Download A. Penicillins

Dr. Bilal Aljaidi
Antibacterial agents:
 Antibiotic: any substance produced by a microorganism
that is antagonistic to the growth of other
microorganisms (mainly bacteria) in high dilution
(Waksman 1942).
 Antimicrobial agent:
 is a substance that kills or inhibits the growth of
microorganisms such as bacteria, fungi, or protozoans.
 Either kills the microbe (microbiocidal) or prevent its
growth (microbiostatic).
 covers both antibiotics and synthetic agents
Antibacterial agents:
 Infection is the colonization of the host organism with
a microorganism like bacteria, parasite, virus, or even a
macro organism like fungi and macro parasites such as
worms and nematodes. The microorganism then will
use the host resources to reproduce and grow that
results in a disease.
 Host system normally use the immune system to fight
against the invading organism, first by the innate
immune system, then by the adaptive immune system.
History of Antibacterial agents:
 The presence of bacteria was first identified in 167o by Van
leeuwenhoek.
 Pasteur was the first who link the bacteria to disease in
1800.
 Lister introduced carbolic acid as antiseptic and sterilizing
agent for the operating wards.
 Koch identified that some bacteria is responsible for
specific infections such as tuberculosis, cholera and
typhoid
History of Antibacterial agents:
 Paul Ehrlich “ the father of chemotherapy” developed
the principle of chemotherapy: a chemical could
directly interfere with the proliferation of
microorganism at a concentration tolerated by the
host .. This was named the magic pullet.. Which is
nowadays called the selective toxicity.
 In 1910, Ehrlich had developed the first synthetic
antibacterial agent; Salvarsan, which is active against
protozoa especially trypanosoma.
History of Antibacterial agents:
 In 1934, Proflavine was used in the wound infections,
due to its systemic toxicity, it did not used systemically.
 In 1935, Prontosil was discovered as effective
antibacterial agent in vivo. later found to be a prodrug
that release sulfanilamide as the active metabolite:
History of Antibacterial agents:
 Sulfonamides were the only effective antibacterial
agents until the discovery of Penicillins in 1940.
 Streptomycin was discovered in 1945 and used against
tuberculosis and other gram –ve bacteria, then lead to
the discovery of other aminoglycosides.
Streptomycin
History of Antibacterial agents:
 Chloramphenicol and tetracycline antibiotics were
discovered in 1947.
 In 1955, Cephalosporins were discovered.
 Many synthetic antibacterial agents were prepared later
such as quinolones (1962), Flourinated quinolones
(1980),Linezolid (2000) and other sulfonamide derivatives
with a broad spectrum activity.
 The extensive studies on the bacterial cell
components, genome and metabolic pathways
considerably helped in better understanding the
essential metabolic stages that are important for the
growth and proliferation of the bacterial cells, which
means that targeting these pathways might result in
killing or attenuating the bacterial growth.
 Also the absence of certain essential bacterial
metabolic pathways means that targeting such
reaction could result in selective bacterial killing
without harming human cells and result in safe
antibacterial agents
Bacterial cell
 Is a prokaryotic cell.
 Differ significantly from eukaryotic cells:
 1-10 µm length whereas eukaryotic length is 10-100 µm
 Has no nucleus.
 Circular DNA, no chromosomal structure.
 Most of the organelles are simpler that eukaryotics.
 Characteristic cell wall which differ from bacteria to
another, but generally, it is thick and fatty envelope
which protect the bacterial cell from lysis and invading
by external environment.
