Rutin

The fresh fruits of sea buckthorn used in this experiment were provided and identified by Qinghai Nationalities University. They were stored in the −18 °C freezer until use.
Reference substances of rutin (purity ≥ 98%), quercetin (purity ≥ 98%), isorhamnetin (purity ≥ 98%) were purchased from Shanghai yuanye Bio-Technology Co., Ltd. (Shanghai, China). Ultrapure water was produced by Barnstead TII super Pure Water System (MA, USA). D101 macroporous resin was purchased from Tianjin Bohong Resin Technology Co., Ltd. (Tianjin, China). All other analytical grade chemicals used in this experiment were purchased from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China).
High-fat feed was purchased from SPF (Beijing, China) Biotechnology Co., Ltd. (Beijing, China). All detection kits (total cholesterol test kit, triglyceride detection kit, protein carbonyl detection kit, high-density lipoprotein cholesterol detection kit, low-density lipoprotein cholesterol detection kit, aspartate aminotransferase detection kit, alanine transaminase detection kit and Bradford protein detection kit) were purchased from Nanjing Jiancheng Bioengineering Research Institute (Nanjing, Jiangsu, China).

Chromatographically pure acetonitrile and acetic acid was purchased from Thermo Fisher (Waltham, MA, USA). Catechins (C), epicatechin (EC), gallocatechin (GC), epigallocatechin (EGC), epigallocatechin gallate (ECG), gallocatechin gallate (GCG), epigallocatechin-3-O-gallate (EGCG), procyanidin B1, procyanidin C1, kaempferitrin, rutin, isoquercetin, myricetin-3-O-galactoside, myricetin-3-O-rutinoside, galloylglucose, gallic acid, cinnamic acid, chlorogenic acid, shikimic acid, p-coumaric acid, quinic acid, pyrogallic acid, theogallin, strictinin, theanine, theobromine, ethyl gallate, tannic acid, caffeine, L-sodium glutamate, and D-glucose were purchased from Yuanye (Shanghai, China). Quercetin, myricetin, kaempferol, and kaempferol-3-O-glucoside were purchased from Shanghai Zzstandard (Shanghai, China). Amino acid mixture standard reagent, borate buffer solution, o-phthalaldehyde (OPA), and 9-fluorene methyl chloroformate (FMOC) were purchased from Agilent Technology (Santa Clara, CA, USA). n-Hexane was purchased from Thermo Fisher (Waltham, MA, USA). 2-Methyl-3-heptanone and n-alkanes (C₇–C₃₀) were purchased from Sigma (St. Louis, MO, USA).

The production of YGYSG was undertaken in The First Clinical Medical College at Anhui University of Chinese Medicine. A Milli-Q water purification system (Millipore, Bedford, MA, USA) was employed to acquire deionized water. For mass spectrometry analysis, formic acid of LC-MS grade was purchased from Sigma-Aldrich (St. Louis, MO, USA). Acetonitrile and methanol were LC-MS grade and obtained from Merck (Darmstadt, Germany). Overall, 27 chemical standards, including chikusetsusaponin IVa, chikusetsusaponin IV, ginsenoside Ro, 25R-inokosterone, β-ecdysone, acacetin, cymaroside, quercetin, gallic acid, oleanolic acid, hyperoside, chlorogenic acid, astragalin, kaempferol, cryptotanshinone, tanshinone II_A, rosmarinic acid, calycosin-7-O-β-D-glucoside, salvianolic acid B, astragaloside IV, formononetin-7-O-β-D-glycoside, calycosin, formononetin, scopoletin, ferulic acid, rutin, and narcissoside, were purchased from Shanghai Yuanye Bio-Technology Co., Ltd. (Shanghai, China), and their purity levels exceeded 98% as determined through high-performance liquid chromatography (HPLC) analysis. Figure 2 displays the chemical configurations of each reference standard.

After one week of natural drought for 3-week-old A. thaliana (WT and transgenic plants) grown in the same pot, the leaves were taken to extract anthocyanin using 1% (v/v) HCl-methanol extraction solution at 4 °C in the dark for 24 h, to determine the anthocyanin content using the pH difference method according to the previous method [57 (link)].
The apple calli treated with 6% PEG for two weeks were taken to extract flavonoids by incubating in 1% (v/v) HCl methanol for 4 h at 4 °C, to measure the flavonoid content using an ultra-sensitive multi-function microchannel plate detector (Biotek, Cytation 1, Winooski, VT, USA). Rutin (Yuanye Bio-Technology, Shanghai, China) was used as the master standard.