High-performance liquid chromatography with time-programmed fluorescence detection for the quantification of Levofloxacin in human plasma and cerebrospinal fluid in adults with tuberculous meningitis.

An accurate and reliable high-performance liquid chromatography with time-programmed fluorescence detection was developed and validated to measure levofloxacin in human plasma and cerebrospinal fluid (CSF). After solid phase extraction process using Evolute® ABN 96 fixed well plate; levofloxacin and internal standard-enoxacin were separated using a mobile phase consisting of phosphate buffer 10mM with 0.025% triethylamine pH 3.0 - acetonitrile (88:12, v/v) on a Purosphere RP-8e column (5μm, 125×4.0mm) at a flow rate of 1.2mL/min at 35°C. The excitation/emission wavelengths were set to 269/400nm and 294/500nm, for enoxacin and levofloxacin, respectively. The method was linear over the concentration range of 0.02 to 20.0μg/mL with a limit of detection of 0.01μg/mL. The relative standard deviation of intra-assay and inter-assay precision for levofloxacin at four quality controls concentrations (0.02, 0.06, 3.0 and 15.0μg/mL) were less than 7% and the accuracies ranged from 96.75% to 101.9% in plasma, and from 93.00% to 98.67% in CSF. The validated method was successfully applied to quantify levofloxacin in a considerable quantity of plasma (826) and CSF (477) samples collected from 232 tuberculous meningitis patients, and the preliminary intensive pharmacokinetics analysis from 14 tuberculous meningitis patients in Vietnam is described in this paper.