Comparative Study on the Active Components of Favonoids Between Peel and Pulp of Aurantii Fructus Immaturus by UPLC-MS/MS

OBJECTIVE  To develop a UPLC-MS/MS method for simultaneous determination of 7 flavonoids between peel and pulp of Aurantii Fructus Immaturus, and to compare the content differences in different parts of Aurantii Fructus Immaturus.

METHODS  The analysis was carried out on a Waters ACQUITY UPLC HSS T3 column(2.1 mm×100 mm, 1.8 μm) at 30 ℃, with the mobile phase of water containing 0.2% formic acid and acetonitrile under gradient elution, the flow rate was 0.4 mL·min^–1, and the injection volume was 2 μL. The electrospray ionization(ESI) source in the positive ion mode and multiple reaction monitoring(MRM) were used for MS quantitative analysis. The content determination results of 24 batches of Aurantii Fructus Immaturus between peel and pulp were analyzed by the cluster analysis(CA), principal component analysis(PCA) and orthogonal partial least squares discriminate analysis(OPLS-DA).

RESULTS  The linearity of the 7 components was good within their linear range. The results of 24 batches of samples showed that the contents of apigenin and kaempferol were higher in the pulp group, while the contents of 5,7,3',4'-tetramethoxyflavone, 6-demethoxytangeretin, 5,7,3',4',5'-pentamethoxyflavone, isosinensetin, and 5-demethylnobiletin were lower in the pulp group than in the peelp group, and all of them were significantly different. The results of multivariate statistical analysis showed that peel and pulp could be divided into 2 categories according to different tissue parts. Among them, 5,7,3',4',5'-pentamethoxyflavone, isosinensetin, 6-demethoxytangeretin and 5-demethylnobiletin contributed more to differentiate between peel and pulp. In particular, 5,7,3',4',5'-pentamethoxyflavone and 5-demethylnobiletin could be the main differentiating components between the peel and pulp.

CONCLUSION  An accurate and reliable UPLC-MS/MS quantitative method for the determination of 7 flavonoid components in the peel and pulp is established. There are significant differences in the contents of 7 flavonoids between peel and pulp, which can provide a more adequate scientific basis for the scientific connotation about the appearance characteristic of Aurantii Fructus Immaturus.