Determination of antiretroviral agents in human serum by capillary electrophoresis

In this work, a simple and rapid electrokinetic chromatography method for the simultaneous separation of different protease inhibitors (indinavir, ritonavir, saquinavir, nelfinavir), nucleoside reverse transcriptase inhibitors (stavudine, zidovudine, didanosine) and non-nucleoside reverse transcriptase inhibitors (nevirapine, efavirenz) was developed. The analyses were performed in a 75 μm i.d. uncoated fused-silica capillary with 48.5 cm length (effective length of 40 cm) using a running buffer consisting of 20 mmol L−1 sodium dodecyl sulfate, 10 mmol L−1 sodium tetraborate, 30% acetonitrile and 5% ethanol. Samples were injected hydrodynamically by applying 50 mbar pressure during 6 s. All analytes were separated within 10 min with a voltage of 20 kV. The proposed method was validated for zidovudine, didanosine and efavirenz in human serum. Serum samples were prepared using a solid-phase extraction procedure (Waters® Oasis HLB cartridges). For quantitative purposes, stavudine was chosen as the internal standard (IS). Method validation parameters were determined revealing good migration time repeatability (<0.7% RSD) and peak area repeatability (<1.2% RSD). Intra- and inter-day precisions were less than 1.7% and 4.4% RSD, respectively. Matrix matching analytical curves for each drug were linear in the 1.0-20.0 μg mL−1 interval (r > 0.998). Limits of detection (LOD) were in range of 0.3-0.5 μg mL−1. The extraction recoveries were higher than 90% with exception of efavirenz, which was 77.4%. Based on the performance characteristics, the proposed method was found suitable for the determination of zidovudine, didanosine and efavirenz in serum samples.