Quantitative Analysis of Catechins and Free Amino Acids

Samples were assigned and extracted as for UPLC-Q-TOF/MS for catechin determination [7 (link)]. Free amino acid samples were analyzed as per the instructions provided by the waters AccQ.Tag chemistry package. HPLC analysis was carried out using an e2695 connecting 2998 photodiode array detector system (Waters) injected with 10 µL and 25 µL sample solutions for catechin and free amino acids, respectively. For catechins, distilled water with 2% formic acid was used as mobile phase A. Mobile phase B consisted of the HPLC solvent ACN. The samples were eluted at a column temperature of 40 ± 1 °C and a flow rate of 1 mL/min and monitored at 278 nm. Similarly, for amino acids, AccQ.Tag eluent from waters was used as mobile phase A. Mobile phase B was can, and the column temperature was set at 37 ± 2 °C. The remainder of the procedure followed the method prescribed in the AccQ. Tag chemistry package instruction manual. Peaks of catechins were identified by comparing sample retention time to those of authentic standards, and amino acid peaks were as prescribed in the manual.