Enantioselective analysis of ibuprofen enantiomers in mice plasma and tissues by high‐performance liquid chromatography with fluorescence detection: Application to a pharmacokinetic study

A direct fluorometric high-performance liquid chromatography (HPLC) method was developed and validated for the analysis of ibuprofen enantiomers in mouse plasma (100 μl) 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 5 μm silica-gel, 250 x 4.6 mm 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 1 ml min−1. A fluorometric detector was set at: λexcit. = 220 nm and λemis. = 290 nm. 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, 0.09 μg g−1 in brain, and 0.25 μg g−1 in for liver and kidney homogenates. The calibration curves showed good linearity in the ranges of each enantiomers: from 0.125 to 35 μg ml−1 for plasma, 0.09–1.44 μg g−1 for brain, and 0.25–20 μg 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 10 mg kg−1 of racemate.