N 2 hydroxylpiperazine n 2 ethanesulfonic acid hepes

Benzyl alcohol (≥ 98% in purity), adenosine 5’-triphosphate (ATP), 3’-phosphoadenosine-5’-phosphosulfate (PAPS), N-2-hydroxylpiperazine-N’-2-ethanesulfonic acid (HEPES), Trizma base, dithiothreitol (DTT), minimum essential medium (MEM), fetal bovine serum (FBS), penicillin G, and streptomycin sulfate were products of Sigma Chemical Company (St. Louis, MO). Ultrafree-MC 5000 NMWL filter units and cellulose thin-layer chromatography (TLC) plates were from EMD Millipore (Billerica, MA). HepG2 human hepatoma cells (ATCC HB-8065) and Caco-2 human colon adenocarcinoma cells (ATCC HTB-37) from American Type Culture Collection (Manassas, VA). Pooled human lung S9 fraction from a mixed-gender group of 4 donors (Lot No. 0710281), liver cytosol from 50 donors (Lot No. 09103970), small intestine (duodenum and jejunum) S9 fraction from 18 donors (Lot No. 0710351), and kidney S9 fraction from 8 donors (Lot No. 0510093) were obtained from XenoTech, LLC (Lenexa, KS). Ecolume scintillation cocktail and carrier-free sodium [³⁵S]sulfate were from MP Biomedicals, LLC, (Irvine, CA, USA). All other chemicals were of the highest grade commercially available.

6-OH-Mel, NAS and 4-OH-Ram were products of Toronto Research Chemicals (Toronto, Ontario, Canada). Adenosine 5’-triphosphate (ATP), 3’-phosphoadenosine 5’-phosphosulfate (PAPS), 2-morpholinoethanesulfonic acid (MES), N-2-hydroxylpiperazine-N’-2-ethanesulfonic acid (HEPES), 3-N-tris-hydroxymethyl methylamino-propanesulfonic acid (TAPS), 2-cyclohexylamino ethanesulfonic acid (CHES), 3-cyclohexylamino-1-propanesulfonic acid (CAPS), Trizma base, dithiothreitol (DTT), minimum essential medium (MEM), fetal bovine serum (FBS), penicillin G, streptomycin sulfate, and silica gel thin-layer chromatography (TLC) plates were from Sigma Chemical Company (St. Louis, MO, USA). Carrier-free sodium [³⁵S]sulfate was from American Radiolabeled Chemicals, Inc. (St. Louis, MO, USA); and Ecolume scintillation cocktail was from MP Biomedicals (Irvine, CA, USA). Ultra free-MC 5000 NMWL filter units were products of Millipore Corporation (Bedford, MA, USA). HepG2 human hepatoma cells (ATCC HB-8065) and Caco-2 human colon adenocarcinoma cells (ATCC HTB-37) were obtained from American Type Culture Collection (Manassas, VA, USA). Pooled human lung, liver, small intestine, and kidney cytosols were purchased from XenoTech, LLC (Lenexa, KS, USA). All other chemicals used were of the highest grade commercially available.

Acetaminophen (≥ 99% in purity), 4-nitrophenol (4NP) (≥ 99% in purity), dimethyl sulfoxide (DMSO), isopropyl β-D-1-thiogalactopyranoside (IPTG), adenosine 5′-triphosphate (ATP), N-2-hydroxylpiperazine-N′-2-ethanesulfonic acid (HEPES), and dithiothreitol (DTT) were products of Sigma-Aldrich (St. Louis, MO, USA). Tapentadol (≥ 98% in purity) was purchased from Cayman Chemical Company (Ann Arbor, MI, USA). O-DMN (≥ 98% in purity) was a product of Toronto Research Chemicals (Toronto, ON, Canada). Carrier-free sodium [³⁵S] sulfate was from American Radiolabeled Chemicals (St. Louis, MO, USA). Recombinant human bifunctional PAPS synthase was prepared as described previously [27 (link)]. Polygram® Cellulose 300 thin-layer chromatography (TLC) plates were from Macherey-Nagel GmbH and Co. KG (Düren, Germany). PrimeSTAR® Max DNA polymerase was a product of Takara Bio (Mountain View, CA, USA). Protein molecular weight markers were from Bioland Scientific LLC (Paramount, CA, USA). Ecolume scintillation cocktail was purchased from MP Biomedicals, LLC (Irvine, CA, USA). X-Ray films were products of FUJIFILM Corporation (Tokyo, Japan). Oligonucleotide primers were custom synthesized by Eurofins Genomics (Louisville, KY, USA). QIAprep® Spin Miniprep Kit was a product of QIAGEN (Düren, Germany). All other chemicals were of the highest standards commercially available.