Potential targets for antibacterial
agents
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Protein synthesis
Nucleic acid synthesis
Cell metabolism (e.g. folate synthesis)
Cytoplasmic membrane
Bacterial cell wall synthesis
Potential targets for antibacterial
agents
Sulfonamides
(on metabolic enzymes)
Penicillins
Cephalosporins
Aminoglycosides
Tetracycline
Chloramphenicol
Quinolones
Rifampicin
Penicillins
Cephalosporins
Cell wall
Monobactams
synthesis
Carbapenems
Cycloserine
Vancomycin
Teicoplanin
Bacitracin
RNA elongation
Dactinomycin
Ansamycins
Protein synthesis
(tRNA)
Cytoplasmic
membrane
structure Polymixins
Mupirocin
Protein synthesis
(30S inhibitors)
Tetracyclines
Aminoglycosides
Antibacterial agents acting on
the cell wall biosynthesis
Penicillins and Cephalosporins
b-lactam antibiotics
A
5-membered thiazolidine ring
6-membered dihydrothiazine
B
Penicillin nucleus
Carbon atom
C
Cephalosporin nucleus
D
monocyclic
Carbapenem nucleus
Monobactam nucleus
Penicillins
Penicillins
 Highly strained structure due to the presence of
bicyclic fused system composed from four membered
lactam ring fused to the five membered thiazolidine
ring.
 Bacteria synthesizes penicillin using cysteine, valine
and some of the fermentation products:
Penicillins
 Difficult to synthesize in the lab due to:
 The unstable highly strained ring system.
 The three chiral centre it has which should be with
certain stereochemistry.
 Beechams was successfully isolated the biosynthetic
precursor; 6-APA that was used as an intermediate for
the synthesis of most of the semisynthetic penicllins.
The bacterial cell wall
 Peptidoglycan = a vital component of bacterial cell walls,
responsible for its shape and integrity
 Peptidoglycan = macromolecule made of sugar (glycan) chains
cross-linked by short peptide bridges
 The nature of the peptidic cross links varies among bacteria
but the essential mechanism is similar
The bacterial cell wall
Peptidoglycan
N-acetylglucosamine (NAG)
Gram –
Only two layers of
peptidoglycan
Gram +
Consists of 50-200
peptidoglycan
layers
Transpeptidase
Involved in crosslinking
N-acetylmuramic acid (NAM)
D-alanine
+
D-ala D-ala (natural substrate)
Penicillins
Bacterial cell lysis
Excellent selective toxicity
Penicillin-enzyme complex
cross-linking inhibited
The wall become fragile
and can no longer
prevent the cell from
swelling and bursting
 Penicillin mimic the structure of D-ala-D-ala, because
of that the transpeptidase mistakenly bind to it
instead of D-ala-D-ala.
 Also this explains the lack of penicillin toxicity, since
D-amido acids are not present in human, only the Lamino acids present.
 Also targeting the cross linking in the peptidoglycan
biosynthesis which is only present in bacteria explains
the selective toxicity.
Structure-activity relationships of
penicillins (SAR)
 The strained β-lactam ring is essential.
 The free carboxylic acid is essential (the carboxylate
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ion binds to the charged nitrogen of the lysine at the
active site.
The bicyclic system is essential.
The acylamino side chain is essential.
Sulfur is not essential.
The stereochemistry of the bicyclic ring with respect
to the acylamino side chain is important.
Structure-activity relationships of
penicillins
Acid sensitivity of penicillins
Acid sensitivity of penicillins
 Three reasons for the acid sensitivity of penicillin G:
 Ring strain: due to the large angle and torsional strain
exist, acid catalyzed ring opening will relief these
strains.
 A highly reactive β-lactam carbonyl group:

This amide bond is exceptionally unstable compared to the
normal amide (why?).. The stabilization of the amide bond by
the resonance is impaired here due to the increase in the ring
strain that will be formed after the delocalization of the
nitrogen lone pair to form a double bond within the four
memeberd lactam ring
Acid sensitivity of penicillins
 The effect of the acyl side chain:
 It has a self-destructive mechanism in which the oxygen
of the carbonyl group will attack the carbonyl carbon of
the lactam ring causing the ring opining just like the
attack of water previously mentioned.
 This gives Penillic acid and penicillenic acid as final
products (explain the mechanism of formation?)
Acid resistant Penicillins
 To reduce the acid instability of penicillins:
 We can not change the β-lactam.
 We can not change the bicyclic system and its ring
strain.
 The only thing that can be modified is the acyl group in
order to reduce the self destructive mechanism (How?).
