**2.5 Method validation**

Calibration curves were constructed from duplicate analyses of GABA and theanine standards at 1.2, 2.4, 4.8, 7.2, 9.6 and 12.0 μM, and triplicate analyses of caffeine and EGCG standards at 5, 50, 100, 160 and 200 μM. To evaluate the intraday and interday precision, 96 μM GABA and theanine standards were analysed in duplicate of two consecutive days, 50 μM caffeine and 160 μM EGCG standards were analysed in triplicate on three consecutive days, and coefficients of variation (CV) for the retention time and peak area were calculated. Accuracy was assessed using a recovery test of spiked samples with 50 μM of each of GABA, theanine, caffeine and EGCG. Recoveries were calculated comparing the obtained amounts with those added using the formula: recovery (%) = [(concentration found – endogenous concentration) / 50] × 100% (Zhao *et al*., 2011). Validation of GABA, theanine, caffeine and EGCG peak identification in the samples was conducted by comparing the retention time of the peak of increased size in the spiked to the unspiked samples.

*GABA-enriched Oolong Tea: Reducing Stress in a Student Cohort May Involve More than Just… DOI: http://dx.doi.org/10.5772/intechopen.106896*
