Quantitative subcellular study of doxorubicin in MCF-7/Adr cells using liquid chromatography-tandem mass spectrometry
Wenzhuan Ma1, Jinling Wang2, Qiang Guo1
1Modern Research Center for Traditional Chinese Medicine, Beijing University of Chinese Medicine, Beijing 100029, People's Republic of China; School of Chinese Materia Medica, Beijing University of Chinese Medicine, Beijing 100102, People's Republic of China.
Abstract:
A rapid, sensitive and selective high-performance liquid chromatography-tandem mass spectrometric (LC-MS/MS) method has been developed and validated for the determination of doxorubicin in intracellular compartments using glibenclamide as internal standard (IS). MCF-7/Adr cancer cells (1×10(6)) were incubated with doxorubicin (8μg/mL) for 0.5, 1, 2 and 4h and then subjected to sequential extraction of cytosolic, membrane/organelle, nuclear and cytoskeleton soluble protein. Samples were extracted using protein precipitation with methanol. Chromatographic separation was carried out on a C18 column with acetonitrile and 0.1% formic acid water as mobile phase and with gradient elution at a flow rate of 0.2mL/min. The method was linear over the range of 1-300ng/mL with a lower limit of quantification (LLOQ) of 1ng/mL. The distribution of doxorubicin in subcellular components of MCF-7/Adr cancer cells was mainly in nucleic protein fraction.
