Pooled human liver microsomes from 150 donors

Methadone hydrochloride (≥ 98%), EDDP perchlorate (≥ 98%), EMDP hydrochloride solution (1.0 mg/ml), Tris (hydroxymethyl) aminomethane (Trizma^® base), ammonium formate and fibronectin were purchased from Sigma-Aldrich (Zwijndrecht, The Netherlands). Methadone and metabolites were ordered under the opium exemption license number 104783 03 WCO, which is registered at Farmatec (executive organization of the Ministry of Health, Welfare and Sport, The Hague, The Netherlands). Dimethyl sulfoxide (DMSO, 99.7%) was obtained from Merck (Schiphol-Rijk, The Netherlands). Phosphate-buffered saline (PBS) was purchased from Gibco (Paisley, Scotland, UK). Acetonitrile (UPLC/MS grade) was obtained from Biosolve BV (Valkenswaard, The Netherlands). hiPSC-CM (Pluricyte^® Cardiomyocytes, cat# PCMI-1031-1, lot# 60151) and Pluricyte^® Cardiomyocyte medium were obtained from Ncardia (Leiden, The Netherlands). Pooled human liver microsomes (from 150 donors), pooled human intestinal microsomes (from 7 donors) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) regenerating system solution A and solution B were purchased from Corning (Woburn, MA, USA). Pooled human plasma and rapid equilibrium dialysis (RED) materials, including RED inserts, RED base plates and sealing tape were obtained from Thermo Fisher Scientific (Bleiswijk, The Netherlands).

Pooled human liver microsomes from 150 donors (75 males; 75 females) were purchased from Corning Life Sciences (Woburn, MA, USA), and (‒)-sophoranone (99.7% purity; SPN) was supplied by SK Chemicals Ltd. (Sungnam, Gyeonggi-do, Korea). β-Nicotinamide adenine dinucleotide phosphate disodium salt (NADP), glucose 6-phosphate disodium salt hydrate, glucose 6-phosphate dehydrogenase, MgCl₂, and all chemicals including the specific substrates, its metabolites, and well-known inhibitors of nine P450s were purchased from Sigma–Aldrich Corporation (St. Louis, MO, USA), Santa Cruz Biotechnology (Dallas, TX, USA), or Cayman Chemicals (Ann Arbor, MI, USA) unless stated otherwise. The purity of all purchased compounds was higher than 97.0%. HPLC-grade acetonitrile and methanol were obtained from Burdick & Jackson Company (Morristown, NJ, USA). Caco-2 cells were supplied by the Korean Cell Line Bank (Seoul, Korea) and cultured according to the supplier’s recommendations. Transwell (24-well, 6.5 mm polycarbonate inserts, 0.4-μm pore) and cell culture reagents were purchased from Corning Life Sciences. Heparinized human plasma was obtained from donors at the Severance Hospital of Yonsei University Health System (Seoul, Korea) and stored at −80 °C prior to use.