Polyphenolic Fingerprinting via HPLC

High-pressure liquid chromatography (HPLC) was used to evaluate the polyphenolic fingerprinting of the extract as described above [50 (link)]. HPLC-DAD analyses were performed using a Shimadzu LC 20 (Kyoto, Japan), which was equipped with a diode array detector (DAD) and with a 150 × 4.6 mm i.d., 2.7 μm Ascentis Express C 18 column. The mobile phases were as follows: H₂O/H₃PO₄ (99:1, solvent A), MeOH/CAN/H₃PO₄ (49,5:49,5:1, solvent B). The gradient used was as follows: concentration of the solvent A of 95% going to 77% (34 min), maintained at 77% (3 min), 74% (60 min), 60% (85 min), 20% (90 min), and 0% (92 min). The total time was 105 min. The column temperature was maintained at 25 °C. The flow was 1 mL/min, and the injection volume was 5 μL. The chromatogram profiles were recorded from 190 to 500 nm and monitored at 280 and 330 nm ± 2 nm.