【摘要】：生物醫(yī)用材料與血液相互接觸時(shí)，首先血漿蛋白質(zhì)會吸附到材料表面，從而觸發(fā)后續(xù)的凝血級聯(lián)過程，因此血漿蛋白質(zhì)的吸附對后續(xù)的凝血過程起到?jīng)Q定性的作用。通過對材料表面改性以提高其阻抗血漿蛋白非特異性吸附能力，是提高生物醫(yī)用材料血液相容性的有效方法之一。 本文使用K5多糖合成具有不同硫酸化位點(diǎn)的硫酸乙酰肝素(Heparan sulfate，HS)類似物，探討不同硫酸化位點(diǎn)的硫酸乙酰肝素類似物在材料表面與血漿蛋白質(zhì)的吸附情況，以期得到一種硫酸乙酰肝素類似物，能夠有效抵抗血漿蛋白質(zhì)的非特異性吸附，用于構(gòu)建蛋白質(zhì)不吸附材料，改善材料表面的血液相容性。 本文以LB為培養(yǎng)基，培養(yǎng)了大腸桿菌K5菌株，采用醇沉分離了K5粗多糖，對K5粗多糖進(jìn)行了脫蛋白處理，采用DEAE陰離子交換樹脂、G75葡聚糖凝膠對K5多糖進(jìn)行了純化，得到了糖醛酸含量為33.1%的K5多糖，并用紅外、核磁對其進(jìn)行了表征。 采用三氧化硫吡啶法對K5多糖N位進(jìn)行了硫酸化，成功制備了NSK5多糖，并用紅外對其進(jìn)行了表征。測定了其抗氧化活性，結(jié)果顯示一定濃度范圍內(nèi)，NSK5多糖的抗氧化活性較K5多糖高。 通過搖瓶培養(yǎng)成功表達(dá)了四種硫酸基轉(zhuǎn)移酶AST-IV、2-OST、6-OST-1、3-OST-1。并用上述硫酸基轉(zhuǎn)移酶對NSK5多糖進(jìn)行了定位硫酸化修飾，合成了3種HS類似物，對合成的產(chǎn)物進(jìn)行了二糖分析，測定了其抗Xa因子活性，結(jié)果顯示硫酸基成功轉(zhuǎn)移到特定的位點(diǎn)上，產(chǎn)物具有良好的抗Xa因子活性。 將HS衍生物通過共價(jià)鍵接枝到硅片表面，并通過水接觸角、XPS、AFM對接枝的情況進(jìn)行了研究，結(jié)果顯示HS衍生物成功接枝到硅片表面。最后采用125I同位素標(biāo)記法研究了改性后硅片表面對纖維蛋白原的吸附情況，結(jié)果顯示接枝了26NSK5和NSK5多糖的硅片，與氨基化處理后的硅片相比，對纖維蛋白原的吸附分別可以降低69.2%、90.5%。細(xì)胞粘附實(shí)驗(yàn)結(jié)果顯示，63NSK5、26NSK5、263NSK5改性硅片對HMEC-1細(xì)胞的粘附效果均得到改善。
[Abstract]:When biomedical material and blood contact with each other, the plasma protein will first adsorb to the surface of the material, thus triggering the subsequent coagulation cascade process. Therefore, the adsorption of plasma protein plays a decisive role in the subsequent coagulation process. It is one of the effective methods to improve the blood compatibility of biomaterial by modifying the surface of the material to improve the nonspecific adsorption ability of impedance plasma protein. In this paper, acetylheparin sulfate (Heparan sulfate,HS) analogues with different sulfation sites were synthesized by using K5 polysaccharide, and the adsorption of heparin sulfate analogues with different sulfation sites on the surface of the materials was studied, and the adsorption of heparin sulfate analogues with plasma proteins was studied. In order to obtain a heparin sulfate analogue, it can effectively resist the non-specific adsorption of plasma protein, and can be used to construct protein-non-adsorptive material and improve the blood compatibility of the surface of the material. In this paper, Escherichia coli strain K5 was cultured on LB medium, the crude K5 polysaccharide was separated by alcohol precipitation, the crude K5 polysaccharide was deproteinized, the K5 polysaccharide was purified by DEAE anion exchange resin and G75 dextran gel. K _ 5 polysaccharide containing 33.1% uronic acid was obtained and characterized by IR and NMR. The N site of K5 polysaccharide was sulfated by thiopyridyl trioxide method. The NSK5 polysaccharide was successfully prepared and characterized by IR. The results showed that the antioxidant activity of NSK5 polysaccharide was higher than that of K5 polysaccharide in a certain concentration range. Four sulfate transferase AST-IV,2-OST,6-OST-1,3-OST-1. were successfully expressed in shaking flask culture. Three kinds of HS analogues were synthesized by sulfation modification of NSK5 polysaccharides with the above sulfate transferase. The synthesized products were analyzed by disaccharide analysis and their anti-Xa factor activities were determined. The results showed that the sulfate group was successfully transferred to a specific site, and the product had good anti-Xa factor activity. HS derivatives were grafted onto the surface of silicon wafers by covalent bonding, and the grafting conditions were studied by water contact angle and XPS,AFM. The results showed that the HS derivatives were successfully grafted onto the surface of silicon wafers. Finally, the adsorption of fibrinogen on the surface of modified silicon wafer was studied by 125i isotope labeling method. The results showed that the modified silicon wafer grafted with 26NSK5 and NSK5 polysaccharide was compared with the modified silicon wafer. The adsorption of fibrinogen decreased by 69.2% and 90.5% respectively. The results of cell adhesion test showed that the adhesion effect of 63 NSK5 and 26NSK5263NSK5 modified silicon wafers on HMEC-1 cells was improved.
【學(xué)位授予單位】：江南大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R318.08

【參考文獻(xiàn)】

相關(guān)期刊論文 前6條

1 馮桂龍,王松,朱鶴孫;絲素改性膠原膜的肝素化及其體外抗凝血性能評價(jià)[J];功能材料;2005年01期

2 王海波;王進(jìn);王曉娜;孫鴻;黃楠;;等離子體聚合沉積聚烯丙胺薄膜改性醫(yī)用不銹鋼及內(nèi)皮細(xì)胞粘附行為研究[J];功能材料;2008年07期

3 許海葉;楊紅軍;王維慈;歐陽晨曦;徐衛(wèi)林;;肝素化絲素/聚氨酯共混膜的制備及其緩釋性能[J];武漢科技學(xué)院學(xué)報(bào);2008年07期

4 李全輝;徐祝軍;李全利;沈軍;張維波;;膠原-肝素自組裝多層膜修飾鈦表面[J];中國組織工程研究與臨床康復(fù);2010年51期

5 孫梅;劉雅君;;蛋白質(zhì)藥物涂層心臟支架的研究進(jìn)展[J];藥學(xué)服務(wù)與研究;2010年02期

6 蔡開勇,姚康德;組織工程生物材料的表面修飾[J];中國康復(fù)理論與實(shí)踐;2002年05期

，

本文編號：2434077