Sensitive LC determination of ciprofloxacin in pharmaceutical preparations and biological fluids with fluorescence detection

Abstract

A simple and highly sensitive isocratic reversed-phase high-performance liquid chromatographic method (RP-HPLC) has been developed for the determination of ciprofloxacin. Separation of ciprofloxacin and anthranilic acid (internal standard) was achieved on a Kromasil 100, C 18, 5 μm (250×4.6 mm i.d.) reversed-phase column, using fluorescence detection with λ exc=300 nm and λ emi=458 nm. The mobile phase consisted of acetonitrile–methanol–acetate buffer (pH 3.60; 0.05 M) (10:30:60 v/v/v) containing 1% v/v acetic acid. The analysis was performed in less than 9 min, with a flow rate of 0.8 ml min −1. A rectilinear relationship was observed for concentrations between 0.005 and 1.0 μg ml −1 of ciprofloxacin in aqueous standard solutions and serum and the detection limit was 20 pg injected on-column. The intra- and inter-day relative standard deviation ( n=8) ranged from 1.6 to 2.6% and from 1.9 to 4.8%, respectively, calculated at three concentration levels of standard solutions. Direct measurements of ciprofloxacin in pharmaceutical preparations and in serum, after precipitation of proteins, were performed with high precision and accuracy. The application of the method to urine samples involved a solid-phase extraction treatment of the samples using C 18 cartridges. The linear working range in urine extended from 0.05 to 2.0 μg ml −1 and the detection limit was 0.2 ng injected on-column.