Zhang G Q, Kai M, Ohkura Y
Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
Chem Pharm Bull (Tokyo). 1991 Sep;39(9):2369-72. doi: 10.1248/cpb.39.2369.
A high-performance liquid chromatographic method involving postcolumn fluorescence derivatization is described for the quantification of five fragment peptides (methionine-enkephalin-Thr-Ser-Glu-Lys, methionine-enkephalin-Lys, methionine-enkephalin-Arg, leucine-enkephalin-Lys and leucine-enkephalin-Arg) released by tryptic digestion from the opioid peptide precursors (proopiomeranocortin and proenkephalins A and B) in rat brain tissues. The tissue proteins containing the precursors are hydrolyzed with trypsin to the fragment peptides. The peptides are separated on an Asahipak ODP-50 column and on-line detected fluorometrically by using hydroxylamine, cobalt(II) and borate buffer reagents. The detection limits (S/N = 3) for the peptides are 0.7-2.8 pmol per 100 microliters injected. The distribution of the precursors in the brain tissues was also discussed on the basis of the determined values of the fragment peptides.
本文描述了一种采用柱后荧光衍生化的高效液相色谱法，用于定量分析大鼠脑组织中阿片肽前体（阿片促黑皮质素原以及脑啡肽原A和B）经胰蛋白酶消化后释放的五种片段肽（甲硫氨酸脑啡肽-苏氨酸-丝氨酸-谷氨酸-赖氨酸、甲硫氨酸脑啡肽-赖氨酸、甲硫氨酸脑啡肽-精氨酸、亮氨酸脑啡肽-赖氨酸和亮氨酸脑啡肽-精氨酸）。含有前体的组织蛋白用胰蛋白酶水解为片段肽。这些肽在Asahipak ODP-50柱上分离，并使用羟胺、钴（II）和硼酸盐缓冲试剂进行在线荧光检测。这些肽的检测限（信噪比=3）为每注射100微升0.7 - 2.8皮摩尔。还根据片段肽的测定值讨论了前体在脑组织中的分布情况。
