1 1 06/20/22 06/20/22 Antibiotics • Hugh B. Fackrell • Filename: antibiot.ppt
Dec 31, 2015
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Antibiotics
• Hugh B. Fackrell
• Filename: antibiot.ppt
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Outline• History
• Ideal properties
• Sources
• “Sulfas”– Antimetabolites– antibiotic synergism
• Major Groups of antibiotics
• Mechanisms of action
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History
• Salvarsan 606
• Prontosil
• Penicillin
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Salvarsan 606
• Paul Ehrlich– early 1900’s– syphilis– arsenic + organic compound
• Aniline dyes -
– wasn't able to find the "magic bullet”
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Prontosil
• 1930's, Gerhard Domagk– Prontosil
• 1935, Jacques and Therese Trefoncel– discovered that the active compound in
Prontosil was Sulfanilamide
• sulfanilamide “ Sulfas”
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Penicillin
• 1928, Alexander Fleming– antibacterial activity in Penicillium mold
(called it Penicillin)
• 1938, Howard Florey and Ernst Chain– developed Penicillin as an effective antibiotic
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Antimicrobial Therapy• Antimicrobics
– substances produced by microbes that inhibit other microbes
• Semi-synthetic antibiotics– naturally produced but altered
• Synthetic antibiotics:
– derived from chemicals
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Ideal Properties of an Antibiotic
• Low toxicity for patient– kills the invading microorganism without damaging the
host– no adverse side reactions– non allergenic
• High toxicity for microbe– bactericidal not bacteriostatic– broad spectrum
• Low risk of other infections
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More Characteristics
• drug can be administered orally or parenterally (by injection)– Soluble in tissue fluids– absorbed by and dissolved in tissues or body
fluids
• levels of active drug sustained long enough to kill the invading agent
• Long “Shelf” life
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Still More Characteristics• Low probability of resistance
• Microbial drug resistance develops slowly
• microbicidal rather than microbistatic
• Not inactivated by organic material
• Assists the host in eliminating the infecting microbe
• Not a powerful allergen
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Sources of AntibioticsMost spore-forming microorganisms
• Fungi
– Penicillium penicillin,
– Cephalosporium griseofulvin
• Bacteria
– Bacillus bacitracin, polymyxin, tyrothricin, colimycin, gramicidin
– Streptomycetes Aminoglycosides, nystatin, chloramphenicol, erythromycin, tetracylcine...
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Mechanisms of Drug Action
• inhibit cell wall synthesis
• inhibit nucleic acid synthesis
• inhibit protein synthesis
• interfere with cell membrane function
Sulfa Drugs
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Sulfa vs PABA
PABAPABA
NHNH22HOOCHOOC
SulfanilamidSulfanilamidee
NHNH22NHNH22SOSO22
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Structure of Sulfa Drugs
ProntosilProntosil
SulfisoxazoleSulfisoxazole
SulfanilamideSulfanilamide
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Folic Acid MetabolismPABA + pteridinePABA + pteridine
ThymidineThymidine
DNADNA
PurinesPurines
DNA, RNADNA, RNA
MethionineMethionine
tRNa, tRNa, ProteinsProteins
Pteridine Pteridine synthetasesynthetase
Dihydropteroic acidDihydropteroic acid
[GTP][GTP]
Dihydrofolic AcidDihydrofolic Acid
Dihydrofolate Dihydrofolate SynthetaseSynthetase
L- GlutamineL- Glutamine
Tetrahydrofolic AcidTetrahydrofolic Acid
DihydrofolatDihydrofolate e synthetasesynthetase
2 NADPH2 NADPH
2 2 NADP+NADP+
SulfonamidSulfonamidee
TrimethopriTrimethoprimm
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ThymidineThymidine
DNADNA
PurinesPurines
DNA, RNADNA, RNA
MethionineMethionine
tRNa, tRNa, ProteinsProteins
Folic Acid InhibitionPABA + pteridinePABA + pteridine
Dihydropteroic acidDihydropteroic acid
Dihydrofolic AcidDihydrofolic Acid
Tetrahydrofolic AcidTetrahydrofolic Acid
SulfonamidSulfonamidee
TrimethopriTrimethoprimm
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Antibiotic Synergism
SulfisoxazoleSulfisoxazole
TrimethopriTrimethoprimm
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Antibiotic Synergism
• Sulfonamide + trimethoprim
• Effective dosage 10% of two separately
