Chemoenzymatic Synthesis Of HS Analogues、Biological Activity And Protein Adsorption Behavior On Mateiral Surface

When biomedical material surface contacted with blood, the plasma protein adsorbed to thesurface at first, thereby triggering the subsequent process of the coagulation cascade, so theadsorption of plasma proteins played a decisive role in the subsequent coagulation process,improving the impedance of the non-specific plasma protein adsorption capacity by surfacemodification of biomaterials is an effective and emerging way to improve the bloodcompatibility of biomedical materials.In this thesis, in order to get a heparan sulfate analogues can effectively resist theadsorption of plasma proteins, which can be used to build anti-biofouling materials, finallyimprove the blood compatibility of materials, a set of heparan sulfate analogues with differentsulfation points were synthesized by chemoenzymatic methods from K5polysaccharide, theheparan sulfate analogues were covalently immobilized to silicon wafers, and its proteinadsorption behavior were investigated.E.coli K5strain was cultivated by fermentation with LB, rough K5polysaccharide wasextracted by ethanol precipitation, and then purified by Sevage method, DEAE anionexchange resin and Sephadex G-75column, finally uronic acid content of33.1%of the K5polysaccharide was obtained, and it was characterized by IR and NMR.N site of the K5polysaccharide was chemically sulfated by Sulfur trioxide pyridinecomplex, NSK5polysaccharide was prepared, and it was characterized by IR. The antioxidantcapacity of N-sulfated K5polysaccharide was investigated, the results indicated that NSK5showed stronger antioxidant activity than K5in certain concentration range.Four sulfotransferases AST-IV,2-OST,6-OST-1,3-OST-1were successfully expressed byshake flask. Finally, three heparan sulfate analogues were synthesized by the abovesulfotransferases from NSK5. Synthetic products were identified by disaccharide analysis andits anti-factor Xa activity was determined, the results showed that sulfate was successfullytransferred to the specific point; the product has a good anti-factor Xa activity.HS analogues were covalently grafted to the silicon wafers, and the modified silicon waferswere investigated by water contact angle, XPS, and AFM, the results showed that the HSanalogues were successfully grafted to the silicon wafers. Finally, the125I isotope labelingmethod was used to study the adsorption behavior of fibrinogen on modified silicon wafers,and the results revealed that when compared to NH2-terminated silicon wafers, the adsorptionof fibrinogen of26NSK5and NSK5grafted silicon wafers were reduced to69.2%and90.5%,respectively. The cell adhesion test results showed that HMEC-1cell adhesion was improvedwhen silicon wafers grafted with63NSK5,26NSK5,263NSK5polysaccharide.