Astragaloside 4

Radix Astragali was collected from Longxi, Gansu Province in China. After being cleaned and dried in a heating-air drying oven at about 40 °C until constant weight, the raw materials were smashed and sieved through a 6-mesh sieve. The powder was sealed in a PE bag and then kept at −20 °C for later use. Astragaloside I, astragaloside II, and astragaloside IV standard compounds were purchased from Shanghai yuan ye Bio-Technology Co., Ltd. (Shanghai, China) with a purity of >98%. Methanol (HPLC grade), acetonitrile (HPLC grade), methanoic acid (HPLC grade), ammonium hydroxide, and ethanol were sourced from Sinopharm Chemical Reagent Co., Ltd. (Shanghai, China). Ultrapure water was used for the preparation of all solutions.

Homogenized liver microsomes of humans and mice were purchased from the Research Institute for Liver Diseases Co., Ltd. (Shanghai, China). As an internal standard (IS), astragaloside IV was obtained from Yuanye Biotech Co., Ltd. (Shanghai, China). Authentic compounds, such as 7-ketolithocholic acid, UDCA, chenodeoxycholic acid (CDCA), ursocholic acid (UCA), β-muricholic acid (β-MCA), and UDCA-d₄, were commercially supplied by Yuanye Biotech Co., Ltd. (Shanghai, China). UDCA-3-sulfate (UDCA-3-S) was previously biosynthesized by SULT2A1 in our laboratory.
LC–MS grade acetonitrile (ACN), methanol, ammonium formate, as well as formic acid were obtained from Thermo-Fisher (Pittsburgh, PA, USA). Ultrapure deionized water was prepared using an in-house Millipore Milli-Q Integral water purification system (Bedford, MA, USA). Nicotinamide adenine dinucleotide phosphate (NADPH), glucose 6-phosphate (G6P), glucose-6-phosphate dehydrogenase (G6PD), uridine 5′-diphosphate glucuronic acid (UDPGA), alamethicin, dithiothreitol (DTT), D-saccharic acid-1,4-lactone monohydrate, 3′-phosphoadenosine-5′-phosphosulfate (PAPS), magnesium chloride, tris hydrochloride (Tris HCl), phosphate-buffered saline (PBS) were purchased from Yuanye Biotech Co., Ltd. (Shanghai, China).