Development of an LC–MS/MS method for quantification of two pairs of isomeric flavonoid glycosides and other ones in rat plasma: Application to pharmacokinetic studies

Abstract

A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for simultaneous determination of six flavonoid glycosides - isoorientin (1), orientin (2), 2″-O-β-d-xylopyranosyl isoorientin (3), 2″-O-β-d-xylopyranosyl isovitexin (4), 6-C-l-α-arabipyranosyl vitexin (5) and vitexin (6) - in rat plasma using isoquercitrin as the internal standard (IS). Plasma samples were prepared by a one-step protein precipitation with acetonitrile. Chromatographic analysis was carried out on a 25 cm C column with a gradient mobile phase consisting of acetonitrile and 0.1% aqueous formic acid. Six analytes and IS were detected through electrospray ionization in negative-ion selection reaction monitoring mode. The mass transitions were as follows: m/z 447.2 → 327.0 for 1, m/z 447.2 → 327.0 for 2, m/z 579.3 → 458.9 for 3, m/z 563.0 → 293.1 for 4, m/z 563.0 → 353.0 for 5, m/z 431.1 → 311.1 for 6, and m/z 463.1 → 300.2 for IS. Calibration curves exhibited good linearity (r > 0.9908) over a wide concentration range for all compounds. Intra- and inter-day precision (RSD, %) at four different levels were both <14.2% and the accuracy (RE, %) ranged from -11.9 to 12.0%. The extraction recoveries of the six components ranged from 88.2 to 103.6%. The validated assay was successfully applied to the pharmacokinetic studies of the six components in male rat plasma after intravenous administration of total flavonoids of Scorzonera austriaca Wild.

Determination of isoorientin levels in rat plasma after oral administration of Vaccinum bracteatum Thunb. methanol extract by high‐performance liquid chromatography‐tandem mass spectrometry

“…When chromatograms of blank rat plasma samples were compared with plasma spiked with IS only and plasma spiked with 1 or 5 ng/mL of isoorientin, no peaks interfered with those of isoorientin and the IS (Figure ), and the peaks of isoorientin and of the IS were symmetrical. The retention times of isoorientin and IS in plasma were ~5.3 and 4.1 min, respectively, and the total run time was 8 min (Figure B–D), which is shorter than that of LC‐MS/MS method reported previously (8 min vs 16 or 20 min; Sheng et al ., ; Zhang et al ., ). The retention times of isoorientin and IS in standard plasma samples were identical to those obtained for plasma after the oral administration of VBME at 150 mg/kg (Figure E).…”

“…Quantitative analyses were performed using MRM modes at m / z 449.0 → 299.1 (for isoorientin) and m / z 417.1 → 297.1 (for IS). Compared with previous LC‐MS/MS methods (Sheng et al ., ; Zhang et al ., ), both previous MS/MS methods for isorientin employed the MRM with the ESI − mode at m / z 447.3 → 326.8 and m / z 447.0 → 327.0, respectively.…”

“…However, few reports have been issued on the use of liquid chromatography tandem mass spectrometry (LC‐MS/MS) in negative electrospray ionization (ESI − ) mode to quantify isoorientin levels in biological fluids. Recently, Zhang et al () developed a LC‐MS/MS method to quantify six flavonoid glycosides, including isoorientin, from Scorzonera ausriaca Wild, in rat plasma, and earlier Sheng et al () developed a LC‐MS/MS method for the simultaneous determination of secoiridoid glycosides and flavonoid glycosides, including isoorientin, in the plasma of rats administered Swertia pseudochinensis extract. However, these methods require a sample preparation process that includes liquid–liquid extraction with ethyl acetate (Sheng et al ., ) or protein precipitation followed by evaporation to dryness (Zhang et al ., ), and utilize complex gradient elution with a long total run time (20 and 10 min followed by 6 min column equilibration, respectively; Zhang et al, ; Sheng et al ., ).…”