Independent Antibiotic Actions of Hetacillin and Ampicillin Revealed by Fast Methods

Abstract

Fast methods for measuring inhibition of growth of Escherichia coli or Staphylococcus aureus by hetacillin or ampicillin showed that the rate of inhibition was a function of the antibiotic concentration. When ampicillin concentrations were measured rapidly, hetacillin was shown to have a half-life of 20 min at 37 C and pH 6.7. Hetacillin was much less susceptible to penicillinase (f8-lactamase) than was ampicillin. An inhibitory effect of hetacillin was shown in the presence of 1l-lactamase, which destroyed ampicillin activity very rapidly.Hetacillin is an acetone-condensation derivative of ampicillin (1, 6). It is unstable under conditions of acidity and temperature suitable for antibiotic action, breaking down to ampicillin and acetone (2, 5, 6). The half-life of hetacillin at 37 C at pH 7.0 has been estimated at 15 to 30 min (2, 6). In none of the previous studies could the authors demonstrate an independent antibiotic action of hetacillin which could not possibly be attributed to ampicillin.In this study, very rapid, instrumented methods were used to investigate the general problem of how to measure antibiotic activity of an unstable compound whose half-life is not much longer than the replication time of the test bacteria and whose breakdown product has an antibacterial action similar to that of the parent antibiotic. To measure bacterial growth and inhibition, a culture was prepared in its logarithmic growth phase by incubating 0.1 ml of an overnight meat infusion broth culture in 50 ml of the same medium at 37 C for 1.5 hr. A volume of 0.25 ml of antibiotic solution at 10 times the final concentration to be used and 0.1 ml of either 0.05 M phosphate buffer (pH 6.7) or a 1:10 dilution of ,-lactamase (2.5 units) were incubated for 5 min in a 1-cm spectrophotometer cuvette. Then 2.25 ml of the log-phase culture was added, containing between 2 x 106 and 4 x 106 bacteria/ml (viable count, corresponding to an optical density of 0.064 for E. coli 014 and 0.060 for S. aureus S1). The cuvette was incubated at 37 C in a water-jacketed holder in a Unicam SP600 series 2 spectrophotometer, and the optical density at 540 nm was recorded on a Watanabe X-YT/XT recorder (model WX431) fitted with a roll chart moving at 2 mm/min. In some experiments, the output from the spectrophotometer was amplified 10 times with a Keithley model 604 differential amplifier before recording. A similar control culture without antibiotic was incubated simultaneously, and recordings of optical density were made alternately from the test and the control for periods of approximately 1 or 2 min throughout a total time of 10 min to 4 hr in different experiments. MATERIALS AND METHODSAmpicillin was measured by adapting the penicillinase iodometric titration method (4) from the micromethod modification of Novick (3). Penicillinase (,-lactamase) reacts with ampicillin to produce penicilloic acid, which decolorizes starch-iodine reagent at a rate corresponding to its concentration. Starchiodide reagent was prepared from a solution of 0.08 M I, in 3.2...

Merging Electrochemical Synthesis and Post-Ugi Cyclization for the Synthesis of Diverse 4-Imidazolidinones

“…4-Imidazolidinones are ubiquitous heterocycles found in many biologically important molecules . Particularly, the spirocyclic variant of 4-imidazolidinones is found in several marketed drugs such as hetacillin, NNC 63-0532, spiperone, and spiroxatrine . More Precisely, MacMillan’s imidazolidin-4-ones organocatalysts are metal-free organocatalysts, widely used in enantioselective organic transformations (Scheme ).…”

Geminal Diheteroatomic Motifs: Some Applications of Acetals, Ketals, and Their Sulfur and Nitrogen Homologues in Medicinal Chemistry and Drug Design

“… The stability of ( N , N )-aminals can be enhanced by incorporating electron-withdrawing groups at C2 or acylating either or both of the basic amines. However, the effect on stability is context-dependent, as exemplified by the cyclic acylated aminal hetacillin ( 583 ), a prodrug of ampicillin ( 585 ), which is unstable in aqueous solution with a half-life of 15–30 min at 37 °C and pH = 7 . The geminal dimethyl groups at C2 contribute to the instability of 583 by stabilizing the carbonium ion intermediate 584 (Scheme ).…”