Synthesis, characterization and antibacterial activity of a tridentate Schiff base derived from cephalothin and sulfadiazine, and its transition metal complexes.

Metal(II) coordination compounds of a cephalothin Schiff base (H2L) derived from the condensation of cephalothin antibiotic with sulfadiazine were synthesized. The Schiff base ligand, mononuclear [ML(H2O)3] (M(II)=Mn,Co,Ni,Zn) complexes and magnetically diluted dinuclear copper(II) complex [CuL(H2O)3]2 were characterized by several techniques, including elemental and thermal analysis, molar conductance and magnetic susceptibility measurements, electronic, FT-IR, EPR and (1)H NMR spectral studies. The cephalothin Schiff base ligand H2L behaves as a dianionic tridentate NOO chelating agent. The biological applications of complexes have been studied on two bacteria strains (Escherichia coli and Staphylococcus aureus) by agar diffusion disc method.

Structure-based optimization of cephalothin-analogue boronic acids as beta-lactamase inhibitors.

Boronic acids have proved to be promising selective inhibitors of beta-lactamases, acting as transition state analogues. Starting from a previously described nanomolar inhibitor of AmpC beta-lactamase, three new inhibitors were designed to gain interactions with highly conserved residues, such as Asn343, and to bind more tightly to the enzyme. Among these, one was obtained by stereoselective synthesis and succeeded in placing its anionic group into the carboxylate binding site of the enzyme, as revealed by X-ray crystallography of the complex inhibitor/AmpC. Nevertheless, it failed at improving affinity, when compared to the lead from which it was derived. The origins of this structural and energetic discrepancy are discussed.

Microbial production of 7-amino-cephalosporanic acid and new generation cephalosporins (cephalothin) by different processing strategies.

The development of beta-lactam antibiotics has been a continuous battle of the design of new compounds to withstand inactivation by the ever-increasing diversity of beta-lactamases. Semisynthetic cephalosporins like cephalothin were synthesized from 7-amino-cephalosporanic acid (7-ACA), and thiophene-2-acetic acid using cephalosporin-C acylase enzyme was studied. The production of cephalosporin-C acylase by Pseudomonas diminuta was used and the growth kinetics studied. The optimum condition of enzyme activity was determined by using response surface methodology. A 2(3) full-factorial composite design was employed for experimental design and the result analyzed. The pH value and temperature for optimum activity were 6.5 and 32 degrees C, respectively. The structural analog compound similar to the side-chain of semisynthetic cephalosporins, e.g., thiophene-2-acetic acid, was added. HPLC data analysis indicate that the concentration of cephalothin was 1.6 mg/mL.

Derivatives of cephalothin that inhibit ampicillin resistant Escherichia coli.

Two derivatives of cephalothin, compound I and II, were synthesized and demonstrated strong growth inhibition of ampicillin resistant Escherichia coli (E. coli). Compound I is the propyl ester of the parent cephalothin antibiotic, while compound II is the butyl ester derivative. The ester substituent replaces the former carboxyl group of cephalothin. Compounds I and II are stable at room temperature and have increased lipophilicity compared to cephalothin due to the presence of the ester substituent. The MIC50 of I and II were determined to be 55 microg/mL and 30 microg/mL, respectively. Cephalothin showed less than 20% growth inhibition of E. coli at all concentrations based on assay of colony forming units. Compounds I and II showed greater than 50% growth inhibition of E. coli at all concentrations greater than 50 microg/mL (more than 65% at 400 microg/mL). Pharmacological properties such as octanol/water partition Log P and polar surface area were determined. Values for polar surface area suggested 35% of I or II present in the intestinal system would be absorbed. The Log P values for I and II are 2.061 and 2.62, respectively, which indicate I and II will penetrate the blood-brain barrier more effectively than cephalothin. The lipophilic substituent constant (pi) for I and II are 1.592 and 1.95, respectively, which indicates the ester substituents contribute a strong lipophilic trait. Properties of molar refractivity, parachor, and molar volume, which describe van-der-Waals interactions, are also determined.

Dynamic reaction design of enzymic biotransformations in organic media: equilibrium-controlled synthesis of antibiotics by penicillin G acylase.

