Abstract: One of the two 5 mL reservoir syringes of a stopped-flow module of a fluorescence spectrometer was filled with a solution containing 0.15 mM borate buffer of pH 9.2, 2 mM o-phthaldialdehyde (OPA) and 3 mM N-acetylcysteine and the other with 0.15 mM borate buffer and an aminoglycoside antibiotic, amikacin (I), kanamicin (II) or tobramycin (III) at a final concentration of 0.05-30 µg/ml. A 0.15 mL portion from each syringe was transferred to a mixing chamber (1 cm path length) at 25°C and the fluorescence measured at 455 nm (excitation at 335 nm) throughout the reaction. The initial-rate procedure, based on the measurement of the slope of the kinetic curve fluorescence intensity vs. time, which is proportional to the analyte concentration was applied. Calibration graphs were linear for 0.05-30 µg/ml of I. Three equations were required to cover the concentration ranges 0.05-1 µg/ml, 1-10 µg/ml and 10^-30 µg/ml. The detection limit was 0.02 µg/ml. RSD (n = 11) were 1.8 and 0.9% for 0.5 and 12 µg/ml of I. I/interferent ratios of 1:50 and 1:4 were tolerated for penicillins and tetracyclines respectively. The mean recovery was 98.7%.

Abstract: Plasma (1 ml) was vortex-mixed with 0.1 mL of 50 mM Na2SO4 (pH 3.5) and 0.5 mL of 10% trichloroacetic acid for 15 s and the mixture was centrifuged at 5°C and 2000 g for 10 min. A 100 µL portion of the supernatant solution was subjected to HPLC on a column (12.5 cm x 4 mm) of LiChrospher RP 18, operated at 45°C, with a mobile phase (1.5 mL min-1) of (50 mM Na2SO4 and 5 mM sodium octylsulfate at pH 3.5) - methanol (7:3). The eluate was merged with a stream (0.3 mL min-1) of phthalaldehyde - 2-mercaptoethanol reagent (details given) before fluorimetric detection at 415 nm (excitation at 340 nm). The calibration graph was rectilinear from 0.1 to 2 µg mL-1 of amikacin (I); the detection limit was 25 ng mL-1. Recoveries ranged from 99.5 to 105%. The intra- and inter-day coefficient of variation for 0.2 µg mL-1 of I were 4.7 and 5.6%, respectivley. A pharmacokinetic study is reported.

Abstract: Biological fluids were deproteinized and I was adjusted before centrifugal filtration (Amicon Centrifree filter; 20 min). The filtrate was analyzed for amino-acids by HPLC on a column (15 cm x 3 mm) of cation-exchange resin (7 µm; Li+ form) at 45°C with Li citrate buffers (pH 2.75 and 7.5) as gradient eluents (0.3 mL min-1), post-column derivatization with phthalaldehyde and fluorescence detection at 425 nm (excitation at 334 nm). Iohexol and trometamol were eluted near or on the tyrosine peak, phenylpropanolamine was co-eluted with tyramine in the following chromatogram and amikacin gave a post-arginine peak which carried over on to the subsequent 2 to 3 chromatograms.