Targeted and non-targeted metabolomics uncovering the effects of Er-Miao-Wan formula on rats with hyperuricemia

Er-Miao-Wan formula (EMW), composed of Phellodendri Chinensis Cortex and Atractylodis Rhizoma, is widely used in the treatment of hyperuricemia (HUA), gout, and related complications as a classic compound formula. However, its mechanisms for the treatment of HUA still need to be further systematically investigated. The study aimed to perform modern analytical techniques to elucidate the mechanisms of EMW in improving the symptoms of HUA from the perspective of metabolomics. We used a high-fructose diet to establish a rat model of HUA to evaluate the effects of EMW on improving HUA. Next, we established a targeted metabolomics analysis method to quantitatively analyze purine metabolites in plasma by using ultra-high-performance liquid chromatography with ultraviolet and triple quadrupole mass spectrometry (UHPLC-UV-QQQ MS), and combined with plasma non-targeted metabolomics analysis by using ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UHPLC-Q/TOF MS) to clarify the potential mechanisms of EMW to improve HUA. Oral administration of EMW could significantly increase the urinary uric acid and decrease the serum uric acid, and exhibited a remarkable effect on improving HUA. Plasma targeted metabolomics analysis showed that six purine metabolites related to HUA, including uric acid, hypoxanthine, xanthine, deoxyadenosine, deoxyguanosine, and deoxyinosine, were changed in the EMW-treated group. Further, principal component analysis (PCA) and partial least squares discrimination analysis (PLS-DA) showed that the mechanism of EMW interfering with purine metabolic pathway in the rats with HUA could be different from that of allopurinol. On the basis of plasma non-targeted metabolomics, PCA and orthogonal partial least squares discriminant analysis (OPLA-DA) screened and identified 23 potential biomarkers in the rats with HUA, and 11 biomarkers showed a trend of reversion after the intervention of EMW. The pathway analysis suggested that EMW might have therapeutic effects on the rats with HUA via the metabolic pathways including phenylalanine metabolism, glycerophospholipid metabolism, and tryptophan metabolism. In this study, a plasma targeted metabolomics method that can simultaneously quantify nine purine metabolites in rats with HUA was established for the first time, which can be used to study diseases closely related to HUA. In addition, we further explored the overall effect of EMW on HUA in combination with the metabonomic method established by non-targeted metabolomics, which was helpful to solve the defect that the pharmacological mechanism caused by multi-components and multi-targets of traditional Chinese medicine was difficult to explain scientifically and comprehensively. In summary, EMW could effectively alleviate the symptoms of high-fructose-induced HUA, and the study provided a reference for the potential therapeutic mechanism of EMW.