LC-ESI-MS-MS Determination of Rat Plasma Protein Binding of Major Flavonoids of Flos Lonicerae Japonicae by Centrifugal Ultrafiltration

Centrifugal ultrafiltration (CU) and a fast liquid chromatography with electrospray ionization tandem mass spectrometric (LC-ESI-MS-MS) method have been developed for drug-protein binding analysis of four flavonoids of Flos Lonicerae Japonicae (FLJ). A rapid separation of free drug from the plasma was achieved by CU and the simultaneous analysis of four flavonoids of FLJ in rat plasma was by fast LC-ESI-MS-MS. The chromatographic analytical time decreased to 6.5 min without sacrificing resolution using a 4.6 mm × 50 mm, 1.8 μm particle size reverse phase column and MS–MS mode performed for multiple reaction monitoring (MRM). The protein binding rates of four flavonoids were in the range of 66.8 ± 7.4 to 81.3 ± 5.1% after oral administration of the flavonoid fraction of FLJ. All values were almost similar to 70.0% at 30, 45 and 80 min. Its result suggests that these flavonoids in rat plasma perform high protein binding rates and present a stable binding in a longer time. Meanwhile, it reveals that the elimination of these flavonoids may be relatively moderate in the body, the performance of a certain degree of accumulation may be speculated and make further impacts on the pharmacokinetic parameters.