Tu Yunfei, Yang Xiufang, Zhang Shikang, Zhu Yuejin
Hangzhou Institute of Tea, All China Federation of Supply and Marketing Cooperatives, Hangzhou 310016, China.
Se Pu. 2012 Feb;30(2):184-9. doi: 10.3724/sp.j.1123.2011.10037.
A method of precolumn derivatization-high performance liquid chromatography (HPLC) for the determination of theanine and gamma-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 x 10(-5) mmol/L and 7.98 x 10(-5) mmol/L, and the limits of quantification (LOQs) were 9.99 x 10(-5) mmol/L and 2.658 x 10(-4) mmol/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.
建立了一种柱前衍生化-高效液相色谱法（HPLC）用于测定茶叶中的茶氨酸和γ-氨基丁酸（GABA）。选用邻苯二甲醛（OPA）和N-乙酰-L-半胱氨酸（NAC）作为衍生化试剂。研究了茶多酚（Tp）、脯氨酸（Pro）和维生素C（Vc）对衍生化产率的影响。结果表明，Vc不仅能稳定OPA储备液，还能提高GABA衍生物的产率。然而，茶氨酸衍生物的产率受影响较小。同时对HPLC分离系统进行了优化。通过调节流动相的pH值和磷酸盐-柠檬酸缓冲液浓度提高了衍生物的分离度。GABA和茶氨酸的检测限（LOD）分别为3.01×10⁻⁵ mmol/L和7.98×10⁻⁵ mmol/L，定量限（LOQ）分别为9.99×10⁻⁵ mmol/L和2.658×10⁻⁴ mmol/L。GABA和茶氨酸的线性范围分别为0.01 - 0.4 mmol/L，相关系数为0.996；0.05 - 0.8 mmol/L，相关系数为0.995。绿茶、乌龙茶和红茶中GABA和茶氨酸的主要回收率分别为99.29%至119.60%和62.88%至141.06%。该方法操作简单、分离高效,适用于茶叶中GABA和茶氨酸的测定。
