Printz H, Emmerich B, Loth H, Göke B
Department of Internal Medicine, Philipps University of Marburg, FRG.
Z Gastroenterol. 1990 Aug;28(8):383-8.
This study analyzed the secretory pattern of pancreatic proteins released from the rabbit pancreas after acute stimulation of secretion by the cholecystokinin analog cerulein. To facilitate this, a new analytical approach utilizing high performance liquid chromatography (HPLC) was considered. Secretin (0.1 CU/kg x h) was intravenously infused in anaesthetized rabbits in combination with cerulein (0.05, 0.2 or 0.05 followed by 0.2 ug/kg x h) over 3 hours. Pancreatic juice was collected from the main pancreatic duct. The release of protein, amylase, trypsin and chymotrypsin was measured by conventional photometric methods, and the protein profiles were analyzed by reversed phase HPLC. Separation of pancreatic juice proteins by HPLC (Nucleosil 300-7 RP column; injection of 50 ul aliquots of samples normalized to 10 mg/ml protein concentration) resulted in a resolution of up to 16 peaks. Peaks representing amylase, prolipase, prophospholipase A2, procarboxypeptidases, chymotrypsinogen, trypsinogen, and glycoproteins were identified with some certainty by SDS-gel electrophoresis. Secretin infusion produced a small and short lasting rise in total protein secretion but lead to a persistent increase of fluid flow. The release of enzymes followed a mainly parallel pattern according to the photometric measurements. The resolution of the whole profile of pancreatic juice proteins by HPLC demonstrated only minor variations without a consistent or increasing tendency towards a preferential release of individual enzymes. Since even microheterogenities in the samples became apparent after HPLC, this approach would be sensitive enough to mirror effects like nonparallel release of enzymes.
本研究分析了在胆囊收缩素类似物雨蛙肽急性刺激胰腺分泌后，兔胰腺释放的胰腺蛋白的分泌模式。为便于进行此项研究，考虑采用一种利用高效液相色谱（HPLC）的新分析方法。在麻醉的兔中静脉输注促胰液素（0.1 CU/kg×h），并在3小时内联合输注雨蛙肽（0.05、0.2或0.05随后为0.2μg/kg×h）。从主胰管收集胰液。通过传统的光度法测量蛋白质、淀粉酶、胰蛋白酶和糜蛋白酶的释放，并通过反相HPLC分析蛋白质谱。通过HPLC（Nucleosil 300 - 7 RP柱；注入50μl等份样品，将蛋白质浓度标准化为10mg/ml）分离胰液蛋白，可分辨出多达16个峰。通过SDS - 凝胶电泳可较为确定地鉴定出代表淀粉酶、前脂肪酶、前磷脂酶A2、前羧肽酶、糜蛋白酶原、胰蛋白酶原和糖蛋白的峰。输注促胰液素使总蛋白分泌出现小幅度且持续时间短的升高，但导致液体流量持续增加。根据光度测量，酶的释放主要呈现平行模式。通过HPLC对胰液蛋白的整个谱进行分辨，仅显示出微小变化，没有一致或增加的趋势表明个别酶优先释放。由于即使样品中的微小异质性在HPLC后也变得明显，这种方法足够灵敏，能够反映出酶的非平行释放等效应。
