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To investigate the effects of P. ginseng C.A. Mey (P. ginseng) on the metabolism of diester alkaloids and explore the potential mechanism. P. ginseng was administered orally to rats for 7 days, after which liver microsome samples were prepared and then incubated with diester alkaloids. Ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry was used to determinate the concentration of diester alkaloids to calculate the clearance rate. The cocktail method was used to evaluate the effects of oral administration of P. ginseng extracts on the activities of cytochrome P450 (CYP) isoforms in rats through the changes in the pharmacokinetic parameters of the probe drugs. The protein and gene expression of CYP3A2 and pregnane X receptor (PXR) in rats were evaluated by western blotting and quantitative PCR. The specific enzyme inhibitor method and human recombinant enzyme method were used to identify the involvement of sub-CYPs in the metabolism of diester alkaloids in human liver microsomes (HLMs). The clearances of aconitine, mesaconitine, and hypaconitine in the P. ginseng groups were lower than those of the control group. The areas under the curve of midazolam were 2.37 ± 1.05, 4.96 ± 0.51, and 6.23 ± 1.30 mg·L−1·h for the low-, medium-, and high-dose P. ginseng groups, respectively, which were higher than that of the control (2.23 ± 0.64 mg·L−1·h). The clearances of midazolam for the medium- (1.87 ± 0.16 L·h−1·kg−1) and high-dose (1.60 ± 0.34 L·h−1·kg−1) P. ginseng groups were lower than that of the control group (4.66 ± 1.43 L·h−1·kg−1). After exposure to P. ginseng extracts, the gene and protein expression levels of CYP3A4 and PXR were decreased. The hepatic metabolism rates of aconitine, mesaconitine, and hypaconitine in HLMs were decreased to 60.37%, 21.67%, and 10.11%, respectively, when incubated with ketoconazole, a specific inhibitor for CYP3A. The kinetic plots indicated that the KM and Vmax values of CYP3A4 were 10.08 ± 3.26 μM and 0.12 ± 0.01nmol·mg protein−1·min−1 for aconitine, 131.3 ± 99.75 μM and 0.73 ± 0.44 nmol·mg protein−1·min−1 for mesaconitine, and 17.05 ± 9.70 μM and 0.16 ± 0.04 nmol·mg protein−1·min−1 for hypaconitine, respectively. The in vitro mean intrinsic clearance rates by CYP3A4 were 0.0119, 0.0056, and 0.0091 mL·nmol CYP−1·min−1 for aconitine, mesaconitine, and hypaconitine, respectively. Therefore we implied that P. ginseng inhibited the metabolism of diester alkaloids in vitro and decreased the CYP3A4 enzyme activity as well as the gene and protein expression of CYP3A4 and PXR in vivo. CYP3A4 had a larger effect on diester alkaloid metabolism than the other human CYP isoforms, CYP1A2, CYP2C9, and CYP2E1.

BackgroundThe side effects caused by Polygoni Multiflori Radix (PMR) and Polygoni Multiflori Radix Praeparata (PMRP) have often appeared globally. There is no research on the changes of endogenous metabolites among PMR- and PMRP-treated rats. The aim of this study was to evaluate the varying metabolomic effects between PMR- and PMRP-treated rats. We tried to discover relevant differences in biomarkers and endogenous metabolic pathways.MethodsHematoxylin and eosin staining and immunohistochemistry staining were performed to find pathological changes. Biochemical indicators were also measured, one-way analysis of variance with Dunnett’s multiple comparison test was used for biochemical indicators comparison among various groups. Metabolomics analysis based on ultra-high performance liquid chromatography-quadrupole time of flight mass spectrometry (UPLC-Q/TOF-MS) was performed to find the changes in metabolic biomarkers. Multivariate statistical approaches such as principal component analysis (PCA) and orthogonal partial least square-discriminant analysis (OPLS-DA) were applied to reveal group clustering trend, evaluate and maximize the discrimination between the two groups. MetaboAnalyst 4.0 was performed to find and confirm the pathways.ResultsPMR extracts exhibited slight hepatotoxic effects on the liver by increasing aspartate and alanine aminotransferase levels. Twenty-nine metabolites were identified as biomarkers, belonging to five pathways, including alpha-linolenic acid metabolism, taurine and hypotaurine metabolism, glycerophospholipid metabolism, arginine and proline metabolism, and primary bile acid biosynthesis.ConclusionThis study provided a comprehensive description of metabolomic changes between PMR- and PMRP-treated rats. The underlying mechanisms require further research.

Objectives To investigate whether Panax ginseng (P. ginseng) could affect the metabolism of Diester Alkaloids (DAs) derived from Aconiti Lateralis Radix in vivo. Methods and Results 24 male Sprague-Dawley rats were randomized for 7-day treatment with P. ginseng (low, middle, and high), or vehicle. Aconiti Lateralis Radix was administered orally to each group on the 8th day. Plasma samples were collected, and Xevo TQ-S was used to detect the concentration of aconitine, mesaconitine, and hypaconitine in plasma. We describe a fast and reproducible method to detect the concentration of aconitine, mesaconitine, and hypaconitine in plasma. Compared to the control group, the AUC(0-t) of three DAs increased in both the middle and high dosing groups. The Vz/F of three DAs in these groups as well as the CLz/F of aconitine in all P. ginseng groups and the CLz/F of mesaconitine and hypaconitine in P. ginseng middle and high groups were decreased compared to the control group. Conclusion Orally administrated P. ginseng potentially inhibits the metabolism of DAs from Aconiti Lateralis Radix in rats.

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This study focused on the differential metabolomic effects between water extracts of Polygoni Multiflori Radix and Polygoni Multiflori Radix Preparata in rats. The extracts were subsequently administered for 28 d. Serum biochemical indicators were tested, hematoxylin-eosin staining and immunohistochemistry staining were used to detect histopathological changes in the livers. Ultra-performance LC/quadrupole timeof-flight mass spectrometry was used to detect the changes in endogenous metabolites. Finally, we performed detailed analysis of the changes in metabolic pathways. Hematoxylin-eosin staining and immunohistochemistry staining results indicated that the water extracts of Polygoni Multiflori Radix and Polygoni Multiflori Radix Preparata had mild liver injury effect. Fifty-two differential endogenous biomarkers were confirmed as potential biomarkers between Polygoni Multiflori Radix and Polygoni Multiflori Radix Preparata groups. In the positive ion mode, the biomarkers included 31 Phosphatidyl cholines (PCs), six lysoPCs, and ceramide. In the negative ion mode, 12 biomarkers were confirmed, including glycodeoxycholic acid, chenodeoxycholic acid, and deoxycholic acid, etc. In Hydrophilic Interaction Liquid Chromatography (HILIC) mode, nine biomarkers were confirmed, including niacinamide, L-palmitoylcarnitine, and butyrylcarnitine, etc. Using MetaboAnalyst 4.0, six related metabolic pathways, including taurine and hypotaurine metabolism, sphingolipid metabolism, glycerophospholipid metabolism, nicotinate and nicotinamide metabolism, arginine and proline metabolism, and tryptophan metabolism and primary bile synthesis, were confirmed as the most differential pathways between the Polygoni Multiflori Radix and Polygoni Multiflori Radix Preparata groups.