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31 protocols using d glucose 6 phosphate

1

Synthesis and Characterization of N-Sulfonylhydrazone Prototypes

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N-sulfonylhydrazone
prototypes, LASSBio-1771 (1), LASSBio-1772 (2), LASSBio-1773 (3), LASSBio-1774 (4),
and the internal standard (IP), biphenyl l-4-carboxylate methyl, were
synthesized at the Laboratório de Avaliação
e Síntese de Substâncias Bioativas
(LASSBio,
UFRJ, Brazil) with ≥98% pure by HPLC (see the Supplemental Data). HPLC grade solvents and reagents, i.e., acetonitrile, methanol, and formic acid (96%), were
purchased from Tedia-Brazil. Type I grade water produced by an ultrapure
water purification system (Master System MS2000, Gehaka, Brazil) was
used in experiments and HPLC analysis. Bis(p-nitrophenyl)
phosphate sodium salt, D-glucose-6-phosphate, glucose-6-phosphate
dehydrogenase, β-nicotinamide adenine dinucleotide phosphate
(NADP+), magnesium chloride hexahydrate, quinidine, sulfaphenazole,
furafylline, p-nitrophenol, ketoconazole, bicinchoninic
acid (BCA) protein assay kit, and salts used for buffer preparation
were purchased from Sigma-Aldrich (St. Louis, MO).
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2

Lysionotin and Testosterone Characterization

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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.
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3

Xanthotoxol Cytochrome P450 Inhibition

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Xanthotoxol (purity > 98%) was purchased from Sichuan Weikeqi Biotechnology Co. Ltd. (Sichuan, China). Paclitaxel, 1-aminobenzotriazole (ABT), phenacetin, sulfaphenazole, chlorzoxazone, quinidine, clomethiazole, furafylline, 8-methoxypsoralen, coumarin, diclofenac, quercetin, dextromethorphan, ketoconazole, testosterone, S-mephenytoin, omeprazole, glucose-6-phosphate dehydrogenase, NADP+, and D-glucose-6-phosphate were obtained from Sigma-Aldrich (St. Louis, MO, USA). All other reagents were the highest purity commercially available or HPLC grade.
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4

Inhibition of Human Liver Enzymes

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Anemarsaponin BII (≥98%) and testosterone (≥98%) were purchased from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). 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.
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5

Human Liver Microsome Characterization

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BFT and BF were purchased from Shanghai Winherb Medical Technology Company (Shanghai, China). ABT, furafylline, sulfaphenazole, clomethiazole, omeprazole, 8-methoxypsoralen, ticlopidine, CYP3cide, glucose-6-phosphate dehydrogenase, D-glucose-6-phosphate, and NADP+ were obtained from Sigma (St. Louis, MO, United States). Montelukast, quinidine, ketoconazole was purchased from Jianglai Biotechnology Co., Ltd. (Shanghai, China). The pooled HLMs (from 50 donors, lot no. X008067) were obtained from BioreclamationIVT (Baltimore, MD, United States). A panel of baculovirus expressed human P450s (CYP1A1, 1A2, 2A6, 2A13, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, 3A4, 3A5, 4F2, and 4F3), co-expressing NADPH-CYP reductase and cytochrome b5 were obtained from BD Gentest Corp (Woburn, MA, United States). All chemicals and solvents were of analytical grade.
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6

Peucedanol Metabolism Assay Protocol

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Peucedanol (≥98%, Figure 1) and testosterone (≥98%) were purchased from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). 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.
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7

Vonoprazan Metabolism in Hepatic Microsomes

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Both vonoprazan acetate (purity 99.5%) and vonoprazan fumarate (purity 99.7%) and their metabolite M-I (purity 98.44%) were provided by Jiangsu Carefree Pharmaceutical Co. Ltd. Research Institute (Nanjing, China). The internal standard repaglinide (purity 99.7%) was purchased from National Institutes for Food and Drug Control (Beijing, China). Dog (male beagle dog, Lot. UKHU, mixed from 3 dogs) and human (male Mongolia human, Co. Ltd. SHQY, mixed from 11 individuals) hepatic microsomes were purchased from Research Institute of Liver Diseases (Shanghai, China). D-glucose-6-phosphate, glucose-6-phosphate dehydrogenase and β-NADP+ were purchased from Sigma-Aldrich (St. Louis, Mo, USA). Other chemicals and reagents were of analytical grade or better.
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8

Characterization of Cornin and Testosterone

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Cornin (≥ 98%, Fig. 1) and testosterone (≥ 98%) were purchased from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). D-glucose-6-phosphate, glucose-6-phosphate dehydrogenase, 4′-hydroxydiclofenac (≥ 98%), 4-hydroxymephenytoin (≥ 98%),NADP+, phenacetin (≥ 98%), acetaminophen (≥ 98%), 6β-hydroxytestosterone (≥ 98%), corticosterone (≥ 98%), 7-hydroxyCoumarin (≥ 98%), sulfaphenazole (≥ 98%), tranylcypromine (≥ 98%), chlorzoxazone (≥ 98%), quinidine (≥ 98%), 6-hydroxychlorzoxazone (≥ 98%), paclitaxel (≥ 98%), clomethiazole (≥ 98%), and furafylline (≥ 98%) were obtained from Sigma Chemical Co. Montelukast (≥ 98%) was obtained from Beijing Aleznova Pharmaceutical (Beijing, China). Coumarin (≥ 98%), dextromethorphan (≥ 98%), diclofenac (≥ 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.

The chemical structure of cornin

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9

Antibiotic Sensitivity Testing of E. coli

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Kirby Bauer disc diffusion antibiotic sensitivity testing was performed on Mueller Hinton II Agar (BD #221275) or BBL™ Columbia Agar with 5% Sheep Blood (BD #221165). An overnight overnight-grown saturated E. coli (DH5α) liquid culture was diluted (1:50) in water and streaked on the agar plate to form a bacterial lawn. d-Glucose-6-phosphate (Sigma #G7250) was made in 100 mM stock solution and added to the diluted bacteria to achieve 2 mM final concentration right before streaking the plate. A 7 mm Whatman™ 3MM Chr Chromatography filter paper disc (Whatman # 3030-6189) was imbued with 7 µL of an aqueous solution of inhibitor, placed on the streaked agar plate and incubated overnight at 37 °C.
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10

HPLC Analysis of Simvastatin and AC

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Standards of AC (purity >98%) and simvastatin (purity >98%) was purchased from the National Institute for the Control of Pharmaceutical and Biological Products (Beijing, China). d-Glucose-6-phosphate, glucose-6-phosphate dehydrogenase and NADP+ were obtained from Sigma Chemical Co. (St. Louis, MO, USA). Pooled RLM were purchased from BD Biosciences Discovery Labware (Woburn, MA, USA). All other reagents and solvents were of analytical reagent grade. DST tablets were purchased from Guangdong Baiyunshan Pharmaceutical Co., Ltd (Guangzhou, China). Acetonitrile and methanol were purchased from Fisher Scientific (Fair Lawn, NJ, USA). Formic acid was purchased from Anaqua Chemicals Supply Inc. Limited (Houston, TX, USA). Ultrapure water was prepared with a Milli-Q water purification system (Millipore, Billerica, MA, USA). All other chemicals were of analytical grade or better.
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