Separation and Quantification of Theanine Enantiomers Using High Performance Liquid Chromatography Coupled with Chiral Derivatization

L-Theanine is a unique non-protein amino acid in tea and shows many physiological functions. To directly extract L-theanine from tea is a lengthy and very costly process. Although the chemical synthesis of theanines is simple the product is unfortunately a mixture of DL-enantiomers. A chiral derivatization method was developed for the separation and quantification of theanine enantiomers by reversed-phase high performance liquid chromatography RPHPLC. 1-Fluoro-2 4-dinitrophenyl-5-L-alanine amide FDAA was used as the chiral reagent. This method showed good linearity for both L-theanine ranging from 1. 732 x 10(-3) to 2. 077 mu g and D-theanine ranging from 1. 696 x 10(-3) to 2. 044 pRg. The recoveries were in the range of 97.3% -102. 0% for L-theanine and 97.2% -103. 2% for D-theanine. This method also showed excellent limit of detection similar to 5 x 10(-4) mu g and limit of quantification similar to 1 x 10(-3) mu g for both L-theanine and D-theanine. The results demonstrated that this method is precise accurate and can be used for the determination of theanine enantiomers.