Discrimination of Aminoglycoside Antibiotics (Dihydrostreptomycin, Kanamycin, Fradiomycin) with Drug-inactivating Enzymes

Abstract

A simple method employing aminoglycoside-inactivating enzymes from Escherichia coil for identification of aminoglycosides (AGs) such as dihydrostreptomycin (DHSM), kanamycin (KM) and fradiomycin (FRM) in food was developed.The crude enzyme (S105) fraction was prepared by ultrasonic treatment of cells of drug-resistant E. coil, followed by centrifugation at 105, 000×g. DHSM was inactivated by the S105 fraction of SM-resistant E. coli in the presence of both adenosine triphosphate (ATP) and Mg2+, KM and FRM were also inactivated by the S105 fraction of KM-resistant E. coli in the presence of both ATP and Mg2+ Discrimination of KM from FRM was done by the inactivation of the former by commercial kanamycin-6′-acetyltransferase (AAC(6′)-1) in the presence of ATP, Mg2+ and acetyl CoA. KM was inactivated by the action of AAC(6′)-1, while FRM was not.Treatment of serum and kidney samples of rats, which had been intramuscularly injected with DHSM and KM and FRM, with these S105 fractions and AAC(6′)-1 resulted in the loss of the respective antimicrobial activities.When this method was applied to 4 bovine kidney samples in which AGs had been detected by the ordinary method, the AGs were identified as DHSM in two samples and KM in two other samples.This method is useful for identification of DHSM, KM and FRM in AGs-contaminated samples.