A simple method for testing the efficacy of a beta-lactamase inhibitor against beta-lactamase-producing gram-negative bacteria.

The synergistic effect of a beta-lactam antibiotic in combination with a beta-lactamase inhibitor (cloxacillin) on Gram-negative bacteria producing various types of beta-lactamases was studied with respect to the properties of the enzymes by using a set of bacterial stains we had developed previously for evaluation of beta-lactamase stability of beta-lactam antibiotics). Cloxacillin showed strong synergy in combination with ampicillin or cephalosporins against typical cephalosporinase-producing bacteria, but not against penicillinase-producing bacteria. Analysis of the results indicated that the efficacy of a competitive inhibitor such as cloxacillin is greatly influenced by the following factors: the affinity of the inhibitor for the enzyme, the difference between the affinity of the inhibitor for the enzyme and that of the combined antibiotic for the enzyme, and the ability of the inhibitor to penetrate the bacterial outer membrane. It was also demonstrated that this information concerning the efficacy of an inhibitor could easily be obtained by using the set of bacterial strains and the methods described.