Parameters relevant to the thermodynamically controlled synthesis of cephalothin utilizing highly active stabilized penicillin G acylase derivatives were studied. These included solubility/stability of substrates, enzyme derivative activity/stability, reaction course and synthetic yields. These parameters were altered by varying the pH, dimethylformamide concentration and temperature. Simultaneous optimization of the selected parameters could not be achieved with a single set of conditions. However, continuous adjustment of conditions throughout the reaction course allowed each parameter to be optimized (dynamic reaction design). This strategy works by optimizing those parameters that are critical to the overall reaction at a given point, whilst leaving others sub-optimal when their contribution to the total is minimal. This strategy has achieved a 90% transformation of antibiotic nucleus to cephalothin at a final concentration of 20 g/l, high enzyme and reactant stability, with a reaction period of 3 h (using 1 ml of derivative/40 ml of reaction solution).

Synthesis and antibacterial activity of new C-10 quinolonyl-cephem esters.

A series of cephalosporins derived from cephalothin containing an ester-linked quinolonyl substituent at the C-10 position (C-10 quinolonyl-cephem esters) has been prepared and evaluated for in vitro antibacterial activity. The C-10 quinolonyl-cephem esters exhibited a broadened spectrum of activity when compared with cephalothin and the corresponding quinolones, including activity against beta-lactamase-producing bacteria.

Monitoring beta-lactamase activity in vivo by 13C nuclear magnetic resonance spectroscopy.

A 13C-labeled cephalothin, 7 beta-(2-thienylacetamido)-3-[acetoxy-13C1]methyl-3-cephem-4- carboxylate (compound 1), has been prepared and used to monitor beta-lactamase activities by 13C nuclear magnetic resonance spectroscopy. Time-elapsed spectral analysis of the reaction of the labeled cephalothin with the TEM-2 beta-lactamase purified from Escherichia coli revealed the progressive loss of the cephalothin acetyl resonance at 176.8 ppm and accumulation of an acetate signal at 184.3 ppm. Spectral results identical to those observed in the in vitro experiment were obtained when compound 1 was incubated with cell suspensions of E. coli JSR-O (pBR322), which contains the plasmid-encoded TEM-2 beta-lactamase, and Enterobacter cloacae strains that contain a class I chromosomal beta-lactamase. Pseudo-first-order rate constants for the lactamase-catalyzed formation of acetate from cephalothin in vivo were obtained by integration of the 13C-acetyl resonances of compound 1 during timed incubations with cell preparations. These results constitute the first demonstration of the ability to monitor beta-lactamase activity in viable cells by nuclear magnetic resonance spectroscopy.

Coupling products of amino acids to penicillin V and cephalothin: synthesis and susceptibility to carboxypeptidases and lysosomal enzymes.

Amino acids have been coupled to the carboxyl group of penicillin V and cephalothin by methods that keep the beta-lactam ring intact. Derivatives were successfully obtained with both neutral (Leu, Val, Ala, Ile, Trp, Tyr, Gly) and one acidic (Glu) amino acids. The new compounds were inactive in vitro against Staphylococcus aureus or Micrococcus luteus. Incubation in the presence of purified carboxypeptidases (A, B), soluble lysosomal fractions from liver, or cellular homogenates from liver, kidney, fibroblasts, and macrophages did not allow recovery of the antibacterial activity. Injection in mice also failed to cause liberation of microbiologically active compounds. HPLC studies confirmed that the amide linkage between the antibiotic and the amino acid was not hydrolyzed in the presence of soluble lysosomal fractions from liver. However, conversion of cephalothin and cephalothin-leucine to desacetyl derivatives was observed in the presence of soluble lysosomal fractions and extracts from liver and semipurified orange peel acetylesterase(s). It is concluded that amino acid derivatives of beta-lactam antibiotics do not offer potential chemotherapeutic use as prodrugs.

7-(D,L-alpha-fluoro-2-thienylacetamido)cephalosporanic acid (L 14655), a new semisynthetic cephalosporin. Synthesis and preliminary biological evaluation.

We describe the synthesis and some preliminary antimicrobial and enzymatic properties of 7-(D,L-alpha-fluoro-2-thienylacetamido)cephalosporanic acid (L 14655), a fluorinated derivative of cephalothin. L 14655 had antimicrobial activity similar to that of cephalothin. Cell-free studies demonstrated that L 14655 was very stable to hydrolysis by type Ia beta-lactamase (325 times more than cephalothin) but not to TEM enzyme. In addition, L 14655 was 480 times more active than cephalothin in preventing the hydrolysis of nitrocephin by type Ia enzyme. Preliminary studies in intact beta-lactamase-producing cells did not however produce evidence of enhanced activity of L 14655 relative to cephalothin.