Determination of amikacin in serum by high-performance liquid chromatography with ultraviolet detection

A procedure for the determination of amikacin in serum is described. The aminoglycoside is extracted from serum by using a disposable cation-exchange column. The eluate of this column is derivatized with 1-fluoro-2,4-dinitrobenzene and subsequently analysed by reversed-phase high-performance liquid chromatography with ultraviolet detection at 365 nm. The absolute recovery of amikacin by this procedure is 72%. Kanamycin is used as the internal standard. The sensitivity is 1 mg/l for amikacin with samples of 200 microliters. Precision, expressed as the coefficient of variation, is about 3% in the therapeutic concentration range. The 2,4-dinitrophenyl derivative of amikacin is synthesized on a preparative scale by a new method and its structure is demonstrated to be the fully derivatized amikacin. The analysis of serum samples obtained in an in vivo experiment correlates well with the results from a microbiological assay.