去铁酮与人有机阳离子转运体及有机阴离子转运体1体外相互作用研究  

In vitro interaction of deferiprone with cellular membrane transporters of hOCTs and hOAT1

作　　者：孔思思 涂美娟[1] 杨希[1] 赵垒[1] 周慧[1] 曾苏[1] 蒋惠娣[1] 

机构地区：[1]浙江大学药学院药物分析与药物代谢研究室浙江省抗肿瘤药物临床前研究重点实验室

出　　处：《浙江大学学报（医学版）》2014年第2期129-134,共6页Journal of Zhejiang University(Medical Sciences)

基　　金：国家科技重大专项(2012ZX09506001-004)

摘　　要：目的：建立细胞裂解液中去铁酮测定的液相色谱-串联质谱（ LC-MS/MS）方法，体外考察去铁酮与人有机阳离子转运体（ hOCTs）及有机阴离子转运体1（hOAT1）的相互作用。方法：以Agilent Eclipse Plus C18柱（2．1 mm ×50 mm，3．5μm）为分析柱；0．1％甲酸-水（ v/v）和0．1％甲酸-乙腈（ v/v）为流动相，梯度洗脱；应用电喷雾离子源（ ESI源）、多反应监测（ MRM）模式进行检测；应用稳定表达hOCTs及hOAT1的细胞模型（ MDCK-hOCTs和MDCK-hOAT1）考察去铁酮对经典底物在细胞内积聚的影响；比较去铁酮在转空载体的mock细胞及MDCK-hOCTs中积聚的差异，以及经典抑制剂对其积聚的影响。结果：去铁酮在5～300 nmol/L的浓度范围内线性关系良好；准确度大于94％，日内相对标准偏差小于2％。300μmol/L去铁酮使hOCTs经典底物MPP＋在MDCK-hOCTs的积聚降低至阴性对照的70．4％～87．1％；去铁酮在MDCK-hOCTs细胞和mock细胞中的积聚无明显差别，且经典抑制剂对去铁酮的积聚无明显抑制作用；100μmol/L去铁酮对hOAT1经典底物6-CF在MDCK-hOAT1细胞中的积聚无显著影响。结论：本实验建立的方法适用于细胞裂解液中去铁酮的定量分析；去铁酮对hOCT1和hOCT3有一定的抑制作用，对hOCT2和hOAT1则无明显抑制作用；hOCTs在去铁酮跨膜转运中不起主导作用。Objective： To develop a LC-MS/MS method for determination of＆nbsp;deferiprone in cell lysate and to study the potential interaction between deferiprone and hOCTs or hOAT1 transporters in vitro.Methods： The determination was performed on an Agilent Eclipse Plus C18 column（3.5 μm, 2.1 mm ×50 mm）.The gradient mobile phase was composed of solvent A：0.1% formic acid in water, and B：0.1% formic acid in acetonitrile .The mass spectrometer with an electrospray interface was operated in positive ion mode with multiple reaction monitoring （ MRM） scan mode monitored the ion pair of deferiprone at m/z 140→96 , or phenacetin at m/z 180→110 .The effects of deferiprone on the accumulation of typical substrates of hOCTs and hOAT 1 were evaluated by MDCK-hOCTs and MDCK-hOAT1 cells respectively .The accumulation of deferiprone was also investigated in MDCK-hOCTs cells and mock cells with or without typical inhibitors .Results： The standard curve was linear over the range of 5-300 nmol/L.The assay recovery of deferiprone was above 94%, and the intra-day precision （RSD） was less than 2.0%.The accumulation of MPP ＋in MDCK-hOCTs cells with 300 μmol/L deferiprone were 73 .5%, 87 .1% and 70 .4%, respectively .The uptake of deferiprone in MDCK-hOCTs and mock cells did not show significant difference . Deferiprone of 100 μmol/L did not significantly affect the accumulation of 6-CF in MDCK-hOAT1 cell. Conclusion： The method is sensitivity and suitable for the determination of deferiprone in cell lysate .Deferiprone can significantly inhibit hOCT 1 and hOCT3, but has no effects on hOCT2 and hOAT1.hOCTs may not play a major role in the transport of deferiprone .

关 键 词：酮类 铁螯合剂 质谱分析法 色谱法 液相 有机阳离子转运子 有机阴离子转运子1 

分 类 号：R96[医药卫生—药理学]