摘要
Objective: To establish a HPLC method using fluorometric detection for quantitatively determinating intracellular accumulation of verapamil (VER). Methods: Chromatography column was packed with spherisorb ODS(250×4.6 mm,10 μm).The mobile phase consisted of the mixture of methanol:NaAC (0.01 mol/L): diethylamine (65:35:0.25). The detect wavelength was 280/310 nm (Ex/Em). Results: The standard curve showed a good correlation between concentration and peak area within the range of 5-50 ng/ml. RSD was 0.86%, and recovery radio of loading sample, 100%. The detection limit for cell sample was 0.2-148 ng/ml. Intracellular accumulation of VER was observed to decrease from a 13 fold to 5 fold in K562/ADM cells, and from a 3.5 fold to 4.3 fold in K562/VER cells and from a 2.1 fold to 6.5 fold in K562/ADM/VER cells, compared with the relevant control cells. Conclusion: HPLC method was proved to be sensitive and specific for using to quantitatively determine the intracellular accumulation of VER.
Objective: To establish a HPLC method using fluorometric detection for quantitatively determinating intracellular accumulation of verapamil (VER). Methods: Chromatography column was packed with spherisorb ODS(250×4.6 mm,10 μm).The mobile phase consisted of the mixture of methanol:NaAC (0.01 mol/L): diethylamine (65:35:0.25). The detect wavelength was 280/310 nm (Ex/Em). Results: The standard curve showed a good correlation between concentration and peak area within the range of 5-50 ng/ml. RSD was 0.86%, and recovery radio of loading sample, 100%. The detection limit for cell sample was 0.2-148 ng/ml. Intracellular accumulation of VER was observed to decrease from a 13 fold to 5 fold in K562/ADM cells, and from a 3.5 fold to 4.3 fold in K562/VER cells and from a 2.1 fold to 6.5 fold in K562/ADM/VER cells, compared with the relevant control cells. Conclusion: HPLC method was proved to be sensitive and specific for using to quantitatively determine the intracellular accumulation of VER.
