色谱

色谱 ›› 2012, Vol. 30 ›› Issue (02): 184-189.DOI: 10.3724/SP.J.1123.2011.10037

• 研究论文 • 上一篇    下一篇

柱前衍生高效液相色谱法测定茶叶中茶氨酸及γ-氨基丁酸含量

涂云飞*, 杨秀芳, 张士康, 朱跃进   

  1. 中华全国供销合作总社杭州茶叶研究院, 浙江 杭州 310016
  • 收稿日期:2011-10-31 修回日期:2011-11-29 出版日期:2012-02-28 发布日期:2012-03-22
  • 通讯作者: 涂云飞,助理研究员，研究领域为茶叶生物化学和分析化学。

Determination of theanine and γ-aminobutyric acid in tea by high performance liquid chromatography with precolumn derivatization

TU Yunfei*, YANG Xiufang, ZHANG Shikang, ZHU Yuejin   

  1. Hangzhou Institute of Tea, All China Federation of Supply and Marketing Cooperatives, Hangzhou 310016, China
  • Received:2011-10-31 Revised:2011-11-29 Online:2012-02-28 Published:2012-03-22
  • Contact: TU Yunfei, assistant researcher. Research field: tea biochemistry and analytical chemistry. Tel: (0571)87016079,
  • Supported by:

    Zhejiang Provincial Natural Science Foundation of China (No. Y3100683)

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摘要： 对茶叶中的茶氨酸和γ-氨基丁酸(GABA)建立了柱前衍生方法,并利用高效液相色谱法(HPLC)对衍生物进行了测定。采用邻苯二甲醛(OPA)及N-乙酰-L-半胱氨酸(NAC)为衍生试剂,考察了脯氨酸(Pro)、茶多酚(Tp)及维生素C(Vc)等对衍生体系的影响,并对色谱分离条件进行了优化。结果表明,茶氨酸与GABA衍生物在色谱柱上的保留行为与分离效率主要受流动相pH及缓冲盐浓度的影响;Pro、Tp及Vc对茶氨酸的衍生产率影响较小。而Vc可以提高GABA检测的灵敏度,并能稳定衍生用储备液。GABA与茶氨酸衍生后的方法检出限(LOD)分别为3.01×10~5mmol/L和7.98×10~5mmol/L;定量限(LOQ)分别为9.99×10~5mmol/L和2.658×10~4mmol/L;线性范围分别为0.01～0.4 mmol/L(相关系数为0.996)和0.05～0.8 mmol/L(相关系数为0.995);方法的回收率分别为99.29%～119.60%和93.18%～141.06%(除本身含茶氨酸量高的绿茶中回收率为62.88%外)。这说明经优化的柱前衍生-高效液相色谱体系可用于茶叶中两种特征性氨基酸的测定。

关键词: γ-氨基丁酸, N-乙酰-L-半胱氨酸, 茶, 茶氨酸, 高效液相色谱, 邻苯二甲醛, 柱前衍生

Abstract: A method of precolumn derivatization-high performance liquid chromatography (HPLC) for the determination of theanine and γ-aminobutyric acid (GABA) in tea was established. o-Phthalaldehyde (OPA) and N-acetyl-L-cysteine (NAC) were chosen as the derivatization reagents. The effects of teapolyphenol (Tp), proline (Pro) and Vitamin C (Vc) on derivatization yields were investigated. The results indicated that Vc not only stabilized the stock solution of OPA, but also enhanced the yield of GABA derivative. However, the yield of theanine derivative was less affected. The HPLC separation system was also optimized. The resolution of the derivatives was improved by adjusting the pH value and phosphate-citric buffer concentration of the mobile phase. The limits of detection (LODs) for GABA and theanine were 3.01×10~5mmol/L and 7.98×10~5mmol/L, and the limits of quantification (LOQs) were 9.99×10~5mmol/L and 2.658×10~4mmol/L, respectively. The linear ranges of GABA and theanine were 0.01~0.4 mmol/L with the correlation coefficient of 0.996 and 0.05~0.8 mmol/L with the correlation coefficient of 0.995, respectively. The main recoveries for GABA and theanine in green tea, Oolong tea, and black tea, ranged from 99.29% to 119.60% and from 62.88% to 141.06% respectively. The method with simple procedure and efficient separation was proved to be suitable for the determination of GABA and theanine in tea.

Key words: γ-aminobutyric acid, N-acetyl-L-cysteine, o-phthalaldehyde, precolumn derivatization, tea, theanine, high performance liquid chromatography (HPLC)