0.20 μm filters

All experiments were performed on a 1260 infinity II LC (Agilent Technologies, Santa Clara, CA, USA). The isolated compounds and extract were dissolved in methanol at the concentration of 1 and 3 mg/mL, respectively. They were filtered through Millipore 0.20 μm filters before injection for chromatographic separation (Millex-LG, Tokyo, Japan). Then, 5 μL of each was injected and the flow rate was 0.4 mL/min. The separation was performed on YMC triart-C18 column (4.6 × 150 mm, 5 μm), (YMC Company, Kyoto, Japan) at 40 °C with the gradient system of 0.1% formic acid in water (solvent A) and 0.1% formic acid in methanol (solvent B) as follows 0–2 min, 20% B; 2–22 min, 20–100% B; 22–30 min, 100% B, finally the B content was reduced to the initial conditions in 5 min and the column was re-equilibrated for 5 min. The diode array detector (DAD) was measured over the rage of 200–600 nm. The evaporation temperature of the ELSD detector was set at 80 °C. The nebulizer temperature of the ELSD detector was set at 30 °C.