Simultaneous Determination of Zalcitabine and Stavudine in Maternal Plasma, Amniotic Fluid, Placental, and Fetal Tissues Using Reversed Phase on Silica Liquid Chromatography/Tandem Mass Spectrometry

Abstract

In order to study the placental transfer of nucleoside reverse transcriptase inhibitors, a quick and simple reversed phase high performance liquid chromatography with tandem mass spectrometry method has been developed and validated using an underivatized silica column for the separation and analysis of DDC and D4T from rat plasma, amniotic fluid, placental, and fetal homogenate. Extraction of DDC, D4T, and their internal standard lamivudine (3TC) from the matrices was processed by protein precipitation using ice cold acetonitrile. Chromatographic separation was achieved on a Waters Spherisorb S3W silica column (4.6 mm × 100 mm) equipped with a Phenomenex guard column. The mobile phase consisted of 20% methanol in 22 mM formic acid. The flow rate was 0.4 mL/min, and MRM was used for detection. The calibration curves for each day of validation showed good linear response over the range from 2 ng/mL to 2000 ng/mL. The absolute recoveries for all the drugs are all higher than 70%, and the matrix effects are all lower than 20%. All the intra- and inter- day assay precision and accuracy were better than 10% for all the matrices.

Keywords:

• Amniotic
• Fetal tissues
• Maternal plasma
• Placental
• Reversed phase
• Silica LC
• Stavudine
• Tandem mass spectrometry
• Zalcitabine