Combination of hollow-fiber liquid-phase microextraction and capillary electrophoresis for pioglitazone and its main metabolites determination in rat liver microsomal fraction.

Pioglitazone (PGZ), a thiazolidinedione antidiabetic agent, is reported as a potent and selective activator of peroxisome proliferator-activated receptor γ (PPAR γ). This drug has been widely prescribed for the treatment of Type 2 diabetes mellitus. In this regard, this manuscript presents, for the first time, an alternative electrophoretic method for PGZ and its main metabolites determination in rat liver microsomal fraction. The electrophoretic analyses were performed using an uncoated fused-silica capillary of 50 μm id, 48 cm in total length and 40 cm in effective length, and 50 mmol/L sodium phosphate buffer solution (pH 2.5). All experiments were carried out under the normal mode. The capillary temperature was set at 35°C and a constant voltage of +30 kV was applied during the analyses. Samples were introduced into the capillary by hydrodynamic injection (50 mbar, 15 s) and detection was performed at 190 nm. The sample preparation procedure, based on hollow-fiber liquid-phase microextraction, was optimized using multifactorial experiments. Next, the following optimal condition was established: sample agitation at 1500 rpm, extraction for 15 min, 0.01 mol/L hydrochloric acid as acceptor phase, 1-octanol as organic phase, and donor phase pH adjustment to 6.0. The method demonstrated LOQs of 200 ng/mL. Additionally, it was linear over the concentration range of 200-25,000 ng/mL for PGZ and 200-2000 ng/mL for the metabolites. Finally, the validated method was employed to study the in vitro metabolism of PGZ using rat liver microsomal fraction.