The mixture of methanol (HPLC grade, filtered with a 0.45mm
membrane filter) with redistilled water (80:20) was used as a
mobile phase. The program of gradient elution was set for a linear
gradient starting from 50% of methanol to reach 100% at 23 min;
the injection volume was 20mL, and the detection wavelength
254 nm. The total flow-rate was 1 mL min1 and the column
temperature was 25 C. The chromatographic data were recorded
and processed by the LC solution program version 1.23 SP.