Quantitation of tenofovir and emtricitabine in dried blood spots (DBS) with LC-MS/MS

J Pharm Biomed Anal. 2014 Jan;88:144-51. doi: 10.1016/j.jpba.2013.08.033. Epub 2013 Aug 31.

Abstract

A reversed-phase high performance liquid chromatographic (LC), tandem mass spectrometry (MS/MS) assay for the determination of tenofovir (TFV) and emtricitabine (FTC) in dried blood spots (DBS) from human whole blood was developed and validated. Whole blood samples were spotted, dried, and a 3mm punch was extracted with methanol for analysis by LC-MS/MS utilizing stable isotope labeled internal standards. The assay was validated over the range of 2.5-1000ng/mL for TFV and 2.5-5000ng/mL for FTC. The method was accurate (within ±15% of control) and precise (coefficient of variation ≤15%) for hematocrit concentrations ranging from 25% to 76%; using edge punches vs. center punches; and spot volumes of 10-50μL. Analytes were stable for five freeze/thaw cycles and up to 6 days at room temperature, whereas long-term storage required -20°C or -80°C. Comparison of TFV and FTC in DBS vs. plasma yielded r(2)≥0.96, indicating that DBS can be used as a plasma alternative for pharmacokinetic analyses in vivo.

Keywords: Analytical method; Antiretroviral therapy; Dried blood spot; LC–MS/MS; Nucleoside analog.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenine / analogs & derivatives*
  • Adenine / analysis
  • Adenine / blood
  • Calibration
  • Chromatography, Liquid
  • Deoxycytidine / analogs & derivatives*
  • Deoxycytidine / analysis
  • Deoxycytidine / blood
  • Dried Blood Spot Testing
  • Drug Stability
  • Emtricitabine
  • Hematocrit
  • Humans
  • Organophosphonates / analysis*
  • Organophosphonates / blood*
  • Quality Control
  • Reproducibility of Results
  • Tandem Mass Spectrometry
  • Temperature
  • Tenofovir

Substances

  • Organophosphonates
  • Deoxycytidine
  • Tenofovir
  • Emtricitabine
  • Adenine