薄层层析和反相高效液相色谱检测木瓜蛋白酶催化合成的Aspartame及其中间产物  

Determination of Papain Catalyzed Synthesis of Aspartame and Its In-termediate by TLC and Reverse- Phase HPLC

作　　者：姚文兵[1] 吴梧桐[1] 许激扬[1] 金生浩[1] 严悦日[1] 李康乐[1] 

机构地区：[1]中国药科大学生化教研室

出　　处：《药物生物技术》1994年第2期9-12,共4页Pharmaceutical Biotechnology

摘　　要：本文报道以ＴＬＣ和ＨＰＬＣ法检测以木瓜蛋白酶在水－乙酸乙酯两相反应体系中合成的Ａｓｐａｒｔａｍｅ及其中间产物（Ⅰ）（Ⅱ）。ＴＬＣ条件为：展开剂（正丁醇：冰醋酸：水＝３：１：１），展开时间（４～５ｈ），检测（荧光检测２５３．７ｎｍ，１％茚三酮丙酮溶液显色）。ＨＰＬＣ条件为：流动相（乙腈：水＝４５：５５），流速１ｍｌ／ｍｉｎ，检测（紫外检测２６０ｎｍ），进佯量２０ｕｌ，灵敏度６，纸速１～４ｍｍ／ｍｉｎ，分析周期１０ｍｉｎ。该方法简便、准确、灵敏度高。用该方法也证实以木瓜蛋白酶催化合成的Ａｓｐａｒｔａｍｅ及其中间产物（Ⅱ）与以嗜热蛋白酶（Ｔｈｅｒｍｏｌｙｓｉｎ）催化合成的Ａｓｐａｒｔａｍｅ及其中间产物（Ｃｂｚ－Ａｓｐ－ＰｈｅＯＭｅ）完全一致。AbstractThe aspartame and its intermediates(Ⅰ)and(Ⅱ)which were synthesized by pal)ain catalyzationwere determined by TLC and reverse-phase HPLC.Samples were dissolved in ethyl acetate and ana-lyzed by TLC on silica gel GF254 in n-BuOH:AcOH:H2O(3:1:1),for 4-5 hour,visualized byUV at 253.7nm and ninhydrin(hatched)Samples were also analyzed by reversed-phase HPLC on aupondpak C18 reverse-phase column with a mobile phase,acetOnitrile:water(45:55),flow rate1ml/min and detected by UV absorbance at 260nm. The result showed that the aspartame and its inter-mediate(Ⅱ) which were catalyzed by papain were similar to the aspartame and its in termediate (Cbz-Asp-pheOMe)which were catalyzed by thermolysin.

关 键 词：木瓜蛋白酶 APM 藻层层析法 甜味剂 HPLC 

分 类 号：TQ464.7[化学工程—制药化工] TS264.9[轻工技术与工程—发酵工程]