The active fraction was finally separated by RP-HPLC (LC-10A, Shimadzu) with an RP ECOSIL C18 column (4.6 mm ± 250 mm, 5 mm, Lubex, Japan). Elution was performed using 100% double-distilled water with 0.1% trifluoroacetic acid. The flow rate was 1 mL/min and the absorbance was recorded at 214 and 280 nm. The active fraction was concentrated and lyophilized.