欢迎登录材料期刊网

材料期刊网

高级检索

建立了大鼠纹状体细胞膜色谱前沿分析法,研究5-羟色胺(5-HT)受体5-HT1D与藁本内酯的亲和作用。通过大鼠纹状体组织制备得到色谱固定相,利用酶联免疫吸附剂测定法( ELISA)分别测定硅胶吸附前后细胞膜悬液中5-HT的量,求得细胞膜固定相上5-HT受体含量为每克硅胶(40.5±2.3)pg。利用细胞膜色谱与液相色谱的离线联用,特异性地识别混合对照品中的舒马普坦和藁本内酯;以不同浓度(24.2~242 nmol/L)的5-HT1D受体激动剂舒马普坦为模型药物,连续通过细胞膜色谱柱,记录舒马普坦的突破曲线,测得舒马普坦与受体作用的平衡解离常数(KD)为389 nmol/L;并将舒马普坦通过不同浓度(37.0~370 nmol/L)的藁本内酯饱和后的细胞膜色谱柱,记录色谱柱饱和前后舒马普坦突破曲线的变化,测得藁本内酯与受体作用的 KD 值为4.21μmol/L。该方法快速、有效,适用于求解存在竞争结合时药物与受体作用的平衡解离常数。

A rat striatum cell membrane chromatography( CMC)frontal analysis method was developed for the determination of the equilibrium dissociation constants ( KD) for 5-hydroxytryptamine(5-HT)receptor 5-HT1D-ligustilide interactions. Rat striatum was used for preparation of the cell membrane stationary phase( CMSP). An enzyme-linked immunosorbent assay( ELISA)was applied to determine the 5-HT level of CMSP before and after the adsorp-tion of cell membrane,and the value was(40. 5±2. 3)pg per gram of silica. The CMC-offline-HPLC system was applied to specifically recognize the mixed standard solution of sumatriptan and ligustilide. Sumatriptan,a 5-HT1D agonist of 24. 2 to 242 nmol/L,was pumped through a CMC column continuously,and the breakthrough curves were recorded. For further competitive studies,the mobile phase that contained ligustilide(37. 0-370 nmol/L)was pumped through the column to saturate the binding sites. Afterwards,sumatriptan was propelled towards the column. The breakthrough curves were recorded and compared with those obtained from the column without saturation. KD values obtained using frontal analysis were 389 nmol/L and 4. 21μmol/L for sumatriptan and ligustilide,respectively. The competitive binding study indicated that the CMC method could be a quick and efficient way for determining the KDvalues in drug-receptor interactions.

参考文献

[1] Leonardi M;Steiner T J;Scher A T.[J].J Headache Pain,20056(6):429.
[2] 杨胜;张定堃;苏柘僮;徐佳丽;杨金辉.川芎-白芷药对不同配比不同剂型对偏头痛动物模型的影响[J].中国实验方剂学杂志,2011(14):225-228.
[3] 郑琴;魏韶锋;伍振峰;万晶;岳鹏飞;胡鹏翼;杨明.大川芎方对偏头痛大鼠模型血浆中CGRP及ET的影响[J].中药药理与临床,2011(4):3-5.
[4] 汪程远;杜俊蓉;钱忠明.藁本内酯的研究进展[J].中国药学杂志,2006(12):889-891.
[5] 左爱华;王莉;肖红斌.藁本内酯药理学和药代动力学研究进展[J].中国中药杂志,2012(22):3350-3353.
[6] 王雄飞;张玉杰;叶静;罗志强;詹雪艳;袁瑞娟.一种凝血酶亲和色谱介质的制备方法[J].色谱,2013(8):813-816.
[7] 胡青红;王超然;李静;王彦;谷雪;阎超.硼酸亲和整体柱在富集中药活性物质中的应用[J].色谱,2013(7):714-717.
[8] He L C;Yang G D;Geng X D.[J].Chin Sci Bull,199944(9):826.
[9] He L C;Wang S C;Geng X D.[J].Chromatographia,200154(1/2):71.
[10] Du, H.;Wang, S.;Ren, J.;Lv, N.;He, L..Revealing multi-binding sites for taspine to VEGFR-2 by cell membrane chromatography zonal elution[J].Journal of chromatography, B. Analytical technologies in the biomedical and life sciences,2012:67-72.
[11] Du, H.;Ren, J.;Wang, S.;He, L..Cell membrane chromatography competitive binding analysis for characterization of α_(1A) adrenoreceptor binding interactions[J].Analytical and bioanalytical chemistry,201110(10):3625-3633.
[12] Hou X F;Wang S C;Zhang T.[J].J Pharm Biomed Anal,2014101:141.
[13] Hou, XF;Zhou, MZ;Jiang, Q;Wang, SC;He, LC.A vascular smooth muscle/cell membrane chromatography-offline-gas chromatography/mass spectrometry method for recognition, separation and identification of active components from traditional Chinese medicines[J].Journal of chromatography, A: Including electrophoresis and other separation methods,200942(42):7081-7087.
[14] Moaddel R;Jozwiak K;Whittington K.[J].Anal Chem,200577(3):895.
[15] Sanghvi, M.;Moaddel, R.;Wainer, I.W..The development and characterization of protein-based stationary phases for studying drug-protein and protein-protein interactions[J].Journal of chromatography, A: Including electrophoresis and other separation methods,201149(49):8791-8798.
[16] Moaddel R;Wainer I W.[J].Nat Protoc,20094(2):197.
上一张 下一张
上一张 下一张
计量
  • 下载量()
  • 访问量()
文章评分
  • 您的评分:
  • 1
    0%
  • 2
    0%
  • 3
    0%
  • 4
    0%
  • 5
    0%