3h estrone 3 sulfate

TSA was supplied by Errant Gene Therapeutics (United States of America). The radiolabeled compounds ([3H] taurocholic acid (specific activity 5 Ci/mmol), [3H] estrone-3-sulfate (specific activity 45.6 Ci/mmol), scintillation vials and scintillation fluid (Ultima Gold MV) were purchased from Perkin Elmer (Belgium). All other chemicals were commercially available products of analytical grade and were obtained from Sigma (Belgium), unless specified otherwise. Probenecid and TSA were dissolved in dimethylsulfoxide (DMSO), with final DMSO concentrations not exceeding 1 % v/v.

The active ingredient of MT921, cholic acid (CA), was synthesized from Medytox Inc. (Suwon, Korea). Cyclosporine A, diclofenac, bromsulphthalein, deoxycholate, chenodeoxycholic acid, and Dulbecco’s Phosphate Buffered Saline (DPBS) were purchased from Sigma-Aldrich (St. Louis, MO, USA). [³H]estrone-3-sulfate (45 Ci/mmol), [³H]estradiol-17β-d-glucuronide (34.3 Ci/mmol), and [³H]taurocholate (5 Ci/mmol) were purchased from Perkin Elmer (Boston, MA, USA). Fetal bovine serum (FBS), non-essential amino acids, penicillin, and streptomycin were purchased from Gibco BRL, Life Technologies (Grand Island, NY, USA). Dulbecco’s Modified Eagle’s Medium (DMEM), poly-d-lysine coated 24-well plates, and poly-d-lysine coated 96-well plates were purchased from Corning-Gentest (Tewksbury, MA, USA). The acetonitrile used was analytical grade and purchased from Merck (Darmstadt, Germany).
The HEK 293-OATP1B1, -OATP1B3, -OAT3, -NTCP, and -ASBT stable cells and mock cells were purchased from Corning Life Science (Woburn, MA, USA).

³H-Estrone-3-sulfate (48 Ci/mmol) and ³H-dehydroepiandrosterone-sulfate (60 Ci/mmol) were obtained from Perkin Elmer (Waltham, MA, US). The fluorescent dye (E)-1-(3-azidopropyl)-4-(2-(7-(diethylamino)-2-oxo-2H-chromen-3-yl)vinyl)pyridine-1-ium-3-sulfonate (ACPS) was synthetized, as described previously (Nagy et al., 2010 (link)). Tissue total RNAs were obtained from Agilent Technologies (Kromat, Hungary). Estrone-3-sulfate, benzbromarone, and all other materials, if not stated otherwise, were purchased from Sigma-Aldrich (Sigma, Merck, Darmstadt, Germany).

[¹⁴C]Tetraethylammonium (TEA, 55 mCi/mmol) and [³H]1-methyl-4-phenylpyridinium (MPP⁺, 85 Ci/mmol) were obtained from American Radiolabeled Chemicals (St. Louis, MO, USA), [³H]atenolol (3.3 Ci/mmol) was from Moravek Biochemicals (Mercury Lane, Brea, CA, USA), and [³H]estrone-3-sulfate (54 Ci/mmol) was from PerkinElmer (Waltham, MA, USA). Unlabeled atenolol and Dulbecco’s modified Eagle’s medium (DMEM) were from Wako Pure Chemical Industries (Osaka, Japan). Fetal bovine serum (FBS) was obtained from Invitrogen (Carlsbad, CA, USA). All other regents were of analytical grade and commercially obtained.