This can be done by decreasing the nucleophilicity of
the carbonyl oxygen which can be done by adding
electron withdrawing group.
Acid resistant Penicillins
β-lactamase (Penicillinase):
 The wide spread use of penicillin G led to the increase in
the number of resistant strains, especially in S. aureus.
 The main mechanism of resistance is the production and
secretion of β-lactamase enzyme.
 β-lactamase is a mutated version of transpeptidase which is
closely related in structure, especially in the active site.
This means that β-lactamase will interact with penicillin
structure in the same manner as transpeptidase.
 β-lactamase can hydrolyze 1000 penicillin molecule per
second because the cleaved penicillin will leave the active
site to react with other molecule.
β-lactamase (Penicillinase):
 Gram +ve bacteria normally release β-lactamase to outside
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of the cell that will cleave penicillin before reaching the
bacteria.
Gram –ve bacteria release β-lactamase into the periplasmic
space, which again will cleave penicillin before reaching the
plasma membrane.
Penicillin has to reach the plasma membrane where the
transpeptidase present to do its antibacterial action.
95% of S.aureus became resistant to penicillins
Most of gram –ve bacteria are β-lactamase producing
bacteria
β-lactamase (Penicillinase):
 There are various types of β-lactamase enzymes:
 Some are selective against penicillins (penicillinase).
 Some are selective against cephalosporins
(cephalosporinase).
 Some are non-selective, acting on penicillins and
cephalosporins at the same time.
β-lactamase resistant Penicillins
 Structural modification has to be made to penicillin
structure to prevent the binding to β-lactamase active
site.
 This was a difficult task since β-lactamase and
transpeptidase have a very similar active site.
 The only successful modification was by adding bulky
group at the acyl side chain (Steric Shield), but not too
bulky because this proved to reduce penicillin activity.
β-lactamase resistant Penicillins
95% of S.aureus in hospitals
became resistant to methicllin
(MRSA) which in turns are resistant
to most of the older generation
penicillins
Isoxazoyl Penicillins
 Effective against β-lactamase producing strains.
 Acid stable due to the electron withdrawing effect of
the isoxazole ring.
 Used against S.aureus resistant bacteria.
Broad Spectrum Penicillins
 Factors affecting the extent of penicillin activity:
 Chemical structure.
 The ability to cross the cell wall.
 Their susceptibility to β-lactamase .
 Their affinity to transpeptidase enzymes.
Broad Spectrum Penicillins
 Chemical modification to get broad spectrum
penicillins:
1.
2.
Addition of hydrophobic group at the acyl side chain
found to improve activity against gram +ve but reduce
it against gram –ve bacteria.
The addition of hydrophilic group at the acyl side
chain causes a reduction in activity.
Broad Spectrum Penicillins
3.
Activity on gram –ve enhanced when a hydrophilic
group was added at the -carbon in the acyl side
chain, such in ampicillin and amoxicillin. This polar
groups help the penicillin to pass through the
porines that exist in the cell envelope of the gram –
ve bacteria
Ampicillin and amoxicillin
 Broad spectrum penicillins.
 Acid resistant penicillins (why?).
 Sensitive to β-lactamase (why?).
 Poorly absorbed through the mucus memebrane, this
is due to the fact that they formed a zwitter ionic
molecule at physiological pH (they contain a
carboxylic acid and an amino group in their structure.
 The oral bioavailability can be improved by masking one
of them, mainly the carboxylic acid…. By preparing a
prodrug esters.
Ampicillin prodrugs
 The methyl ester did not give the same improvement in absorption and activity (why?).
Ureidopenicillins
 They all have a urea group at the -carbon in the acyl side
chain.
 They have better activity
compared to amoxicillin
and they are more resistant to
β-lactamase.
 Used parenterally for gram –ve
infections especially P.aeruginosa.
 the ureido group though to mimic
Some of the peptidoglycan structure,
which means that it can bind to
penicillin-binding protein
Carboxypenicillins
 They have a carboxylic acid at the -carbon of the acyl
side chain.
 They have broad spectrum activity.