[Detection of β-Lactamase Produced by Klebsiella pneumoniae Using High Performance Liquid Chromatography].

Abstract

In order to develop a new method to detect β-lactamase, we determined degradation of β-lactam antibiotics, ampicillin sodium (ABPC) and cefotaxime sodium (CTX), by β-lactamase-producing Klebsiella pneumoniae (K. pneumoniae) in terms of high performance liquid chromatography (HPLC). Using HPLC with an ODS column and an eluent composed of phosphate buffer and methanol, we could detect ABPC and CTX within 10 min. After cultured with K. pneumoniae, ABPC and CTX were degraded. The degradation rate corresponding to the rate of peak area incubated with and without bacteria increased with increasing McFarland No. of bacterial suspension and incubation time. Under the culture condition of McFarland No. 3.0 and 90-min incubation, the degradation rate of ABPC and CTX was 52.6 and 70.8% by K. pneumoniae, whereas it was less than 10% by susceptible bacteria. Consequently, these results confirmed that β-lactamase produced by ABPC- and CTX-resistant bacteria could be detected within about 120 min through HPLC measurement.