3.4. Sample Extraction and Preparation

The procedure of extraction of total flavonoids, as described in the literature, has been modified [45]. The fresh flower of P. ostii was collected, and the peduncle, receptacles, and calyx were removed from the large flower, leaving only the petals of the flower. The 30 Kg petals were powered, and extracted thrice, each time with 20 L of ethanol at room temperature for 48 h. The combined ethanol extract was filtered and evaporated at 60 °C under reduced pressure using a vacuum rotary evaporator (Yuhua RE-2000A, Zhengzhou, China). The weight of the crude extract of this flower was 93.5 g.

The crude extract (90 g) was suspended in distilled water and subsequently filtered using filter papers. The filtrates were loaded onto the polyamide column (inner diameter 15 cm, length 120 cm, 8000 mL) for total flavonoid enrichment at a flow rate of 4 BV/h. After the polyamide resin was saturated with flavonoid compounds, the resin column was washed with distilled water and 20%, 30%, 40%, 50%, 60%, 70%, 80%, and 90% ethanol (v/v) with isocratic modes at a flow rate of 4.0 BV/h. The content of total flavonoids in each desorbed fraction was analyzed by UV-2600 (Ultraviolet-visible 2600) spectrophotometer. The desorbed fractions were concentrated in an evaporator at 60 °C under vacuum. The dried products of each fraction were weighed, and the content and recovery yields of total flavonoids were calculated.