and Ag historically

10. Disinfectants -3 Alkylating agents
Formaldehyde, glutaraldehyde (effective at high pH)
Ethylene oxide: common, toxic, explosive
Acids and alkalis: often microbiostatic
Acetic and lactic acids in foods;
Salts of benzoic, sorbic, and propionic
Other stuff
Sulfites (control regrowth in wine), nitrites (botulism or cancer?), various dyes (selective growth media)

11. Chemical structures Na+ OCl- sodium hypochlorite Ethylene oxide
Soap Na salt of a fatty acid
Benzalkonium chloride (quat)

12. Physical methods Temperature
Cold: slows or prevents growth, may kill slowly
Ultra-cold used to preserve bacteria long term
Heat: denatures enzymes, kills cells
Moist heat
Traditional pasteurization does not sterilize;
Standard protocols, time and temperature e.g. 62.9°/ 30 min
flash pasteurization ° / 15”
Newer UHT sterilizes: ° in 5”

13. More on temperature Sterilizing with moist heat: boiling, autoclave
Boiling will not necessarily kill endospores
Autoclave is steam heat under pressure, so above boiling
121°C, psi. Very effective.
Compare moist heat and dry heat
Dry heat: 170 deg C, near 350 F, for 2 hours
Water conducts heat much more effectively, sterilizes at lower temperature for shorter time.
Dry heat not useful for liquids!
Incineration (e.g. flaming loop) has its place too.

14. Physical methods-2 Drying: cells need water. Remove it, and no growth.
Freeze drying: lyophilization; also used to preserve cultures for long term storage.
Osmotic pressure/high salt
Sucks water out of cytoplasm; salted meat, jellies, etc.
Radiation
UV used to sterilize air, surfaces in hospitals, etc.
Ionizing radiation: x-rays (electron beam) and gamma rays; important treatment of plastics, various foods.
Irradiation of meat important tool in food safety.
Microwaves only boil;

15. Physical methods-3 Filtration
Membrane filtration: thin plastic disks with holes of 0.22 or 0.45 micrometers, separate liquid from bacteria
Used to collect bacteria or sterilize liquids
Solutions of vitamins or proteins can be destroyed by heating
Air filtration: HEPA filters used in hospitals and also homes to help remove dust-borne bacteria, allergens.
High efficiency particulate air = HEPA