The development and validation of an HPLC-MS/MS method for the determination of eriocitrin in rat plasma and its application to a pharmacokinetic study

Abstract

Eriocitrin is one of the major active constituents of lemon fruit, and it possesses strong antioxidant, lipid-lowering, anticancer and anti-inflammatory activities and has long been used in food, beverages and wine. In this study, for the first time, a rapid, selective, and sensitive liquid chromatography-tandem mass spectrometry method (LC/MS/MS) with protein precipitation was developed and validated for the analysis of eriocitrin in rat plasma. Chromatographic separation was achieved using a mobile phase, comprising 0.1% formic acid aqueous solution and acetonitrile eluted at a flow rate of 0.8 mL min^-1. In multiple reaction monitoring (MRM) modes, eriocitrin and internal standard (IS) were quantified using precursor-to-product ion transitions of m/z 595.4 → 287.1 and m/z 252.0 → 155.9, respectively. The intra- and inter-day precision (RSD) were below 6.79% in plasma, while accuracy (RE) was within ±7.67%. The matrix effect, recovery and stability were also demonstrated to be within acceptable limits. This method was successfully employed in the pharmacokinetic study on rats after the oral administration of eriocitrin. The pharmacokinetic parameters show that the maximum plasma concentration (C _max) of eriocitrin was 299.833 ± 16.743 μg L^-1, while the corresponding time to reach C _max(T _max) was 0.094 ± 0.019 h, and the half-time (T _1/2) was 1.752 ± 0.323 h. The present results would be valuable for further research and development of eriocitrin.

Fig. 1. MS/MS spectra of eriocitrin and IS (sulfamethoxazole) in negative mode.

Fig. 2. MRM chromatograms of the blank plasma (A), plasma spiked with eriocitrin and IS (B), and plasma samples after oral administration of eriocitrin (C).

Fig. 3. Plasma concentration–time profile after a single oral administration of eriocitrin (50 mg kg⁻¹) to rats. Data are expressed as the mean ± SD (n = 6).