• Broader spectrum of action
• Reduce emergence of resistant strains
Major Groups of Antibiotics
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Major Groups of Antibiotics
• Aminoglycosides– streptomycin, kanamycin, neomycin,
gentamicin, spectinomycin, tobramycin, amikacin
• Beta lactams– Penicillins, cephalosporins
• Lincomycins– lincomycin clindamycin
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Major Groups of Antibiotics• Macrolides
– erythromycin, carbomycin
• Polypeptides– polymyxin, colimycin, bacitracin, tyrothricin
• Polyenes– amphotericin B, nystatin
• Rifamycins– Rifampin
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Major Groups of Antibiotics• Synthetic
– pyridine• isoniazid, ethambutol
– sulfonamides• sulfanilamide, sulphisoxazole
– misc• nitrofurans, metronidazole, nalidixic acid
• Tetracyclines– oxytetracline, chlortetracycline
• Unclassified– Chloramphenicol, vancomycin
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PENEMS• Carbapenems• “Ideal” antibiotics
– non toxic– broad spectrum– good “Shelf” life– effective at very low conc
• Attach to Penicillin Binding Proteins– found in cell membrane– Gm+ve lysis through loss of cell wall integrity– Gm -ve filamentous bacteria loss of septum formation
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Adverse Effects of Antibiotics• Aminoglycosides
– Ototoxic- destroys cochlear hair cells
– renal toxic
• Chloramphenicol– depresses bone marrow
– aplastic anemia
– fatal “Grey baby” syndrome
• Penicillins– allergy anaphylaxis
• Vancomycin– thrombophlebitis
– ototoxic
– renal toxic
• Polymyxin, bacitracin colimycin– renal toxic
• Sulfas– skin allergy
– anemia
– renal toxic
– hepato toxic
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Adverse Effects of Antibiotics
• Broad spectrum– Super infections – Candida albicans– Clostridium difficle– Staphylococcus– Gram -ve
TetraclycineTetraclycine– Depress bone Depress bone
marrowmarrow– ““Yellow teeth”Yellow teeth”
Pregnant womenPregnant women children <7 yearschildren <7 years
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Mode of Action of Antibiotics
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Mode of Action of Antibiotics• Inhibit Synthesis of Cell Wall
• Damage Cell Membrane
• Inhibit Protein Synthesis
• Inhibit Nucleic acid Synthesis
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Bacterial Cell Wall
• Peptidoglycan– many layers in gram positives– thin in gram negative
• protects the cell against rupture from hypotonic environments
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Synthesis of peptidoglycan (1/4)
• Uridine diphosphate (UDP) derivatives of NAM and NAG are synthesized in the cytoplasm
• Amino acids are sequentially added to UDP-NAM to form the pentapeptide chain using ATP as an energy source. The two terminal D-alanines are added as a dipeptide (Cycloserine)
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Synthesis of peptidoglycan (2/4)
• The NAM- pentapeptide is transferred from UDP to a bactoprenol PO4 at the membrane surface. Bactoprenol is a 55-Carbon alcohol that attaches to NAM by a pyrophosphate group and moves peptidoglycan components through the hydrophobic membrane
• UDP-NAG adds NAG to the NAM-pentapeptide to form the peptidoglycan repeat unit
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Synthesis of peptidoglycan (3/4)
• The completed NAM-NAG peptidoglycan repeat unit is transported across the membrane to its outer surface by the bactoprenol pyrophosphate carrier
• The peptidoglycan unit is attached to the growing end.
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Synthesis of peptidoglycan (4/4)
• The bactoprenol carrier returns to the inside of the membrane to collect another NAM-pentapeptide. Bactoprenol pyrophosphate must give up phosphate to connect Bacitracin
• Finally, transpeptidization - interbridges formed
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Inhibit Synthesis of Cell Wall
• penicillin, bacitracin, vancomycin, cephalosporin, carbapenems
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Inhibition of Cell Wall Synthesis• Cycloserine - inhibits peptidoglycan sub-unit
formation
• Vancomycin - inhibits peptidoglycan elongation
• Beta-lactam antibiotics - Penicillins – lactam antibiotics block peptidases required to connect
inter bridges
• Cephalosporins bind to the peptidases that are essential to cross link the glycan molecules.
