摘要
目的:研究使用Wang树脂与Fmoc保护策略固相合成抗凝血药比伐卢定(bivalirudin)的实验方法。方法:比伐卢定的合成经过了逐步偶联、TFA裂解、脱保护、HPLC纯化等一系列步骤。为了优化反应条件,选择了不同氨基酸取代度的树脂(0.29、0.48、0.59mmol/g)和不同的缩合试剂(HBTU、DCC、HATU)进行试验。结果:成功得到了纯度为98%的比伐卢定。活性测定表明,不同方法合成的比伐卢定均具有与标准样品相当的抗凝活性,并且显示:①氨基酸取代度为0.3～0.4mmol/g的Wang树脂可以得到产率及纯度均较高的产品,取代度太大(>0.5mmol/g)得到的产品副产物多、纯度低;②用HBTU/HOBt为缩合试剂具有操作方便、成本经济、合成效率高等优点。根据以上实验结果选择优化条件,用5gWang树脂合成克级单位的比伐卢定,得到的粗品产率可达84%,纯度可达68%以上。结论:通过固相合成的方法,考察了不同氨基酸取代度、不同缩合试剂及规模大小对合成比伐卢定的产率、纯度及活性的影响,对比伐卢定的大量合成具有一定的指导作用。
Objective：To study the solid-phase synthesis of the thrombin inhibitor bivalirudin using Wang Resin and Fmoc-protection strategy.Methods：Bivalirudin was synthesized through the process of coupling reaction,TFA cleavage /deprotection and HPLC purification.Different Fmoc-Leu substitution resins（0.29 mmol/g,0.48 mmol/g,0.59 mmol/g） and different coupling reagents（HBTU,DCC,HATU） were used to synthesize the bivalirudin.Results：The purity of bivalirudin could be 〉98%.The bioactive assay showed that the synthetic bivalirudin had the same extent anticoagulant activity as the standard sample.The results showed：① the high Fmoc-Leu substitution resin（〉0.5 mmol /g） results in the low purity product,which was also difficult to purify;the substitution 0.30.4 mmol/g of Fmoc-Leu-Wang Resin was recommended;②using HBTU/HOBt as the coupling reagent had the advantages of convenient operation,economic and high efficiency.Following the results above,the optimized condition was used to synthesize gram grade of the Bivalirudin using 5 g Fmoc-Leu-Wang Resin.The crude product with the 84% total yield and 68% purity was obtained.Conclusion：The solid-phase synthesis method was used to evaluate the effect of different Fmoc-Leu substitution resins,different coupling reagents and different synthesis size on the total yield,purity and activity of bivalirudin.This research has instruction for the synthesis of bivalirudin in large amount.
出处
《南京医科大学学报（自然科学版）》
CAS
CSCD
北大核心
2010年第9期1282-1286,共5页
Journal of Nanjing Medical University(Natural Sciences)
基金
南京大学人才引进项目资助
关键词
凝血酶
固相合成
比伐卢定
缩合试剂
取代度
thrombin
solid-phase synthesis
bivalirudin
coupling reagent
substitution
