Abstract
A direct fluorometric high-performance liquid chromatography (HPLC) method was developed and validated for the analysis of ibuprofen enantiomers in mouse plasma (100μl) and tissues (brain, liver, kidneys) using liquid-liquid extraction and 4-tertbutylphenoxyacetic acid as an internal standard. Separation of enantiomers was accomplished in a Chiracel OJ-H chiral column based on cellulose tris(4-methylbenzoate) coated on 5μm silica-gel, 250 x 4.6mm at 22°C with a mobile phase composed of n-hexane, 2-propanol, and trifluoroacetic acid that were delivered in gradient elution at a flow rate of 1ml min-1 . A fluorometric detector was set at: λexcit . = 220nm and λemis. = 290nm. Method validation included the evaluation of the selectivity, linearity, lower limit of quantification (LLOQ), within-run and between-run precision and accuracy. The LLOQ for the two enantiomers was 0.125 μg ml-1 in plasma, 0.09μg g-1 in brain, and 0.25μg g-1 in for liver and kidney homogenates. The calibration curves showed good linearity in the ranges of each enantiomers: from 0.125 to 35μg ml-1 for plasma, 0.09-1.44μg g-1 for brain, and 0.25-20μg g-1 for liver and kidney homogenates. The method was successfully applied to a pharmacokinetic study of ibuprofen enantiomers in mice treated i.v. with 10mg kg-1 of racemate.
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