Flurbiprofen

Caco-2 cells were purchased from Sigma Aldrich (Schnelldorf, Germany) and cultured in Minimum Essential Medium Eagle medium supplemented with 10% fetal calf serum (FCS) (Thermo Fisher Scientific, Schwerte, Germany), 2 mM L-glutamine (Thermo Fisher Scientific, Schwerte, Germany) and 1× non-essential amino acid solution (Sigma Aldrich, Schnelldorf, Germany). All media contained 1% penicillin/streptomycin (Thermo Fisher Scientific, Schwerte, Germany), and cells were cultured at 37 °C in a 5% CO₂ atmosphere. Ibuprofen was obtained from Sigma Aldrich (Schnelldorf, Germany) and flurbiprofen, etoricoxib and paracetamol were purchased from Biomol (Hamburg, Germany). Ibuprofen was dissolved in water. For in vitro experiments, flurbiprofen, etoricoxib and paracetamol were dissolved in 100% dimethyl sulfoxide (DMSO) (Sigma Aldrich, Schnelldorf, Germany) and further diluted in media (c_stock = 25 mg/mL (for flurbiprofen and paracetamol), c_stock = 20 mg/mL for etoricoxib; maximum DMSO concentration during experiments 0.1% (etoricoxib, paracetamol); 0.2% (flurbiprofen)). To exclude DMSO effects in the infection experiments, flurbiprofen and paracetamol were dissolved in water and further diluted in media (c_stock = 500 µg/mL and 10 mg/mL for flurbiprofen and paracetamol, respectively).

Flurbiprofen (FLUR), Hyaluronic acid (HA), Bovine serum albumin (BSA), Cetyltrimethylammonium bromide (CTAB), and Complete Freund’s adjuvant (CFA) were purchased from Sigma-Aldrich (St. Louis, MO, USA). Chloroform, Methanol, and Citric acid were purchased from Al-Gomhureya Co. (Cairo, Egypt). All other reagents and chemicals were of analytical grade.

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).