为了提高胶原耐酶解能力,以N-羟基琥珀酰亚胺-己二酸酯(NHS-AA)为交联剂(浓度CNHS-AA为0.25~4.0mmol/L)交联胶原,采用质量法、三硝基苯磺酸法、差示扫描量热法和十二烷基硫酸钠-丙烯酰胺凝胶电泳等对胶原耐胰蛋白酶解的性能进行了研究。结果显示,当CNHS-AA为1.5 mmol/L时,胶原酶解重量保持率达到最大,自由氨基浓度达到最低,胶原的变性温度达到最高;酶解产物电泳谱带的颜色随CNHS-AA增加而逐渐变浅。研究表明,NHS-AA与胶原的自由氨基反应形成的交联网状结构有效地阻碍了胰蛋白酶渗透进入胶原材料,从而提高了胶原抗酶解能力,当CNHS-AA=1.5 mmol/L时,胶原抗酶解能力达到最强。由此可见,可以通过改变交联度来控制胶原胰蛋白酶降解的速率。
In order to improve the enzymatic resistance of collagen,collagen was cross-linked with N-hydroxysuccinimide activated adipic acid(NHS-AA) at concentrations ranging from 0.25 mM to 4 mM.The results demonstrated that the digested collagen cross-linked with 1.5 mM NHS-AA had the maximum weight remaining,minimum amino groups determined by trinitrobenzensulfonic acid(TNBS) assay,and the maximum denaturation temperature,obtained by means of differential scanning calorimeter(DSC).Meanwhile,with increasing NHS-AA concentration,color of peptide maps of collagen digested by trypsin,obtained by means of sodium dodecyl sulphate polyacrylamide gel electrophoresis(SDS-PAGE),gradually became lighter.All shown enzymatic resistance of collagen was improved significantly owing to interference of the penetration of the enzyme into the cross-linked collagen network.The results demonstrated that collagen cross-linked with 1.5 mM NHS-AA had optimum enzymatic degradability due to the highest cross-linking density of collagen.The susceptibility of collagen towards enzymatic degradation could be controlled by varying the degree of cross-linking of the samples.