使用混合物设计和响应面方法对α-胰凝乳蛋白酶催化的N-Ac-Phe-Tyr-NH2合成进行溶剂选择和优化,Biotechnology Progress

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使用混合物设计和响应面方法对α-胰凝乳蛋白酶催化的N-Ac-Phe-Tyr-NH2合成进行溶剂选择和优化
Biotechnology Progress ( IF 2.5 ) Pub Date : 2012-10-18 , DOI: 10.1002/btpr.1623
Shih-Hao Hu , Chia-Hung Kuo , Chieh-Ming J. Chang , Yung-Chuan Liu , Wen-Dee Chiang , Chwen-Jen Shieh

的肽，Ñ -Ac-PHE-酪氨酸- NH ₂，与血管紧张素I转化酶（ACE）抑制剂活性通过的α胰凝乳蛋白酶催化的缩合反应合成ñ -乙酰基苯基丙氨酸乙酯（Ñ -Ac-Phe- OEt）和酪氨酸酰胺（Tyr-NH ₂）。三种溶剂：Tris-HCl缓冲液（80 mM，pH 9.0），二甲基亚砜（DMSO）和乙腈被用于本研究中。通过单纯形质心混合物设计确定最佳反应溶剂组分。在有机水溶液（Tris-HCl缓冲液：DMSO：乙腈= 2：1：1）中，其中N -Ac-Phe-Tyr-NH ₂的产率为73.55％，合成效率得到了提高。可以实现。此外，使用中央复合可旋转设计（CCRD）通过响应面方法（RSM）评估了反应参数对产率的影响。基于最大峰分析，该肽合成的最佳条件包括：反应时间为7.4分钟，反应温度为28.1°C，酶活性为98.9 U，底物摩尔比（Phe：Tyr）为1： 2.8。预测的和实际的（实验）产率分别为87.6和85.5％。实验设计和RSM在优化N -Ac-Phe-Tyr-NH ₂合成方面表现良好，因此有望成为获得高产酶促肽的有效方法。©2012美国化学工程师学会生物技术研究所。Prog。，2012年

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Solvent selection and optimization of α‐chymotrypsin‐catalyzed synthesis of N‐Ac‐Phe‐Tyr‐NH2 using mixture design and response surface methodology

A peptide, N‐Ac‐Phe‐Tyr‐NH₂, with angiotensin I‐converting enzyme (ACE) inhibitor activity was synthesized by an α‐chymotrypsin‐catalyzed condensation reaction of N‐acetyl phenylalanine ethyl ester (N‐Ac‐Phe‐OEt) and tyrosinamide (Tyr‐NH₂). Three kinds of solvents: a Tris–HCl buffer (80 mM, pH 9.0), dimethylsulfoxide (DMSO), and acetonitrile were employed in this study. The optimum reaction solvent component was determined by simplex centroid mixture design. The synthesis efficiency was enhanced in an organic‐aqueous solvent (Tris‐HCl buffer: DMSO: acetonitrile = 2:1:1) in which 73.55% of the yield of N‐Ac‐Phe‐Tyr‐NH₂ could be achieved. Furthermore, the effect of reaction parameters on the yield was evaluated by response surface methodology (RSM) using a central composite rotatable design (CCRD). Based on a ridge max analysis, the optimum condition for this peptide synthesis included a reaction time of 7.4 min, a reaction temperature of 28.1°C, an enzyme activity of 98.9 U, and a substrate molar ratio (Phe:Tyr) of 1:2.8. The predicted and the actual (experimental) yields were 87.6 and 85.5%, respectively. The experimental design and RSM performed well in the optimization of synthesis of N‐Ac‐Phe‐Tyr‐NH₂, so it is expected to be an effective method for obtaining a good yield of enzymatic peptide. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012

更新日期：2012-10-18

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