Penam Nucleus Research Articles

Novel Penicillin Analogues as Potential Antimicrobial Agents; Design, Synthesis and Docking Studies.

A number of penicillin derivatives (4a-h) were synthesized by the condensation of 6-amino penicillinic acid (6-APA) with non-steroidal anti-inflammatory drugs as antimicrobial agents. In silico docking study of these analogues was performed against Penicillin Binding Protein (PDBID 1CEF) using AutoDock Tools 1.5.6 in order to investigate the antimicrobial data on structural basis. Penicillin binding proteins function as either transpeptidases or carboxypeptidases and in few cases demonstrate transglycosylase activity in bacteria. The excellent antibacterial potential was depicted by compounds 4c and 4e against Escherichia coli, Staphylococcus epidermidus and Staphylococcus aureus compared to the standard amoxicillin. The most potent penicillin derivative 4e exhibited same activity as standard amoxicillin against S. aureus. In the enzyme inhibitory assay the compound 4e inhibited E. coli MurC with an IC50 value of 12.5 μM. The docking scores of these compounds 4c and 4e also verified their greater antibacterial potential. The results verified the importance of side chain functionalities along with the presence of central penam nucleus. The binding affinities calculated from docking results expressed in the form of binding energies ranges from -7.8 to -9.2kcal/mol. The carboxylic group of penam nucleus in all these compounds is responsible for strong binding with receptor protein with the bond length ranges from 3.4 to 4.4 Ǻ. The results of present work ratify that derivatives 4c and 4e may serve as a structural template for the design and development of potent antimicrobial agents.

Improved X-ray diffraction from Bacillus megaterium penicillin G acylase crystals through long cryosoaking dehydration.

Penicillin G acylase from Bacillus megaterium (BmPGA) is currently used in the pharmaceutical industry as an alternative to PGA from Escherichia coli (EcPGA) for the hydrolysis of penicillin G to produce 6-aminopenicillanic acid (6-APA), a penam nucleus for semisynthetic penicillins. Despite the significant differences in amino-acid sequence between PGAs from Gram-positive and Gram-negative bacteria, a representative PGA structure of Gram-positive origin has never been reported. In this study, crystallization and diffraction studies of BmPGA are described. Poor diffraction patterns with blurred spots at higher resolution were typical for BmPGA crystals cryocooled after a brief immersion in cryoprotectant solution. Overnight soaking in the same cryo-solution substantially improved both the mosaicity and resolution limit through the establishment of a new crystal-packing equilibrium. A crystal of BmPGA diffracted X-rays to 2.20 Å resolution and belonged to the monoclinic space group P2(1) with one molecule of BmPGA in the asymmetric unit.

Serum leptin levels reduced in non-obese type 2 diabetic patients

A generic enzyme immunoassay (EIA) for penicillins containing the intact penam nucleus (PEN EIA), and an EIA for penicillin G derivative after alkaline hydrolysis of the betalactam ring (benzylpenicilloic acid, BPO EIA) were used to analyse raw goat milk. The detection limit for penicillin G in the PEN EIA was at 0.2–0.4 ng/ml, the recoveries in artificially contaminated goat milk samples (2–32 ng/ml) were at 90–140% (RSD 25–37%). The detection limit for hydrolysed penicillin G in the BPO EIA was at 1–2 ng/ml, the recovery in artificially contaminated goat milk (4–16 ng/ml) was at 95–123% (RSD 12–30%). A total of 577 goat milk samples (composite samples) collected from 13 goat milk farms in Hesse, Germany, were analysed for penicillins. Nine samples were positive in the PEN EIA, but at low levels (1 ng/ml to <4 ng/ml) in seven samples. One sample was positive at a level of 7 ng/ml, and also contained BPO at a concentration of 4.6 ng/ml. One sample yielded extremely high concentration levels in the PEN EIA (mean: 80.000 ng/ml) and in the BPO EIA (mean: 30.000 ng/ml). The relatively high levels of BPO indicate that penicillin G was used by parenteral but not intramammary application. The results show that the immunochemical methods are applicable to analyse goat milk, and that penicillin G may occasionally occur in goat milk samples in Hesse.

