Quantification of Epoxyeicosatrienoic Acids

EDTA-free Halt protease inhibitor cocktail, dithiothreitol, iodoacetamide, Acros Organics’ ammonium bicarbonate buffer (98% purity), and peptides of interest listed in Supplementary Table S1 were obtained from Thermo Fisher Scientific (Waltham, MA, USA) and used without further purification. Dulbecco’s phosphate-buffered saline, pH 7.4 (DPBS, 10×), Pierce bicinchoninic acid (BCA) protein assay kit, sodium chloride, potassium phosphate, ammonium acetate, ACS-grade ethyl acetate, chloroform, optima-grade acetonitrile, water, methanol, formic acid, and acetic acid were purchased from Fisher Scientific (Hampton, NH, USA). Terfenadine, hydroxy terfenadine, carboxy terfenadine hydrochloride, triphenylphosphine, azacyclonol, NADPH, and PLA₂ from Naja mossambica mossambica were obtained from Sigma-Aldrich (St. Louis, MO, USA). A 1 mg/mL ethanolic solution of midazolam was purchased from Cerilliant Corporation (Round Rock, TX, USA) and used as an internal standard. Precellys 24 homogenizer, reinforced homogenization tubes and stainless-steel beads were obtained from Bertin Instruments (Rockville, MD, USA). Human serum albumin (HSA) and bovine serum albumin (BSA) were purchased from Calbiochem (Billerica, MA, USA). 14,15-EET and deuterated 14,15-EET, 11,12-EET, 8,9-EET and deuterated 8,9-EET, 5,6-EET and deuterated 5,6-EET, and 14,15-DHET-d₁₁ were obtained from Cayman Chemical (Ann Arbor, MI, USA).