柱前衍生化RP-HPLC测定白芷中氨基酸含量

Determination of Amino Acids Content in Angelica by Pre- column Derivatization RP- HPLC

目的建立柱前衍生RP-HPLC法测定白芷中游离氨基酸含量的方法，并测定不同产地的白芷样品中游离氨基酸含量。方法样品以异硫氰酸苯酯（PITC）为衍生化试剂，经与白芷中氨基酸柱前衍生，并以高效液相色谱法测定其含量，色谱条件采用WatersSymmetryRPl8色谱柱（4．6mm×250mm，5μm），流动相以[50mmol／L乙酸钠缓冲液（以冰醋酸调至pH6．5）]（A）-[乙腈-甲醇-水（60：20：20）]（B）进行梯度洗脱，柱温35℃，流速1．0mL／min，检测波长254nm。结果17种氨基酸均能达到基线分离，17种氨基酸的平均回收率在95．48％～104．30％之间，RSD在3．2％-4．9％之间（n=6）。结论柱前衍生RP-HPLC法灵敏，准确，重现性和稳定性良好，适用于对白芷中的氨基酸含量进行研究，为白芷药材的质量控制和评价提供了科学依据，不同产地的白芷样品中氨基酸的含量存在差异，必需氨基酸的比例也存在差异。

Objective： To establish a pre -column derivatization RP- HPLC method for the determination of free amino acid content inAngelica and measure free amino acid content inAngelica samples from different origins. Methods： Phenyl isothiocyanate （PITC） as deri- vatization reagent samples were derivatized by pre - column amino acids inAngelica and HPLC was used to determine the content. Chroma- tographic column was Waters Symmetry RP18 （4. 6mm ×250mm, 5μm） and mobile phase was 50mmol/L sodium acetate buffer （adjusted with glacial acetic acid pH6. 5 ） （A） and acetonitrile - methanol - water （60 ： 20 ： 20） （ B ） as gradient elution, at 35 ℃ of column tempera- ture, 1.0 mL/min of flow rate and 254nm of detection wavelength. Results： 17 kinds of amino acids could reach baseline separation, the average recovery at the range of 95.48% to 104. 30% , RSD from 3.2% to 4. 9% （ n = 6）. Conclusion ： The pre - column derivatization RP - HPLC method is sensitive and accurate with reproducibility and good stability, suitable for the determination of amino acid content inAngelica, and provide a scientific basis for the quality control and evaluation of Angelica medicinal material. The amino acid contents inAngelica samples from different origins have differences and the proportions of essential amino acids also have differences.