7 hydroxycoumarin

Kurarinone (>98%) was purchased from BioBioPha Co., Ltd. (Kunming, China). Glucose-6-phosphate, Glucose-6-phosphate dehydrogenase, NADP⁺, D-Glucose-6-phosphate, Tris-HCl, 7-hydroxycoumarin, 4-methylumbelliferone (4-MU), 4-methylumbelliferone-β-D-glucuronide, uridine 5′-diphosphoglucuronic acid (UDPGA; trisodium salt), phenacetin, diclofenac, dextromethorphan, chlorzoxazone, testosterone, (S)-mephenytoin, sulfaphenazole, quinidine, clomethiazole, furafylline, ketoconazole, and omeprazole were purchased from Sigma Aldrich (St. Louis, MO, USA). Metabolites of the probe substrates including 6-hydroxylated chlorzoxazone (2E1), O-deethylated phenacetin (CYP1A2), 4′-hydroxylated diclofenac (2C9), 4′-hydroxylated (S)-mephenytoin (2C19), O-demethylated dextromethorphan (2D6), and 6β-hydroxylated testosterone (3A4) were provided from the Research Institute for Liver Disease Co., Ltd. (Shanghai, China). Pooled liver microsomes from humans (HLMs), monkeys (MLMs), rabbits (RAMs), rats (RLMs), mouse (MOMs), dogs (DLMs), and minipigs (PLMs) were provided by the Research Institute for Liver Disease Co., Ltd. (Shanghai, China). Recombinant human supersomes (UGT1A1, UGT1A3, UGT1A6, UGT1A7, UGT1A8, UGT1A9, UGT1A10, UGT2B4, UGT2B7, UGT2B15, CYP1A2, CYP2C9, CYP2D6, CYP2E1, CYP3A4, and CYP2C19) were obtained from BD Gentest Corp. (Woburn, MA, USA).

Lysionotin (≥98%) and testosterone (≥98%) were obtained from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). The chemical structure of lysionotin is shown in Figure 1. d-Glucose-6-phosphate, glucose-6-phosphate dehydrogenase, corticosterone (≥98%), NADP⁺, phenacetin (≥98%), acetaminophen (≥98%), 4-hydroxymephenytoin (≥98%), 7-hydroxyCoumarin (≥98%), 4′-hydroxydiclofenac (≥98%), sulfaphenazole (≥98%), quinidine (≥98%), tranylcypromine (≥98%), chlorzoxazone (≥98%), 6-hydroxychlorzoxazone (≥98%), paclitaxel (≥98%), 6β-hydroxytestosterone (≥98%), clomethiazole (≥98%), and furafylline (≥98%) were obtained from Sigma Chemical Co. Montelukast (≥98%) was obtained from Beijing Aleznova Pharmaceutical (Beijing, China). Coumarin (≥98%), diclofenac (≥98%), dextromethorphan (≥98%), and ketoconazole (≥98%) were purchased from ICN Biomedicals. Pooled HLMs were purchased from BD Biosciences Discovery Labware. All other reagents and solvents were of analytical reagent grade.

7-Hydroxy-coumarin (Umbelliferone, Co., Ltd., USA) was purchased from Sigma-Aldrich Co., Ltd. Epichlorohydrin (AR), potassium hydroxide (AR), sodium hydroxide (AR), chloroform (AR), anhydrous sodium sulfate (AR) and N,N-dimethylformamide (DMF, AR) were purchased from Sinopharm Chemical Reagent Co., Ltd (China). HA (Mn = 100 kDa) was purchased from Zhenjiang Dongyuan Biotech Co., Ltd. (China). Methacrylic anhydride (MA) was purchased from Aladdin Chemical Co., Ltd. LPS (from Escherichia coli O111: B4) was purchased from Sigma-Aldrich Co., Ltd. IFN-γ was purchased from PROSPEC Co., Ltd. Sprague–Dawley rats (120 g) were bought from Zhejiang Academy of Medical Science. Recombinant human macrophage colony stimulating factor (MCSF) was purchased from Peprotech PeproTech, Inc. The transwell molds and 24-well culture plates were bought from Corning Co., Ltd., USA. Fetal bovine serum (FBS) was purchased from Sijiqing Inc., China.

