Tandem use of carboxypeptidase Y reactor and displacement chromatograph for peptide synthesis.
Tandem use of carboxypeptidase Y reactor and displacement chromatograph for peptide synthesis.
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串联使用羧肽酶 Y 反应器和置换色谱仪进行肽合成。
DOI：
10.1016/s0021-9673(01)94182-2
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发表时间：
1987
期刊：
影响因子：
--
通讯作者：
Horváth,C
中科院分区：
文献类型：
--
作者：
Cramer,SM;elRassi,Z;Horváth,C
将具有固定化羧肽酶 Y 的填充床酶反应器与置换色谱仪串联使用，从 N-苯甲酰-L-精氨酸和 L-甲硫氨酰胺制备 N-苯甲酰-L-精氨酰-L-甲硫氨酰胺。耦合酶反应器和置换色谱仪的泵和阀门由微处理器控制。该酶通过戊二醛固定在微粒氨基二氧化硅上，并装入 60 × 4.6 mm I.D. 中。柱子。填充床反应器以再循环模式使用，随后使用丁氧基乙氧基乙醇作为置换剂，在250×4.6mm十八烷基二氧化硅柱上通过置换色谱法分离反应混合物的组分。将未反应的1-甲硫氨酰胺返回到反应混合物中。通过高效液相色谱分析监测反应进程和置换色谱分离程度。该系统的设计使得酶促肽合成、置换色谱分离和柱再生通过并联使用两个相同的柱同时进行。该系统在24小时内可得到460mg纯度大于99％的N-苯甲酰基-1-精氨酰基-1-甲硫氨酰胺。酶反应器和以置换模式操作的液相色谱仪的串联操作为肽的合成和纯化提供了一种手段。
A packed-bed enzyme reactor with immobilized carboxypeptidase Y was used in tandem with a displacement chromatograph for the preparation of N-benzoyl-l-arginyl-l-methioninamide, from N-benzoyl-l-arginine andl-methioninamide. The pumps and valves of the coupled enzyme reactor and displacement chromatograph were controlled by a microprocessor. The enzyme was immobilized on microparticulate amino-silica by glutaraldehyde and packed into a 60 × 4.6 mm I.D. column. The packed-bed reactor was used in the recirculating mode and components of the reaction mixture were subsequently separated by displacement chromatography on a 250 × 4.6 mm octadecyl-silica column using butoxyethoxyethanol as the displacer. Unreactedl-methioninamide was returned to the reaction mixture. Both the progress of the reaction and the extent of separation by displacement chromatography were monitored by high-performance liquid chromatographic analysis. The system was designed so that enzymatic peptide synthesis, separation by displacement chromatography, and column regeneration were carried out simultaneously by using two identical columns in parallel. An amount of 460 mg of N-benzoyl-l-arginyl-l-methioninamide having purity greater than 99% could be obtained in 24 h with this system. The tandem operation of the enzyme reactor and liquid chromatograph operated in the displacement mode offers a means for the synthesis and purification of peptides.