Determination of penicillin and benzylpenicilloic acid in goat milk by enzyme immunoassays

A generic enzyme immunoassay (EIA) for penicillins containing the intact penam nucleus (PEN EIA), and an EIA for penicillin G derivative after alkaline hydrolysis of the betalactam ring (benzylpenicilloic acid, BPO EIA) were used to analyse raw goat milk. The detection limit for penicillin G in the PEN EIA was at 0.2–0.4ng/ml, the recoveries in artificially contaminated goat milk samples (2–32ng/ml) were at 90–140% (RSD 25–37%). The detection limit for hydrolysed penicillin G in the BPO EIA was at 1–2ng/ml, the recovery in artificially contaminated goat milk (4–16ng/ml) was at 95–123% (RSD 12–30%). A total of 577 goat milk samples (composite samples) collected from 13 goat milk farms in Hesse, Germany, were analysed for penicillins. Nine samples were positive in the PEN EIA, but at low levels (1ng/ml to <4ng/ml) in seven samples. One sample was positive at a level of 7ng/ml, and also contained BPO at a concentration of 4.6ng/ml. One sample yielded extremely high concentration levels in the PEN EIA (mean: 80.000ng/ml) and in the BPO EIA (mean: 30.000ng/ml). The relatively high levels of BPO indicate that penicillin G was used by parenteral but not intramammary application. The results show that the immunochemical methods are applicable to analyse goat milk, and that penicillin G may occasionally occur in goat milk samples in Hesse.

Exploring the chemistry of penicillin as a β-lactamase-dependent prodrug

The penam nucleus can be modified to behave as a beta-lactamase-dependent 'prodrug' by incorporation of a vinyl ester side chain at the 6-position. Enzyme-catalysed hydrolysis of the beta-lactam ring uncovers the thiazolidine-ring nitrogen as a nucleophile that drives a rapid intramolecular displacement on the side chain. Attachment of 7-hydroxy-4-methylcoumarin as the releasable group of this side chain generated a penicillin structure that can function as a fluorescence-based reporter substance/diagnostic for the presence of low levels of beta-lactamase enzyme in solution. Mechanistic details of the reaction pattern are documented and the scope and limitations of exploiting the structural modification are discussed.

Synthesis and elastase inhibitory activity of 6α-chloro-2,2-dimethyl-3α-(pivaloyloxy)methylpenam sulfone, 6α-chloro-2,2-dimethyl-3- exo-methylenepenam sulfone, benzyl and methyl 6α-substituted penicillanate sulfones

The triflates and pivalates of 3α-hydroxymethyl-6-substituted-2,2-dimethylpenam sulfones 3, 5; methyl and benzyl 6-substituted penicillanates 6–9 and 3- exo-methylene-6-substituted-2,2-dimethylpenam sulfone 4 were synthesized. These novel compounds were evaluated as elastase inhibitors using porcine pancreatic elastase. The effects that structural modifications of substituents on C-3 and C-6 in the penam nucleus have on elastase activity were examined and several similarities and distinctions were identified when compared to the reported penicillin esters and amides elastase inhibitors.

Stereospecific synthesis, 1H and 13C NMR spectroscopy, and X-ray crystallographic studies of 6,6-dibromo-3α-cyano-2,2-dimethylpenam-(1R)-S-oxide

A stereospecific synthesis of 6,6-dibromo-3α-cyano-2,2-dimethylpenam-(1R)-S-oxide (2) is described. The X-ray single crystal investigation of this compound shows that it crystallizes in the monoclinic system, space group P21, with cell parameters a = 6.194(1), b = 9.363(1), c = 9.869(1) Å, β = 96.55(2)°, V = 568(1) Å3, and two molecules per unit cell. Thestructure was solved, from low temperature data, by direct methods, and refined, by least-squares procedures, to a final R = 3.17%, Rw = 3.38%, for 1221 observed independent reflections. The sulfoxide configuration is R. The thiazolidine presents an envelope shape with the C(3) atom 0.56 Å above the other four members of the ring mean plane. Therefore, it exhibits the conformation with α-CH3 pseudoequatorial and β-CH3 and α-CN both pseudoaxial. This conformation is different from the one reported for other penicillin sulfoxide derivatives. The geometry of the β-lactam ring is typical of the fused bicyclic penam nucleus. The ring is non-planar. The N(4) atom displays a pyramidial arrangement. The examination of crystal packing shows that molecules along a are connected by a short C—H … O contactReported single crystal studies showed two dominant conformations for the thiazolidine ring in the fused bicyclic penam nucleus: the C(3) or 2β-3α substituents both pseudoaxial, as found in 2, and the S(1) or 2β-3α substituents both pseudoequatorial, adopted by most sulfoxides. Neither of these conformations can explain the nuclear Overhauser effect data. A conformation with the substituents C2-β-methyl-C3-α-H and C2-α-methyl-C3-α-CN fully eclipsed is in accordance with the experimental results in the solution. The 1H nuclear magnetic resonance data reflect the influence of the magnetic anisotropy of the CN triple bond on the gem-dimethyl groups, particularly in relation with the geometrical features of the molecule. Comparison of 13C nuclear magnetic resonance chemical shifts for 2 and its parent penicillin sulfide (1) indicates configurationally dependent substituent effects, the magnitude of which can be used for the stereochemical assignment of sulfoxide groups in this series. Key words: penicillin sulfoxide derivatives, di-halogen penicillanates, stereospecific synthesis, NMR studies, absolute configuration determination.