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Murray 2.4 & 5.4, p. 10
Inhibition of cell wall synthesis
• Cycloserine - inhibits the addition of the two terminal D-alanines
• Bacitracin - inhibits the transport of the subunits to their position in the cell wall
• Vancomycin - inhibits the elongation of the peptidoglycan to form connecting units
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Inhibition of Cell Wall Synthesis
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Natural Penicillins
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Semi Synthetic Penicillins
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Semi Sythetic Penicillins -2
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Structure of Penicillin
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Hydrolysis of Beta Lactam Ring
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Comparison of Structures
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Pen G in Blood
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Damage Cell Membrane
polymyxin, colimycin, nystatin, amphoteracin, tyrothricin
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Injury of Plasma Membrane
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Polymyxin
Membrane
Cytoplasm
Polymyxin action
• Polymyxin B binds to the cell membrane to disrupts its structural and permeability properties
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Inhibit Protein Synthesis• Binds to 50S ribosomal subunit
– prevents peptide chain elongation• clindamycin, chlorampenicol, erythromycin
– block rRNA(23S)• lincomycin, macrolides
• Binds to 30S ribosomal subunit– misreading of mRNA
• aminoglycosides- genetamcin– Blocks binding of tRNA-AA to 30S
• tetraclyclines
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Inhibition of Translation
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Translation
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Inhibition of Peptide Bond
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Inhibition of Ribosome Movement
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Inhibition of tRNA Attachment
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Misreading mRNA
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Inhibit Nucleic acid Synthesis
Quinolones – Ciprofloxacin and other quinolones
• Inhibits DNA gyrase
• Blocks DNA replication
• Inhibits mitochondrial DNA– conc in tissues too low for toxicity
• Urinary and intestinal infections
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Inhibition of DNA Replication
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Inhibit Nucleic Acid Synthesis
• Rifamycin– Inhibits DNA dependent RNA polymerase– Blocks transcription DNA ->RNA
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Acylovir vs Deoxyguanosine
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Inhibition of Transcription
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Antimetabolites• Sulfonamides• Donald D. Woods• Sulfanilamide blocks folic acid
– folic acid is essential to the synthesis of DNA and RNA
– Para amino benzoic acid (PABA) not incorporated into folic acid
• Reversible inhibition– High [PABA] competitively inhibit sulfanilamide
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Inhibited metabolites,Synthesis
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Drug Resistance
• synthesis of enzymes that inactivate the drug
• decrease in cell permeability and uptake of the drug
• change in the number or affinity of drug receptor sites
• modification of an essential metabolic pathway
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Development of Drug Resistance
• intrinsic– chromosomal mutations - low probability
• acquired– transfer of extra chromosomal DNA from a
resistant species to a sensitive one– Plasmids– Transposons
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Plasmids
– resistance factors or R factors transfered by conjugation, transformation or transduction
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Transposons
– sequences that can move from – plasmid >> chromosome – plasmid>> plasmid
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Clinical Trials• patient - has a diagnosed infection - two possibilities:
– a) the new drug is the drug of choice by testing– b) the patient has not responded to other drugs and the new drug
is testing well in the lab
• samples of blood etc. taken to determine all the possible parameters:– level of antimicrobial and presence of agent– cultures of infecting agent taken 2 times per day
• disappearance of the bacteria and patient recovery conclude a successful trial
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Reference: lab manual p. 270
Minimum Inhibitory Concentration• 1. test for antimicrobial activity
• 2. dilute antibiotic (pictures of tubes)
• 3. range selected obtained from therapeutic index
• 4. add to medium
• 5. add pure culture of isolated bacteria
• 6. incubate - tubes that are clear after 16 hours incubation at 35° C are subcultured– 0.1 ml removed and plated on suitably rich medium -
usually the agar version of the liquid growth medium
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Reference: lab manual p. 270
Kirby-Bauer Plate Sensitivity• disks impregnated with various concentrations of
appropriate antibiotics are placed aseptically on innoculated plates
• measurement of drug concentrations in the blood preclinical trials
• subjects receive varying dose levels and intervals of dosage
• pretrials usually determine the route of entry - oral or parenteral (injected subcutaneously, intramuscularly, etc.)
• pretrials determine the carrier substance
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Subjects Tested for Antimicrobial Levels
• blood, lymph, urine, feces tested for effective levels depending on disease
• also of concern is rapid metabolism (catabolism) of the drug and also rapid excretion
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Disk Diffusion Tests
• diffusion of antibiotic from disk controlled by agar concentration
• Zone of Inhibition– controlled by diffusion rate– level of sensitivity
• each antibiotic is unique