Silica gel thin layer chromatography

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.

Morphine, hydromorphone, oxymorphone, butorphanol, levorphanol, nalbuphine, nalorphine, naltrexone, 6β-naltrexol, buprenorphine, pentazocine, and naloxone, were purchased from Cerilliant. Adenosine 5’-triphosphate (ATP), sodium dodecyl sulfate (SDS), dithiothreitol (DTT), dimethyl sulfoxide (DMSO), 3-(N-morpholino)propanesulfonic acid (Mops), Trizma base (Tris), isopropyl-β-D-thiogalactopyranoside (IPTG), and silica gel thin-layer chromatography (TLC) plates were from Sigma Chemical Company. Polyethyleneimine (PEI) cellulose TLC plates were from Brinkmann. Ecolume scintillation cocktail was purchased from MP Biomedical. Carrier-free sodium [³⁵S]sulfate was from PerkinElmer. HepG2 human hepatoma cell line (ATCC HB-8065) was obtained from American Type Culture Collection. Pooled human lung S9 fraction, liver cytosol, small intestine (duodenum and jejunum) S9 fraction, and kidney S9 fraction were purchased from XenoTech, LLC. All other chemicals were of the highest grade commercially available.