Bupropion

Bupropion, metroprolol, phenacetin, midazolam and tolbutamide (all > 98%) and the internal standard diazepam (IS) were obtained from Sigma-Aldrich Company (St. Louis, MO). Ultra-pure water was prepared by Millipore Milli-Q purification system (Bedford, MA). Methanol and acetonitrile (HPLC grade) were obtained from Merck Company (Darmstadt, Germany).

Phenacetin, acetaminophen, 2-acetamidophenol, coumarin, 7-hydroxy-comuarin (umbelliferone), flurbiprofen, dextromethorphan, dextrorphan, trazodone, thymol, bupropion, 6-hydroxy-bupropion, alprazolam, 1′-hydroxy-alprazolam, NADP⁺, isocitrate dehydrogenase, and DL-isocitrate were purchased from Sigma-Aldrich (St. Louis, MO, USA). Pronethalol was from Tocris (Minneapolis, MN, USA). Propofol was provided by Zeneca Pharmaceuticals (Wilmington, DE, USA). 4′-hydroxy-flurbiprofen, 2-fluoro-4-biphenyl-acetic acid, 4-hydroxy-propofol, chlorzoxazone, and 6-hydroxy-chlorzoxazone were purchased from Toronto Research Chemicals (Toronto, ON, Canada). S-Mephenytoin and 4-hydroxymephenytoin were purchased from Gentest (Corning, Corning, NY, USA). GW340416A, a chemical analogue of bupropion, was kindly provided by GlaxoSmithKline (Research Triangle Park, NC, USA). Omeprazole was purchased from BeanTown Chemical, Inc. (Hudson, NH, USA) and omeprazole sulfone was purchased from Cayman Chemical Company (Ann Arbor, MI, USA). Sodium hydroxide, potassium phosphate monobasic and potassium phosphate dibasic and EDTA were purchased from J.T. Baker (Center Valley, PA, USA). HPLC-grade acetonitrile and methanol were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Ultra-pure water was obtained using a Milli-Q^® Q-POD Millipore System (EMD Millipore, Burlington, MA, USA).

The permeability of LMDS-1 and -2 was determined by a PAMPA-BBB assay. Firstly, 300 μl of LMDS-1 or -2 (1 μM) solution were added to the donor well (Millipore). Bupropion (a high permeability marker), piroxicam (a low permeability marker), and lucifer yellow (an integrity marker) (Sigma-Aldrich) were used as quality control (QC) compounds for comparison. The sandwich with aqueous donor on the bottom, artificial lipid membrane (PVDF filter coated with porcine polar brain lipid) in the middle, and aqueous acceptor (5% DMSO in PBS) on the top was assembled as stated [69 (link)]. Each compound was tested in triplicate. After incubation for 18 h at the ambient temperature, the PAMPA sandwich plate was separated. The concentrations of test and QC compounds in the donor and acceptor wells were measured as stated [69 (link)], and the effective permeability coefficient (P_e) was computed [70 (link)].