Study On The Bioactive Peptides Of Sargassum Fulvellun

Water-soluble substances isolated from the Sargassum fulvellun by semi-permeable membrane (MWCO: 10KD) dialysis, dialysis using dextran gel material (Sephadex G-75) were purified by TLC and purified by Tricine-SDS-PAGE confirmed the results. Dry clean 200 g Sargassum fulvellun and cut into 0.5 cm long, at room temperature (25℃), 10 r / min under the conditions of 5 times the amount of deionized water, water-soluble substances extracted 12 h. Extract obtained in the 10000 rpm, 4℃, 10 min supernatant under the conditions of centrifugation, the supernatant under the conditions of 60℃was evaporated to 50 ml, molecular weight of 10 KD by the interception of the semi-permeable membrane dialysis After dialysis the material was used after column Sephadex G-75 Sephadex and flow quite different ionized water, distilled water - methanol (3:2), 10 mmol.L-1 PBS, 10 mmol.L-1 PBS shows the best effect, and the purified material in methanol - ethyl acetate as the agent, stained with ninhydrin on TLC shows one spot in the Tricine- SDS-PAGE showed the only band. 200 g Sargassum by extraction, centrifugation, concentration, dialysis, chromatography, freeze-dried purified peptide of Sargassum is 20 g, the non-peptide, tentatively entitled SFPP (Sargassum fulvellun polypeptide).Biochemical characteristics of the SFPP found that the molecular weight of SFPP about 17 KDa, thermal denaturation temperature of 51.4℃, with the characteristic amino acid composition. Added in the separation gel with urea Tricine-SDS-PAGE method, with silver stain out of SFPP molecular weight of about 17 KDa; by differential scanning calorimetry determination the thermal denaturation temperature of SFPP is 51.4℃; application of amino acid analyzer amino acid composition of the SFPP. Which contains a high content of aspartic acid (Asp), glutamic acid (Gly) and lysine (Lys), and other algae peptides have significantly different amino acids. can not detect the amino acid sequence of SFPP.Studying the SFPP was added rat adrenal tumor cells (PC12), Third Generation of the rat bone marrow stromal cells (BMSCs) and spinal cord cells cultured cells to study the role of the biological activity of SFPP. SFPP and crude extracts of the Sargassum fulvellun could promote tumor cell rat adrenal pheochromocytoma (PC12) proliferation and neural differentiation, and there are similar trends in dose-dependent. SFPP and NGF can promote tumor cell rat adrenal pheochromocytoma (PC12) Neural differentiation and synergy. In the 3rd generation of PC12 cell culture system with 5 groups separately with different doses (50ug/mL, 100ug/mL, 150ug/mL, 200ug/mL, 250ug/mL) of SFPP and crude extract of Sargassum fulvellun and the blank the control group, the application MTT assay detected concentrations from 50ug/mL to 150ug/mL SFPP and no rib when the crude extract of Sargassum could promote the proliferation of PC12 and PC12 with the increase of concentration of the proliferation of the rise, when the concentration value for the 150ug/mL the highest activity when the PC12. From 150ug/mL to 250ug/mL downward trend is the proliferation of PC12. SFPP by inverted electron microscope can promote neuronal differentiation of PC12, the trend remains the same dose. Add 5 separate groups of different dose levels (5ng/mL, 10ng/mL, 15ng/mL, 20ng/mL, 25ng/mL) of NGF compared with the control group, NGF can promote neuronal differentiation of PC12, there are dose dependent, from 5ng / mL to a concentration of 20ng/mL NGF can promote neuronal differentiation of PC12 and PC12 with the increase of concentration of an upward trend in the degree of differentiation, when the concentration of 20ng/mL reached the highest value when the regeneration rate. From 20ng/mL 25ng/mL the PC12 differentiation rate to show a downward trend. 20ng/mL NGF concentrations and 5 different concentrations of SFPP dose group interaction, detected a dose-dependent differentiation of PC12 and synergy.SFPP 3rd generation can contribute to bone marrow stromal cells (BMSCs) and spinal cord cells, and a dose dependent manner. Normal feeding SD rats 2 weeks, isolated rat bone marrow stromal cells (BMSCs) and spinal cord cells in primary culture to the stable after the passage of the 3rd generation, add five groups of different doses (50ug/mL, 100ug/mL, 150ug/mL , 200ug/mL, 250ug/mL) of SFPP, and compare the L-DMEM and RPMI-1640 complete medium in two different effects on their proliferation. Application of MTT assay detection SFPP on bone marrow stromal cells (BMSCs) and there is proliferation of spinal cord cells in a dose dependent manner. 10ug/mL to 50ug/mL from the concentration of bone marrow stromal cells (BMSCs) proliferation on the rise, the concentration of 50ug/mL is the highest proliferation value. The decline from 50ug/mL to 250ug/mL. System of the middle of the spinal cord cells in a dose dependent manner, SFPP concentration 100ug/mL dose group is to promote the optimal dose of spinal cord cells. L-DMEM medium on bone marrow stromal cells (BMSCs) in the proliferation rate higher than the RPMI-1640, but in the spinal cord cell culture system of spinal cord cells of two medium effect was the opposite.The study of the biological activities peptides from the Sargassum fulvellun , hoping for direction from the cellular and molecular treatment of brain disease to help.