Identification and assay of 3′-O-methyltaxifolin by UPLC–MS in rat plasma

A new metabolite of taxifolin: 3′-O-methyltaxifolin (3′-O-MTAX) in Caco-2 cells and in rat plasma was identified. The chemical structure of 3′-O-MTAX was determined by MS and 1H NMR. A rapid, sensitive and specific UPLC–MS method to determine 3′-O-MTAX in rat plasma was also developed. Following ethyl acetate extraction, 3′-O-MTAX in plasma was separated on a Sunfire™ (2.1 mm × 50 mm, 3.5 μm) column and analyzed in the selected ion recording with a negative electrospray ionization mode using puerarin as the internal standard. The lower limit of quantification (LLOQ) was 2.75 ng/mL. Intra- and inter-day precisions (% RSD) were all within 7.2% and accuracy (% deviation) ranged from −5.0 to 4.7%. The overall recoveries at four concentrations were all >72.0%. This validated method was successfully applied to measure 3′-O-MTAX in rat plasma after oral administration of taxifolin.