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高效毛细管电泳法直接测定红豆杉悬浮培养细胞中游离芳香族氨基酸
杨新,梅兴国,汪爱顺
华中科技大学生命科学与技术学院,湖北,武汉,430074
摘要:
建立了红豆杉悬浮培养细胞中游离芳香族氨基酸的高效毛细管电泳的分离直接紫外吸收测定方法.在优化的实验条件下,以间苯二酚为内标,未涂层融硅毛细管(75 μm i.d.,总长37cm,有效分离长度30cm)为分离通道,40 mmol/L磷酸缓冲液(pH10.46)为电泳介质,3sec(0.5psi)压力进样,15 kV恒压电泳,200 nm检测,在9 min内三种氨基酸得到分离测定.色氨酸、苯丙氨酸和酪氨酸的线性范围分别为2.50~250 μmol/L(r=0.9962,RSD=2.52%)、1.25~250μmol/L(r=0.9966,RSD=2.12%)和2.50~250 μmol/L(r=0.9940,RSD=2.93%),苯丙氨酸回收率为96.8%±1.37%,酪氨酸回收率为99.2%±3.04%.
关键词:  高效毛细管电泳  芳香族氨基酸  直接测定
DOI:
分类号:Q503
基金项目:国家科技攻关项目,国家高技术研究发展计划(863计划)
Direct Determination of Free Aromatic Amino Acids in Suspension of Taxus chinensis Cell by High Performance Capillary Eletrophoresis
Abstract:
A method for separation and direct determination of free aromatic amino acids in the suspension of Taxus chinensis cell by high performance capillary eletrophoresis with UV-absorbance was established. The optimum experimental conditions ware the following: separation column: bare fused-sillica capillary (75μm i.d.,total length 37cm,30cm from the inlet to UV detector); electrolyte: 40mmol/L phosphate buffer,pH 10.46;detection wavelength:20nm;sampling:5sec(0.5 psi);running voltage:15kV,polarity from anode(inlet) to cathode(outlelt);temperature:20℃;internal standard:resorcin. The linear ranges of determination for Trp,Phe and Try were 2.50~250μmol/L(r=0.9962,RSD=2.52%),1.25~250μmol/L(r=0.9966,RSD=2.12%)and 2.50~250μmol/L(r=0.9940,RSD=2.93%) respectively. The recoveries of Phe and Try were 96.8%±1.37% and 99.2%±3.04%.
Key words:  high performance capillary eletrophoresis  aromatic amino acid  direct determination