Comparative inhibition of beta-lactamases by cephamycin antibiotics.

Cefoxitin (CFX), cefmetazole (CMZ) and YM093301), and 6059-S2) have a 7a-methoxyl group (a unique structural feature of the cephamycin family) in the cephalosporin nucleus and 1oxa cephalosporin nucleus, respectively (Fig. 1). These compounds are resistant to hydrolysis by various bacterial i3-lactamases. This resistance character also provides the antibiotic with a broad antibacterial spectrum, including activity against indole-positive Proteus strains, Serratia spp. and Bacteroides spp. (including B. fragilis and all its subspecies) as well as against all the bacterial pathogens normally sensitive to the cephalosporins. There is great interest in antibiotics that are resistant to (3-lactamase hydrolysis and in compounds that are inhibitors of (:-lactamases3,4,5). CFX and CMZ, cephamycin antibiotics, have been reported to be potent competitive inhibitors of some (3-lactamases6,7). We wished to assess the (3-lactamase-inhibiting activity of the compounds of the cephamycin family. We have investigated the chromosomallymediated jS-lactamase-inhibiting activity of CFX, CMZ, YM09330 and 6059-S by determining the inhibitor constant (Ki) with cephalothin (CET) as a substrate. The plasmid-mediated penicillinase-inhibiting activity of CFX, CMZ, YM09330 and 6059-S was investigated by determining the Ki with cephaloridine (CER) as a substrate. The maximal rate of reaction (Vmax) and the dissociation constant (Km) were derived from a LINEWEAVER-BURK plot by standard procedures. The minimal inhibitory concentrations (MICs) of the strains were determined by an agar dilution technique. For the Enterobacteriaceae and Pseudomonas strains, the test medium was MUELLER-HINTON agar (Nissui Pharm. Co., Ltd.). For B. fragilis, the test medium was GAM agar (Nissui Pharm. Co., Ltd.). Plates were inoculated with one loopful of 106 colony-forming units (CFU) per ml. Anaerobic incubation was for 18 hours at 37°C in a Gas-Pak jar. Chromosomally-mediated (3-lactamases were extracted and purified from Escherichia coil GN5482, Enterobacter cloacae GN7471, Citrobacter freundii GN7391, S. marcescens GN10857, P. aeruginosa GN10362, P. rettgeri GN4430, P. morganii GN5407, P. vulgaris GN7919, P. cepacia GN 11164 and B. fragilis GN11477. Plasmid-mediated penicillinase type I (Rms212), type II (Rms213) and type III (Rte16)8,9,10) were extracted and purified from the transconjugants of these plasmids in E. coli W3630. Plasmid-mediated Fig. 1. Chemical structures of cefoxitin, cefinetazole, YM09330, and 6059-S.