The Protective Effect Of Wheat Oligopeptides On Oxidative Stress In Intestinal Epithelial Cells In Vitro

The department of agriculture has put forward the activities and the action plans of agricultural products processing, in order to promote the pretreating of agricultural products and the development of the products processed. Though wheat production is high, but it still faced with the problems of a single product structure, short wheat product processing chain, less value-added, no high value-added products. So in a great extent, this problems restricts the ability of agriculture to contribute to increase the farmers’ income. Wheat processing is important to solve the problems of agriculture, rural areas and farmers in China. This study focuses on the subject of wheat oligopeptides, the bowel glues the film repair in vitro against oxidative stress injury and repairing mechanisms, mining more function of wheat oligopeptides, providing favorable foundation support for the effective development and utilization of wheat oligopeptides, and providing new ideas and ways for the development of high value-added food of wheat oligopeptides research. The test contents and results of this test are as follows:1. The main composition analysis of wheat oligopeptides. Experimental results show that the moisture content of wheat low poly peptide is 4.35%, protein content(wet basis) is 89.64%, fat content is 2.18%, ash content is 4.90%, glutamine content is 24.99%.Hydrolytic amino acid composition is balanced. With the content as high as 18.874 g / 100 g, wheat low poly peptide is rich in essential amino acid and a high quality source of amino acids. With the average molecular weight of 433.6 Da, component with molecular weight less than 1000 Da of wheat oligopeptides accounted for 93.45% in which the molecular weight of 140-500 Da accounted for 57.94% which accounted for the most part of below 1000 Da, and this part mainly is dipeptide and tripeptide, so we can conclude that dipeptide and tripeptide is the body of wheat low poly peptide composition. Therefore the main components in wheat oligopeptides this experiment used is protein which have excellent quality.2. The construction of the model of intestinal mucosa in vitro cell. The model of Caco-2 cells in vitro intestinal mucosa is established in this experiment. we verified the Caco-2 this experiment used is in line with the index of intestinal mucosa in vitro by determining cell growth curve, observing whether the cell is closely connected or not, and cell polarity optimization. The results showed that the Caco-2 cells grow in good condition during the process of cell culture for 21 days, and across the membrane resistance value of Caco-2 cells in culture to 21 days is as high as(522±31) ?.cm2, the rate on both sides of the alkaline phosphatase activity ratio is 3.76:1, the validation of phenol red apparent permeability coefficient which was found in 1, 3, 5 h incubation are less than 10-6 cm.s- 1, which means that Caco-2 cells in culture to 21 days form close connection, grow in good condition and form some certain polarity, so in this pilot training optimization of Caco-2 cells can be used for the study of small intestinal epithelial cells.3. The protection of wheat oligopeptides on Caco-2 cells oxidative stress damaged. The experimental results indicate that 1, 5, 10mg/m L of wheat oligopeptides promotes cell proliferation obvious mostly, and 100 μmol/L of H2O2 can cause significantly oxidative stress damage to Caco-2 cells. Considering of Caco-2 cell cytotoxicity mediated by H2O2 and oxidative stress wheat oligopeptides has significant protective effect, and can significantly reduce the cell death rate and significantly improve cell survival, significantly reduce the content of ROS, MDA significantly improve the content of SOD, significantly improve the tightness of cells, significantly increased the membrane-crossing resistance, significantly reduced the through rate of phenol red. Test results show that wheat oligopeptides can improve the monolayer epithelial barrier function injury of Caco-2 cell mediated by H2O2.4.The comparison of the protective effect of wheat oligopeptides and glutamine on oxidative stress injury of intestinal mucosa in vitro. The protective effect of wheat oligopeptides and glutamine on Caco-2 cells with oxidative stress damage had been analyzed. By the results of the experiment, wheat oligopeptides of 5 mg/m L improve the activity of Caco-2 cells with oxidative stress injury more significantly than glutamine of 1.25 mg/m L, besides, compared with glutamine of 5 mg/m L, there is no significant difference until 48h; Wheat oligopeptides of 5 mg/m L have the higher level of ability to improve the activity of Caco-2 cells with oxidative stress injury than glutamine of 1.25 mg/m L, and is slightly less than glutamine of 5 mg/m L and there was no significant difference in individual working time; Wheat oligopeptides of 5 mg/m L have the higher level of ability to improve barrier damage of Caco-2 cells with oxidative stress injury than the glutamine of 1.25 mg/m L, and in some time there was no significant difference compared with the ability of glutamine of 5 mg/m L.5. The protection mechanism of wheat oligopeptides optimization on Caco-2 cells oxidative stress damaged. Cell is closely connected with a variety of related closely connected protein, ZO-1, ZO-2 can form stable complexes and can be combined with occludin proteins, forming a stable network support in the process of signal transmission, control cells closely connectivity, so as to control cell barrier around; HO-1 particles widely exists in various tissues and cells, and is the rate-limiting enzyme of heme catabolism, it can be generated by the degradation of heme producing the same Moore of carbon monoxide, carbon, free iron and biliverdin which can quickly be reduced into bilirubin by biliverdin reductase. Inducing the expression of HO-1 is the important protection mechanism in the oxidative stress state. HO-1 in many disease can abundantly expressed in the process of development, have protective effects on the body, reducing cell death and lipid peroxidation and protein oxidation, HO-1 is a kind of extremely important stress proteins.The experimental results indicate that the wheat oligopeptides can promote the expression of ZO-1, ZO-2 m RNA, especially in the 5 mg/m L, and positively related to the present time concentration in a certain range; And wheat oligopeptides can regulate the expression of HO-1 through optimization to repaire the oxidative stress injuried Caco-2 cells.