Determination of intracellular tenofovir-diphosphate and emtricitabine-triphosphate concentrations in dried blood spots for pre-exposure prophylaxis adherence

Objectives: We measured the intracellular concentrations of tenofovir-diphosphate (TFV-DP) and emtricitabinetriphosphate (FTC-TP) in dried blood spots (DBS) for pre-exposure prophylaxis (PrEP) adherence using sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS). Methods: A total of 191 DBS were obtained from 85 participants who were receiving tenofovir disoproxil fumarate (TDF; 300 mg) and emtricitabine (FTC; 200 mg) as PrEP at the Sexual Health Clinic, National Center for Global Health and Medicine, Tokyo, Japan. DBS punch (3 mm) added to 25 mu L of 50% methanol and 400 mu L of internal standard solution was used for solid phase extraction. Chromatographic separation was achieved on an Atlantis Premier BEH C18 AX Column (50 mm x 2.1 mm i.d.; particle size 1.7 mu m) using gradient elution (flow rate: 0.6 mL/min); injection volume: 7 mu L and run time: 5.5 min. Calibration curves for the two drugs were linear in the range 0.05-12.5 ng/punch. Results: We determined the intracellular TFV-DP and FTC-TP concentrations in 191 DBS obtained from 85 patients administered with TDF and FTC as PrEP. The analytical performance data (calibration curve and QC samples) for all the analytical runs met the acceptance criteria. Intracellular concentrations of TFV-DP and FTCTP in the DBS remained stable for at least 24 h after oral administration. Conclusions: A multiplex LC-MS/MS method was successfully developed for DBS, which can be useful for monitoring the levels of TFV-DP and FTC-TP in individuals receiving PrEP.