Zinc Mediated Reduction of Bromohydrins in Azetidinones and Penems. Application to the Synthesis of FCE 22891

A route to penems is described in which dehalogenative reduction of bromohydrins is carried out after ring opening of the penam nucleus. Reduction on some newly synthesized bromopenems suffeerd from low yields. Better yields and acceptable stereoselectivity were obtained on free-hydroxyethyl azetidinone substrates. Some parameters of the reduction are discussed

Antibacterial properties of (2,3)-alpha- and (2,3)-beta-methylene analogs of penicillin G.

The penam nucleus can assume two conformations; these are designated open and closed. The synthetic (2,3)-alpha- and (2,3)-beta-methylenepenams can be regarded as analogs of the open and closed conformations, respectively. It has been shown that the beta-methylenepenams are essentially inactive, suggesting that the closed conformation of penams is also inactive. In this study, we investigated a series of beta-lactams, all of which contained phenylacetamido side chains: penicillin G, the (2,3)-alpha- and (2,3)-beta-methylenepenams, and the 3-acetoxymethyl- and 3-methylcephalosporins. The alpha-methylenepenam and penicillin G were the most active compounds, while the beta-methylene isomer was only poorly active. Results with permeability mutants suggested that the alpha-methylene compound penetrated the outer membrane somewhat more readily than penicillin G did. The intrinsic potency of the alpha-methylenepenam appeared to be similar to that of penicillin G, on the basis of their affinities for penicillin-binding proteins and their abilities to inhibit peptidoglycan synthesis in ether-permeabilized Escherichia coli, while the beta-methylene analog had very poor intrinsic potency. The alpha-methylene analog was about 10-fold more efficient (Vmax/Km) than penicillin G as a substrate for the cephalosporinases from Enterobacter cloacae and Proteus vulgaris, but it was about 40-fold less efficient with penicillinase from Staphylococcus aureus. These results strongly support the hypothesis that the active conformation of penams is the open conformation and suggest that the position in space of the carboxyl group relative to the beta-lactam carbonyl is an important determinant of cephalosporinlike character, as distinct from penicillinlike character.

A multifaceted approach to the study of the side-chain conformation in beta-lactamase-resistant penicillins.

The resistance of some penicillins to beta-lactamase enzymes was previously attributed to the nature of their C(6) side chain. In order to find explicitly the influence of the conformation of this side chain in the enzymatic mechanism, we have analyzed by experimental and theoretical methods (X-ray diffraction, NMR, IR, PCILO) the molecular structure of six resistant penicillins and derivatives: oxacillin, cloxacillin, dicloxacillin, flucloxacillin, methicillin, nafcillin, cloxacillin sulfoxide, and oxacillinpenicilloic acid. X-ray crystallography of flucloxacillin and nafcillin is fully described. We observe that the side chains of these penicillins have no influence on the electronic properties of the penam nucleus but are much more rigid than in the sensitive ones. The molecular conformations are mostly governed by the nonbonded Van der Waals interactions and, in the oxacillins, partly by the conjugation between exocyclic groups. The lack of flexibility could result in a distorting effect on the structure of the active site of the beta-lactamase, leading to the deactivation of the enzyme.

Studies on the circular dichroism of penam and penam sulfoxides

Optical rotatory properties of models of 6-aminopenicillanic acid (6-AP A) as obtained from earlier molecular orbital and rotatory strength calculations are corroborated by new CD data for 6-AP A. The new spectra extend down to 180 nm, revealing a positive Cotton effect near 185 nm due to carbonyl π→π * transitions. The pH sensitivity of the optical activity of the β-lactam amide π→π * and the 203-nm n→π * transitions is explainable in terms of electrostatic effects and orbital interactions present in α-aminoketones and 4-membered rings which tend to flatten out the NH 2 group. These same interactions are responsible for the unusually low p K a of 4.75 for the amino group; (the carboxyl group of 6-AP A has a normal p K a of 2.5). The previous calculational procedures are extended to the sulfoxides of 6-AP A, and the predicted CD spectra are compared to new experimental curves and also to available data for penicillin sulfoxides. The MO's associated with the principal bands are illustrated with the aid of electron density maps. The positive Cotton effect observed and calculated near 230 nm is due to two charge transfer transitions within the asymmetric penam nucleus. Similar to the situation with the unoxidized penams, the sulfoxide of 6-AP A displays a very low p K a for the amino group (3.8) and a change toward a less positive CD band at 202 nm upon protonation of this group.