Zhang G Q, Kai M, Nakano M, Ohkura Y
Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
Chem Pharm Bull (Tokyo). 1991 Jan;39(1):126-9. doi: 10.1248/cpb.39.126.
A high-performance liquid chromatographic method involving fluorescence derivatization followed by separation on a reversed-phase polymer (octadecylated polyvinylalcohol copolymer gel) column is described for the determination of opioid peptides in rat brain tissues. The peptides extracted from brain tissues were converted into fluorescent derivatives by reaction with hydroxylamine, cobalt(II) ion and borate. The derivatives were separated on an Asahipak ODP-50 column by gradient elution of acetonitrile in the mobile phase containing borate buffer (pH 9.5). The detection limits (S/N = 3) for the peptides were 0.33-1.21 pmol per 100 microliters injected. The method actually permit the determination of leucine enkephalin, methionine enkephalin, methionine enkephalin-Arg-Phe and methionine enkephalin-Arg-Gly-Leu in the tissues. The method is also applied to the characterization of the peptides in the tissues by means of enzymatic degradations with carboxypeptidase A and trypsin.
本文描述了一种高效液相色谱法，该方法包括荧光衍生化，然后在反相聚合物（十八烷基化聚乙烯醇共聚物凝胶）柱上进行分离，用于测定大鼠脑组织中的阿片肽。从脑组织中提取的肽通过与羟胺、钴（II）离子和硼酸盐反应转化为荧光衍生物。衍生物在Asahipak ODP-50柱上通过在含有硼酸盐缓冲液（pH 9.5）的流动相中用乙腈进行梯度洗脱来分离。肽的检测限（S/N = 3）为每注射100微升0.33 - 1.21皮摩尔。该方法实际上可以测定组织中的亮氨酸脑啡肽、甲硫氨酸脑啡肽、甲硫氨酸脑啡肽-Arg-Phe和甲硫氨酸脑啡肽-Arg-Gly-Leu。该方法还通过用羧肽酶A和胰蛋白酶进行酶促降解来用于组织中肽的表征。
