Antimicrobial AgentsMohammad Reza Fazeli, PharmD, PhD

Professor of Microbiology

Department of Drug and Food Control

Faculty of Pharmacy

Tehran University of Medical Sciences

Overview of Antibiotics as Therapeutic Agents Selective Inhibition/Toxicity

• Due to the differences in structure and metabolic pathways

• Harm microorganisms, not the host Four major sites:

• Cell wall• Ribosomes• DNA• Cell membrane

Spectrum of Activity

Broad Spectrum Antibiotics:• Effective against many types

• Example: Tetracycline

Narrow Spectrum Antibiotics:• Effective against very few types

• Example: Penicillin

Bactericidal v. Bacteriostatic

Bactericidal:

• Kill bacteria

• Used when the host defense mechanisms are impaired

Bacteriostatic:

• Inhibit bacteria

• Used when the host defense mechanisms are intact

MECHANISMS OF ACTION OF MAJOR GROUPS OF ANTIBIOTICS

STRUCTURE OF -LACTAM ANTIBIOTICS

PENICILLIN HOME

“Penicillin Home”

Looks like a house with a new room added to the side

Think of the R-group as of an antenna Changing “antennae” and or finishing the

“basement” will create better “homes” (penicillins)

[Penicillin] Home Improvement Project Adding a new antenna creates broad

spectrum penicillins • Example: Ampicillin

Adding additional antennae and finishing the basement creates cephalosporins• Example: 1st, 2nd, 3rd, & 4th generation

cephalosporins

SPECTRUM OF ACTIVITY OF CEPHALOSPORINS

MECHANISM OF ACTION OF -LACTAM ANTIBIOTICS

β-Lactam antibiotics inhibit the formation of peptidoglycan cross-links in the bacterial cell wall, but have no direct effect on cell wall degradation

The β-lactam moiety of penicillin binds to the enzyme (transpeptidase) that links the peptidoglycan molecules in bacteria. The enzymes that hydrolyze the peptidoglycan cross-links continue to function, which weakens the cell wall of the bacterium

Bacteria constantly remodel their peptidoglycan cell walls, simultaneously building and breaking down portions of the cell wall as they grow and divide

Gram-positive bacteria are called protoplasts when they lose their cell walls. Gram-negative bacteria do not lose their cell walls completely and are called spheroplasts after treatment with penicillin

-lactam antibiotics are ineffective against protoplasts and spheroplasts:

MECHANISMS OF ACTION OF ANTIRIBOSOMAL ANTIBIOTICS

Inhibition of Protein Synthesis

Anti-ribosomal antibiotics impair ribosomes by binding to either 50S or 30S ribosomal subunits

Ribosomes are essential for translation of mRNA into proteins

No translation No protein synthesis No protein synthesis No growth

MECHANISM OF ACTION OF SULFONAMIDES AND TRIMETHOPRIM

Folic acid (also known as folate , vitamin M, vitamin B9, vitamin Bc or folacin are forms of the water-soluble vitamin B9.

Folic acid is itself not biologically active, but its biological importance is due to tetrahydrofolate and other derivatives after its conversion to dihydrofolic acid in the liver.

Vitamin B9 (folic acid and folate) is essential for numerous body functions. The human body needs folate to synthesize DNA, repair DNA, and methylate DNA as well as to act as a cofactor in certain biological reactions. It is especially important in aiding rapid cell division and growth, such as in infancy and pregnancy. Children and adults both require folic acid to produce healthy red blood cells and prevent anemia.

Mechanisms of Resistance

Genetic Mechanisms

Non-Genetic Mechanisms

Genetic Mechanisms Chromosome-mediated

• Due to spontaneous mutation: • in the target molecule • in the drug uptake system

Plasmid-mediated• Common in Gram-negative rods• Transferred via conjugation• Multidrug resistance

Transposon-mediated

Non-Genetic Mechanism

Inaccessibility to drugs (e.g., abscess, TB lesion)

Stationary phase (insusceptible to inhibitors of cell wall synthesis)

Protoplasts and spheroplasts (insusceptible to inhibitors of cell wall synthesis)

The End Result of Genetically Conferred Resistance

Production of drug-inactivating enzymes

Modification of target structures

Alteration of membrane permeability

Resistance to Beta-Lactams

Gram +

-lactamase (Penicillinase)

• Alteration of the transpeptidase enzyme

Gram -

-Lactamase (Penicillinase)

• Alteration of porins

How can we test for susceptibility/resistance?

Antibiotic Susceptibility Testing

Dilution Method

Disc Diffusion Method

E-test

High-Tech Methods

Dilution Method

Prepare two fold [antibiotic] dilutions Add 1/2 a million bacterial cells per tube Incubate overnight Check for turbidity Establish the MIC:

• The lowest concentration of the drug that prevented the bacterial growth (no turbidity)

Disc Diffusion Method

Seed agar plates with bacteria in question Place antibiotic-discs over the seeded plate Incubate overnight Measure the inhibition zones Relate the results to the zones given in the

interpretive chart There is an inverse relationship between the

MICs and zone diameters

Therapeutic Index =

Max. Safely Achievable Level MIC