Cyanidin 3 o galactoside

The 26 Chinese prickly ash peels materials were collected from eight provinces of China (the main production areas: Guide, Xunhua, Hanyuan, Jiuzhaigou, Wenxian, Wudu, Qin’an, Fengxian, Fuping, Hengshan, Hancheng, Yongji, Lingbao, Jiaocheng, Shexian, Zaozhaung and Laiwu) at different altitudes (201–2188 m) from July to August 2020 (Figure 1). The 26 samples were divided into 6 groups based on the geographical location and altitude of the sample collection sites, as shown in Table S1. Under the premise of protecting local germplasm resources, Chinese prickly ash mature fruits from each location were collected in three replicates, and the distance between each plant was more than 50 m. Fruits without pests and mechanical damage were dried in the laboratory at room temperature until they reached a constant weight.
A total of 15 flavonoid compound standards (hyperoside, luteolin, kaempferol, quercitrin, catechin, rutin, chlorogenic acid, quercetin, hesperetin, apigenin, peonidin O-hexoside, peonidin 3-O-glucoside, cyanidin 3-O-glucoside, cyanidin O-syringic acid, cyanidin 3-O-galactoside) were bought (Beijing Solarbio Science & Technology, Beijing, China). HPLC-grade methanol, acetic acid and acetonitrile were bought (TEDIA Chemical Co., Ltd., Fairfield, OH, USA). Deionized water (18 MΩ cm) was used to prepare aqueous solutions (MilH-Q Advantage A1, Millipore, Billerica, MA, USA).

The samples for anthocyanin determination were finely ground in liquid nitrogen. About 1 g of powder was added to 5 mL of 1% HCl–methanol, leached overnight at 4 °C in the dark and then centrifuged; the sediment was washed twice with 5 mL of 1% HCl–methanol, and the supernatants were combined. The supernatant volume was adjusted to 20 mL, filtered through a microporous membrane (diameter 13 mm, pore size 0.22 μm, Advantec, CA, USA). Anthocyanins were separated in an HPLC system (Agilent 1220, Waldbronn, Germany). A Waters Symmetry C18 column (5 μm, 4.6 mm × 150 mm) was used; the loading volume was 20 μL, mobile phase A was 10% formic acid, mobile phase B was acetonitrile. The linear gradient elution design was: 0–13 min – acetonitrile 0–20%, 20 min – acetonitrile 40%, 25 min – acetonitrile 0%; column temperature was 25 °C, flow rate was 1 mL/min, and detection wavelength was 520 nm. A standard curve was prepared using cyanidin 3-O-galactoside (Beijing Solarbio Science & Technology Co. Ltd., Beijing, China).