Quantitation of tenofovir and emtricitabine in dried blood spots (DBS) with LC-MS/MS.
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.
Skaggs School of Pharmacy and Pharmaceutical Sciences Department of Pharmaceutical Sciences, University of Colorado Denver, Anschutz Medical Campus, United States.
Jia-Hua
JH
Zheng
Louis A
LA
Guida
Caitlin
C
Rower
Jose
J
Castillo-Mancilla
Amie
A
Meditz
Brandon
B
Klein
Becky Jo
BJ
Kerr
Jacob
J
Langness
Lane
L
Bushman
Jennifer
J
Kiser
Peter L
PL
Anderson
D000225:Adenine / Q000031:analogs & derivatives* / Q000032:analysis / Q000097:blood; D002138:Calibration; D002853:Chromatography, Liquid; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000032:analysis / Q000097:blood; D059788:Dried Blood Spot Testing; D004355:Drug Stability; D000068679:Emtricitabine; D006400:Hematocrit; D006801:Humans; D063065:Organophosphonates / Q000032:analysis* / Q000097:blood*; D011786:Quality Control; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry; D013696:Temperature; D000068698:Tenofovir
D063065:Organophosphonates; D003841:Deoxycytidine; D000068698:Tenofovir; D000068679:Emtricitabine; D000225:Adenine
Analytical method; Antiretroviral therapy; Dried blood spot; LC–MS/MS; Nucleoside analog