Box-Behnken设计毛细管液相色谱法拆分α-氨基酸对映体的研究

将L-脯氨酸通过3-缩水甘油丙基三甲氧基硅烷键合到球型多孔硅胶基质上,制备了手性配体交换色谱固定相.采用高压匀浆法将制备的填料填充到毛细管柱中,在自组装的毛细管液相色谱仪上对3种α-氨基酸对映体进行了拆分.利用Box-Behnken设计对影响因素(流动相中乙腈的体积分数、中心离子Cu~(2+)的浓度、流速)进行优化,以最大保留时间和分离度为响应值,借助二次多元回归方程,建立了相应的数学模型.响应曲面图表明:DL-天冬酰胺在5%乙腈、5 mmol/L Cu~(2+)、流速0.139 μL/min,苏氨酸在10%乙腈、5 mmol/L Cu~(2+)、流速0.167 μL/min,丝氨酸在5%乙腈、7 mmol/L Cu~(2+)、流速0.167 μL/min的条件下均能得到基线分离.表明通过所制备的手性固定相,采用配体交换色谱模式可获得拆分DL-氨基酸的理想效果.

Enantioseparation of α-Amino Acids by Capillary Liquid Chromatography Using Box-Behnken Design

A chiral stationary phase was prepared by treating porous silica gel with 3-glycidoxypropyl-trimethoxysilane and bonding L-proline. The prepared chiral materials were packed into a capillary column by slurry packing technique and applied for the enantioseparation of three α-amino acids using the self-installed capillary liquid chromatography. Box - Behnken experimental design based on three-level, three-variable designs was used for the optimization with respect to the percentage of eluent, copper( Ⅱ ) concentration and flow rate. Statistical interpretation of the variables on different responses such as resolution and retention time of the last component was performed. The optimum conditions of these variables were obtained by using a second-order polynomial model. The result indicated that, under pH 7. 0 buffer, a good baseline was obtained by using ACN percentage of 5% , copper ( Ⅱ ) concentration of 5 mmol/L and a flow rate of 0. 139 μL/min for asparagine, ACN percentage of 10% , copper(Ⅱ) concentration of 5 mmol/L and a flow rate of 0. 167 μL/min for threonine, and ACN percentage of 5% , copper(Ⅱ) concentration of 7 mmol/L and a flow rate of 0. 167 μL/min for serine, respectively. Therefore, the method could applied in the separation of a-amino acids enanti-omer.