RGE was purchased from the Punggi Ginseng Cooperative Association (Youngjoo, Kyungpook, Korea). The ginsenosides Rb1, Rb2, Rc, Rd, Rg1, Rg3, Rh1, Rh2, compound K, Re, PPD, and PPT were purchased from the Ambo Institute (Daejeon, Korea). Berberine, caffeine, valsartan, probenecid, rifampin, TEA, sodium dodecyl sulfate (SDS), and Hank’s balanced salt solution (HBSS) were obtained from Sigma-Aldrich (St. Louis, MO, USA). Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS), non-essential amino acids, and poly-D-lysine-coated 96-well plates were purchased from Corning Life Sciences (Woburn, MA, USA). [³H]Methyl-4-phenylpyridinium ([³H]MPP⁺; 2.9 TBq/mmol), [³H]para-aminohippuric acid ([³H]PAH; 0.13 TBq/mmol), [³H]estrone-3-sulfate ([³H]ES; 2.1 TBq/mmol), and [³H]estradiol-17β-d-glucuronide ([³H]EG; 2.2 TBq/mmol) were purchased from Perkin Elmer Inc. (Boston, MA, USA). Acetonitrile, methanol, and water were obtained from Fisher Scientific Co. (Fair Lawn, NJ, USA). All other chemicals and solvents were of reagent or analytical grade.

7-Cl-KYNA was obtained from
Abcam and dissolved in a small volume of 1 M HCl, and the pH was then
titrated to 7.4 in PBS. 4-Cl-KYN and ac-4-Cl-KYN were kind gifts from
Vistagen Therapeutics, Inc. All other compounds were purchased from
Sigma-Aldrich and solubilized in dimethyl sulfoxide according to the
manufacturer’s instructions.
[¹⁴C]-4-Cl-KYN
(specific activity = 55.5 mCi/mmol) and [¹⁴C]-7-Cl-KYNA
(specific activity = 77.7 mCi/mmol) were purchased from Moravek Inc.
[¹⁴C]-Tetraethylammonium specific activity = 3.5 mCi/mmol)
and [³H]-estrone-3-sulfate specific activity = 49.19 Ci/mmol)
were purchased from PerkinElmer. [³H]-Phenylalanine (specific
activity = 100 Ci/mmol), [³H]-para-aminohippuric acid (specific
activity = 40 Ci/mmol), and [³H]-cyclic guanosine monophosphate
(specific activity = 25 Ci/mmol) were obtained from American Radiolabeled
Chemical Inc. Finally, [³H]-l-DOPA (specific activity
4 Ci/mmol) was acquired from American Radiolabeled Chemicals, Inc.

All reagents were purchased from commercial suppliers (Wako Pure Chemical Industries, Osaka, Japan; Sigma-Aldrich, Japan, Tokyo, Japan; and Nacalai Tesque, Kyoto, Japan) unless indicated otherwise.
In all experiments, ³H- or ¹⁴C-labeled tracers were used because their long half-lives (12.3 and 5700 years, respectively) make them more suitable for in vitro experiments than ¹⁸F (110 min). trans-1-Amino-3-fluoro[1-¹⁴C]cyclobutanecarboxylic acid ([¹⁴C]fluciclovine, 2.09 GBq/mmol) was synthesized [9 (link)] by Nemoto Science (Tokyo, Japan). l-[¹⁴C]alanine (5.9 GBq/mmol, for system A and ASC AATs); l-[¹⁴C]leucine (12.0 GBq/mmol, for system L AATs); d-[1-³H(N)]mannitol (455.1 GBq/mmol, for the evaluation of paracellular transport on LLC-PK1 monolayer experiment); [³H]digoxin (1104 GBq/mmol, for P-gp); [³H]estrone-3-sulfate ([³H]ES; 1655 GBq/mmol, for BCRP and OAT3); and p-[¹⁴C]aminohippurate ([¹⁴C]PAH; 2.04 GBq/mmol, for OAT1) were purchased from PerkinElmer (Waltham, MA, USA). [³H]estradiol-17β-d-glucuronide ([³H]E2G; 1269 GBq/mmol, for MRP4, OATP1B1, and OATP1B3) and [¹⁴C]metformin (3.7 GBq/mmol, for OCT2) were purchased from American Radiolabeled Chemicals (St. Louis, MO, USA).