The Hepatoprotective Properties of the Revised Formulation of Dahuang Xiaoshi Tang, an Ancient Chinese Herbal Decoction, Are Probed by Integrated Metabolomics and Network Pharmacology

<b>Background</b>: Dahuang Xiaoshi Tang (DXT), an ancient Chinese herbal remedy dating back to 220 AD, as documented initially in "Treatise on Febrile and Miscellaneous Diseases," is used to treat damp-heat jaundice with interior sthenia syndrome. In DXT, anthraquinones and alkaloids form insoluble complexes, reducing its effectiveness. A revised herbal extract, DXT-M, was developed, and its hepatoprotective properties were demonstrated in animal models using pharmacodynamic, metabolomic, network pharmacological, and toxicological approaches. <b>Methods</b>: The α-naphthalene isothiocyanate was utilised to establish the acute liver injury rat model. The assays of glutamate pyruvate transaminase, glutamic oxalacetic transaminase, alkaline phosphatase, bilirubin, total bile acid, complement 3 (C3) and C4, interleukin-2 (IL-2) and IL-6, tumour necrosis factor α (TNF-α), and pathological morphology were used to evaluate the hepatoprotection of DXT in comparison to DXT-M. The ¹H-NMR-based serum and urine metabolomics were performed to identify potential biomarkers and metabolic pathways of DXT-M in treating hepatitis. The intrinsic regulatory mechanisms of DXT in liver protection, as well as the combination of network toxicology, were elucidated. Statistical analyses included RM two-way ANOVA with Geisser-Greenhouse correction and Dunnett's post hoc test for longitudinal data, and one-way ANOVA with Dunnett's post hoc test for group comparisons. Data were shown as mean ± SD. <b>Results</b>: Liver-injured animals exhibited weight loss, ruffled fur, and liver damage, accompanied by elevated liver function indicators. DXT-M effectively improved these symptoms, repaired liver damage, restored liver function, and regulated immune status by modulating complement 3. Metabonomics and other analyses indicated the CYP/GST-ROS axis is key to its hepatoprotective effects. DXT-M outperformed DXT in efficacy. <b>Conclusions</b>: DXT-M demonstrated significant effectiveness in restoring liver pathological damage, correcting abnormal biochemical indicators of liver function, and regulating complement factors. The pathway of CYP/GST-ROS served as the shared regulatory axis and transformation site for DXT-M's liver protective effects. These findings suggest that DXT-M has potential as a treatment for acute liver injury, highlighting the need for further research into its underlying molecular mechanisms as well as its complete material basis. This study's main limitation is its focus on acute models; future research should include other liver diseases and clinical observation to evaluate its full potential.