Intra-intestinal kinetics and absorption mechanism of digestion-resisitant maclomolecular food components

卵白蛋白(OVA)对蛋白水解性具有抵抗力，被认为是鸡蛋过敏的主要致敏蛋白。以卵白蛋白为模型大分子食物成分，研究了小鼠肠道卵清蛋白在体内的吸收及循环外周血中卵清蛋白的代谢动力学。小鼠口服卵清蛋白(OVA)10 mg后，采集外周血，从凝血中分离血清。EUSA法测定血清中OVA浓度为10~50 ng/ml，用抗OVA抗体偶联琼脂糖用免疫亲和沉淀法浓缩血清中的OVA。在SDS-PAGE分析中，从外周血中回收的OVA显示单一的主要蛋白条带，其分子质量与完整的OVA相同，而次要条带的相对分子质量略小。小分子的OVA片段是小鼠小肠中OVA抗原的主要成分，小分子OVA片段没有相应的条带。双向凝胶电泳法显示，从血液中回收的卵蛋白由4-6个等电点不同但相对分子质量相同的蛋白质点组成，且与完整的卵蛋白相比，碱性蛋白质点占主导地位，这表明在肠道吸收或循环血液迁移过程中，OVA的等电点向更碱性的蛋白质点移动。在体外与小鼠血清孵育的OVA中，未观察到这种等电点的变化。综上所述，口服OVA的一小部分可以从肠道蛋白水解性消化中逃脱，作为一个完整的形式被吸收，但略有修改，通过肠道上皮进入血流。因此，口服生物活性蛋白可能在外周组织和器官中发挥其生物学功能。

Egg albumin (ovalbumin:OVA) is resistant to proteolytic hydrolysis and known to be a major allergenic protein in egg allergy. By using egg albumin as a model macromoleculer food component, the absorption of OVA from mouse intestinal tract into body and the kinetics of OVA in circulating peripheral blood were investigated. After oral administration of OVA (10 mg) to mice, peripheral blood was collected and then serum was separated from coagulated blood. The OVA concentration in the serum was determined to be 10-50 ng/ml by EUSA, and the OVA in the serum was concentrated by immuno-affinity precipitation using anti-OVA antibodies coupled to Sepharose. In SDS-PAGE analysis, the OVA recovered from peripheral blood revealed a single major protein band with a molecular mass identical to intact OVA and a minor band with a slightly smaller molecular mass. No band corresponding to OVA fragments with smaller molecular masses, which had been demonstrated to be the major components of OVA antigens in the mouse small intestine. Furthermore, two-dimensional gel electrophoresis demonstrated that the OVA recovered from blood was composed of 4-6 protein spots with different pI but with the same molecular mass, and that the basic protein spots were dominant as compared with those of intacl OVA, suggesting that the pI of OVA was shifted to more basic ones during the intestinal absorption or the migration in circulating blood. Such a pI shift was not observed for OVA incubated in vitro with mouse serum. Taken together, it is suggested that a small portion of orally administered OVA could escape from intestinal proteolytic digestion and be absorbed as an intact form, but with a slight modification, across intestinal epithelium into the blood stream. Thus, it might be possible that orally administered bio-active proteins exert its biological function in peripheral tissues and organs.