Cytochrome c (cyt c) (horse heart), glucose-6-phosphate (G6P), glucose-6-phosphate dehydrogenase (G6PD), nicotinamide-adenine dinucleotide phosphate (NADPH), ethoxyresorufin, methoxyresorufin, resorufin, coumarin, 7-hydroxy coumarin, 3-cyano-7-ethoxycoumarin, 3-cyano-7-hydroxycoumarin, fluorescein, trypsin, penicillin-streptomycin (10,000 units penicillin and 10 mg streptomycin per mL), Dulbecco’s Modified Eagle’s Medium-low glucose (DMEM), fetal bovine serum (FBS), phosphate buffered saline pH 7.4 (PBS) and doxorubicin were obtained from Sigma Aldrich (St. Louis, MO, USA). Nicotinamide adenine dinucleotide phosphate (NADP+) was obtained from Gerbu (Heidelberg, Germany). Bradford reagent was obtained from Bio-Rad (Hercules, CA, USA) and Quiazol from Qiagen (Hilden, Germany). Dibenzylfluorescein, sodium dithionite, dimethyl sulfoxide (DMSO), acetonitrile (ACN) and sodium hydrogencarbonate (NaHCO₃) were purchased from Merck (Kenilworth, NJ, USA). Insulin was obtained from Cell Applications, Inc. (San Diego, CA, USA). All other chemicals and solvents were of the highest grade commercially available.

EAM-2201 was obtained from Cayman Chemical Company (Ann Arbor, MI, USA). Acetaminophen, N-acetylserotonin, alamethicin, chenodeoxycholic acid, coumarin, diclofenac, dimethyl sulfoxide (DMSO), 7-hydroxycoumarin, midazolam, mycophenolic acid, naloxone, naloxone 3-β-d-glucuronide, NADPH, phenacetin, trifluoperazine, Trizma base and uridine 5′-diphosphoglucuronic acid (UDPGA) were purchased from Sigma-Aldrich (St. Louis, MO, USA). ¹³C₂,¹⁵N-Acetaminophen, bufuralol, N-desethylamodiaquine, 1′-hydroxybufuralol, d₉-1-hydroxybufuralol, 4-hydroxydiclofenac, 4-hydroxymephenytoin, 1′-hydroxymidazolam, [S]-mephenytoin and ultrapooled human liver microsomes (150 donors) were purchased from Corning Life Sciences (Woburn, MA, USA). N-Acetylserotonin β-d-glucuronide, chenodeoxycholic acid 24-acyl-β-glucuronide, mycophenolic acid β-d-glucuronide and SN-38 glucuronide were obtained from Toronto Research Chemicals (Toronto, ON, Canada). SN-38 was obtained from Santa Cruz Biotechnology (Dallas, TX, USA). Acetonitrile, methanol and water were of liquid chromatography–mass spectrometry (LC-MS) grade and purchased from Fisher Scientific Co. (Fair Lawn, NJ, USA). All other chemicals were of the highest quality available.

E. supina was purchased in mid-April 2012 from a market in Jinju, South Korea. The plant was authenticated by Professor Moo Ryong Huh, a plant taxonomist with the Research Institute of Agricultural Life Science, Gyeongsang National University. A voucher plant was deposited in the herbarium at this institute. The plant was washed with water, lyophilized, and stored in dark containers at −70°C until needed. All chemicals were purchased from Sigma-Aldrich Co., LLC (St. Louis, MO, USA). Gallic acid, protocatechuic acid, 7-hydroxycoumarin, quercetin 3-O-glucoside, and kaempferol, which were purchased from Sigma-Aldrich Co., LLC (St. Louis, MO, USA), were used as external standards after recrystallization in ethanol. The purity of all standards was confirmed to be >99% by using HPLC. All solvents and water were obtained from Duksan Pure Chemicals Co., Ltd. (Ansan, Republic of Korea).

Synthesis and characterization of the coumarin derivatives were performed as described previously [40 (link)]. Briefly, series of coumarin derivatives were synthesized via Knoevenagel condensation starting from various substituted salicylaldehydes and ethyl acetoacetate (series 1; compounds 1–7), ethyl benzoylacetate (series 2; compounds 8–15), ethyl cyanoacetate (series 3; compounds 16–23), diethyl malonate (series 4; compounds 24–31), and dimethyl malonate (series 5, compounds 32–38). 7-hydroxycoumarin and coumarin (compounds 39 and 40 respectively) were purchased from Sigma Chemical Co. (St. Louis, MO, USA). The structures of the tested compounds are presented in Table 1.