Structure Identification Of Mung Bean Peptides And Its Effect On The Immunoreactive Substances Of Macrophage In Mice

Bioactive peptide enzymatic hydrolysised from protein is proved to be an important material can be involved in a variety of cellular functions in the human body. The functional and mechanism research of bioactive peptide is an important theoretical base for functional food and medicine. In our study, we used mung bean peptides(MBPs), getting from the Alcalase 2.4L alkaline protease hydrolyse the mung bean protein, as the research object. Analysed the amino acid composition, molecular weight distribution, structure characteristics and stability,by means of automatic amino acid analyer, SDS-PAGE electrophoresis, FTIR, HPLC and MS. Then studied its influence to the glycogen, DNA and RNA, ATPase, LZM and SOD activity in macrophages. Finally discussed its effect on the immune activity of macrophages by measuring macrophage production of NO and iNOS activity and iNOSmRNA expression quantity alteration.The research content and results are as follows.1. The structure identification and stability analysis of MBPs. MBPs hydrolysis degree is32.25%, it contains 17 kinds of amino acid, total nonpolar amino acid 28.37%, polarity of neutral amino acids 15.03%, acidic amino acid 32.27% alkaline amino acid 24.33%, nonpolar and basic amino acid which is proved to be associated with immune related content is higher has a higher proportion. The determination results of SDS-PAGE and HPLC shows that its molecular weight is under 6.5 KDa. According to the results of FTIR, MBPs may contain a variety of unsaturated group and still contain protein secondary structure of ?-helix and irregular curl, but ?-folding and?-corner structure have been digestion and open. According to the results of MS, 216 kinds of peptides getting from MBPs mainly belongs to 24 kinds of protein. The ends of peptide nonpolar and basic amino acid content is higher.2. The influence of MBPs to active substances in macrophages. Determined the macrophage proliferation by MTT method, showing that MBPs can significantly(P<0.05) promote macrophage proliferation and no cytotoxicity, Can also significantly(P<0.05) ease the LPS inhibition to macrophage proliferation. Macrophage cultured in MBPs its glycogen and nucleic acid staining obviously deepened, To some extent reflects the macrophages activity enhancement after MBPs culturing. MBPs concentration for more than 200?g/m L can significantly(P<0.05)promoting the ATPase activity of macrophages, MBPs can significantly ease the inhibition ofATPase activity by LPS. MBPs concentrations greater than 100?g/mL can significantly(P<0.05)strengthen the LZM activity in macrophages. Significantly reduce(P<0.05) the LZM activity weakened caused by LPS. Only when the concentration of MBPs more than 200?g/mL, its effect to promote SOD activity and inhibition to LPS stimulation will be obvious.3. The influence of MBPs to NO, iNOS and iNOSmRNA in macrophages. Using MBPs to cultivate the macrophages, its NO secretion increased significantly(P<0.05) after 24 h. NO have enough time to play a role of immunity. MBPs can promote the NO secretion of macrophage.When the MBPs concentration is 10?g/mL, NO secretion of macrophage is not significant(P>0.05), When the MBPs concentration is 200?g/mL or 400?g/mL, their NO secretion of macrophage have no statistical difference(P>0.05). The NO promoting effect of MBPs was lower than those of LPS. MBPs can significantly(P<0.05) reduce the LPS stimulation of macrophages. MBPs and LPS can also significantly(P<0.05) increased the activity of iNOS, But MBPs can also inhibit the excessive expression of iNOS caused by LPS, the concentration of MBPs higher, the effect more obvious.The real-time fluorescent quantitative PCR analysis on iNOSmRNA showed that the MBPs cultured macrophage iNOSmRNA expression increased, but lower than that of LPS stimulation. MBPs can also reduce the excessive expression of macrophage iNOSm RNA caused by LPS.The molecular weight of MBPs mostly concentrated below 6.5 KDa, Still contain ?-helix and irregular curl, but ?-folding and ?-corner structure have been digestion and open. Promote the NO secretion, iNOS activity and iNOSmRNA expression of macrophage. Prompting macrophages play a role of immunity.