柱前衍生反相高效液相色谱法测定 草本水杨梅叶中氨基酸含量

Determination of amino acids in leaves of Geum aleppicum Jacq. by precolumn derivatization reversed-phase high performance liquid chromatography

中文关键词:  草本水杨梅  柱前衍生  氨基酸  高效液相色谱法

英文关键词:Geum aleppicum Jacq.  precolumn derivatization  ammo acid  high performance liquid chromatography

中文摘要:

      目的 采用柱前衍生反相高效液相色谱法对草本水杨梅叶中氨基酸进行测定。方法 样品以异硫氰酸苯酯(PITC) 为衍生化试剂, 与草本水杨梅叶中氨基酸柱前衍生, 采用Ultimate Amino Acid氨基酸分析专用柱(4.6 mm×250 mm, 5 μm); 流动相: A为乙腈-水(80:20, V:V), B为乙腈-三水合乙酸钠缓冲溶液(7:93, V:V, pH 6.5), 梯度洗脱; 柱温35 ℃, 流速1.0 mL/min, 检测波长为254 nm。结果 20种氨基酸在0.039~1.250 mmol/L内呈良好的线性关系(r2>0.9996)。20种游离氨基酸平均回收率在96.51%~108.06%之间, RSD在0.36%~2.08%之间(n=6); 水解氨基酸平均回收率在95.29%~102.13%之间, RSD在0.85 %~2.45%之间(n=6)。结论 该方法可用于草本水杨梅中氨基酸的检测, 其灵敏度高且准确, 具有良好的重复性及稳定性。

英文摘要:

      Objective To determinate and investigate the amino acids in leaves of Geum aleppicum Jacq. after derivatization by a simple and sensitive reversed-phase high performance liquid chromatography method. Methods The amino acids in leaves of G. aleppicum were pretreated with phenyl isothiocyanate (PITC). The separation of G. aleppicum derivatives was carried out on a Cltimate Amino Acid of specific column (4.6 mm × 250 mm, 5 μm). Eluent A was acetonitrile-water (80:20, V:V) and eluent B was a mixture of acetonitrile and sodium acetate trihydrate (7:93, V:V, pH6.5), eluting with a gradient. The column temperature was 35 ℃, flow rate was 1.0 mL/min and the detection wavelength was 254 nm. Results The linear range of 20 amino acids was 0.039~1.250 mmol/L (r2>0.9996). The average recovery rates of 20 free amino were ranged from 96.51% to 108.06 % and RSD values were from 0.36% to 2.08% (n=6). The average recovery of rates of 20 total amino acids was ranged from 95.29% to 102.13% and RSD values were from 0.85% to 2.45% (n=6). Conclusion This method is flexible, accurate, and repeatable, and stable for the determination of amino acids in leaves of G. aleppicum.