Daidzin

Gallic acid, p-hydroxybenzoic acid, ferulic acid, trans-cinnamic acid, p-coumaric acid, benzoic acid, ellagic acid, protocatechuic acid, sinapic acid, caffeic acid, quercetin, rutin, taxifolin, isoquercitrin, catechin, gallocatechin, epicatechin, vitexin, genistin, daidzin, phloroglucinol, quinic acid, and 3,4-dihdyroxyphenylaceticacid were obtained from Yuanye Bio-Technology Co., Ltd. (Shanghai, China). Procyanidin B1, procyanidin B2, and kaempferol-3-O-rutinoside were obtained from Chengdu Pusi Biotechnology Co., Ltd. (Chengdu, China). 2,20-azinobis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,4,6tri(pyridin-2-yl)-1,3,5-triazine (TPTZ), (±)-6-Hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid (Trolox), fluorescein, 3′,6′-dihydroxyspiro [isobenzofuran-1 (3 H),9′-(9 H) xanthene]-3-one (FL), 2,2′-azobis (2-amidinopropane) dihydrochloride (AAPH), and EDTA-2Na were purchased from Aladdin Biochemical Technology Co., Ltd. (Shanghai, China). Thermo Fisher Scientific (Waltham, MA, USA) provided the methanol (HPLC quality), formic acid (99.9%, LC-MS grade), and acetonitrile (LC-MS grade). Cellulase (T richoderma Vride G, 400 u/mg), hemicellulase (Aspergillus niger, 20 u/mg), and pectinase (Aspergillus niger, 500 u/mg) were obtained from Yuanye Bio-Technology Co., Ltd. (Shanghai, China).

An SI mixture containing 51.81% daidzin, 19% glycitin, 7.94% genistin, 2.45% daidzein, 0.81% glycitein, and 0.52% genistein, and their standards, were obtained from Shanghai Yuanye Bio-Technology Co., Ltd. (Shanghai, China) [15 (link)]. Penicillin/streptomycin, fetal bovine serum (FBS), and Dulbecco’s modified Eagle medium (DMEM) were purchased from Gibco (CA, USA). PC12 cell lines were purchased from Nanjing Keygen Biotech. Co., Ltd. (Nanjing, China). The kits for ROS, apoptosis, the cell cycle, and the protein content were obtained from the Shanghai Beyotime Institute of Biotechnology (Shanghai, China). Antibodies for β-tubulin, Bax, cyclin A, p21, Bcl-xL, hypoxia-inducible factor-1 alpha (HIF-1α), LC3, p62, Nrf2, p-Akt, Akt, p-mTOR, mTOR, p-p38 MAPK, p38 MAPK, and LamB were obtained from Shanghai Abmart Inc. (Shanghai, China).

Ferulic acid (98% purity), liquiritin (98% purity), magnolol (98% purity), and ginsenoside Rb2 (93.8% purity) were obtained from the National Institutes for Food and Drug Control (NIFDC, Beijing, China). Borneol (98% purity), ginsenoside Rc (98% purity), artemisinin (99% purity), chenodeoxycholic acid (98% purity), daidzin (98% purity), and bacopaside I (98% purity) were purchased from Shanghai yuanye Bio-Technology Co., Ltd. Other reagents were of analytical purity.

An Ultimate 3000 Ultra High Performance Liquid Chromatography (UPLC) system coupled with LTQ-Orbitrap velos pro mass spectrometer was used (Thermo-Fisher, Waltham, MA, USA), as well as an Acquity UPLC I-Class UPLC system coupled with Xevo TQ-S micro triple quadrupole mass spectrometer (Waters, Milford, MA, USA).
The reference compounds of formononetin, ononin, daidzein, daidzin, genistin, genistein, calycosin-7-O-β-d-glucoside, biochanin A, and chlorogenic acid (purity > 98% for all) were all purchased from Shanghai Yuanye Biotechnology Co., Ltd. (Shanghai, China). Acetonitrile and formic acid were purchased from Fisher Scientific, San Jose, CA, USA Company (mass spec-grade). Deionized water was prepared by Milli-Q ultrapure water preparation system (Millipore Co., Ltd., Billerica, MA, USA). All other chemicals were purchased from Beijing Chemical Works (Beijing, China, analytical grade).

Reference standards of puerarin, 3′-methoxypuerarin, daidzin and daidzein (purity ≥ 98%) were supplied by Shanghai Yuanye Biotechnology Co., Ltd, China. Methanol and formic acid (Analytical Grade, Tianjin, China) were purchased from Tianjin Kermel Chemical Reagent Co., Ltd. Acetonitrile (Tedia Company Inc., Fairfield, USA) was used for HPLC analysis. Deionized water (>18 MΩ cm) was prepared using an Elga Purelab water system (Elga, England).
Dry Puerariae lobata was purchased from the Zhonglu Hospital. The roots of Puerariae lobata were washed, dried and ground in a grinder (Zhongxing Weiye, Beijing, China). The powder was passed through a 60 mesh sieve and stored at −4 °C.