Lonicerae japonicae Flos (LJF) possesses a good anti-respiratory syncytial virus (RSV) effect. However, the material basis of LJF in treating RSV is still unclear. In this study, a sensitive and accurate quantitative method based on UHPLC-QQQ MS was established and validated for the simultaneous determination of the 15 ingredients from LJF in RSV-infected mice plasma. Multiple reaction monitoring was performed for quantification of the standards and of the internal standard in plasma. All the calibration curves show good linear regression within the linear range (r² > 0.9918). The method validation results, including specificity, linearity, accuracy, precision, extraction recovery, matrix effect, and stability of 15 ingredients, are all within the current acceptance criteria. This established method was successfully applied to the pharmacokinetic study of 15 compounds from LJF. Furthermore, the repair rate of lung index and the improvement rate of IFN-γ and IL-6 improved after administration of the LJF, indicating that LJF possessed a positive effect on the treatment of RSV infection. Finally, by combining Spearman and Grey relation analysis, isochlorogenic acid B, isochlorogenic acid C, secoxyloganin, chlorogenic acid, and loganic acid are speculated to be the main effective ingredients of LJF in treating RSV. This study lays the foundation for attempts to reveal the mechanisms of the anti-RSV effect of LJF.

Benzylisoquinoline alkaloids (BIAs) exhibit diverse biological activities, such as neuroprotective effects. The delta-opioid receptor (DOR) has emerged as a promising therapeutic target due to its potential role in enhancing neuroprotection and regeneration. However, reports on the binding of BIAs to the DOR remain scarce. Here, neferine, a BIA from Nelumbo nucifera, as a potential DOR agonist. Molecular docking ranked neferine among the top of 15 BIAs. Initial binding was detected by cellular membrane chromatography and quantitatively confirmed by bio-layer interferometry, with a K_D value of 37.4 μM. ONE vector G protein Optical biosensor revealed that G_i2, G_i3 and G_Z signaling could be activated by neferine through DOR modulation. Consistent with the G_i/z activation, neferine dose-dependently inhibited cAMP accumulation with an EC₅₀ of 0.25 µM. Transcriptomic analysis in DOR-overexpressing HEK293T cells indicated that neferine stimulation predominantly regulates gene networks governing cell cycle and stress adaptation. However, direct transcriptional signature for neuroprotection was not predominant in our system, suggesting that DOR signaling may exhibit context-dependent effects. In conclusion, we identified the neferine as a natural DOR agonist through in silico and in vitro approach, providing a reference for further investigation into its pharmacological potential.

Objectives: This study aimed to investigate the therapeutic effects and potential mechanisms of Lonicerae japonicae Flos (Jinyinhua, JYH) against respiratory syncytial virus (RSV)-induced pneumonia by integrating lung tissue metabolomics with gut microbiota analysis. Methods: An RSV-infected mouse model was established through intranasal inoculation. Lung pathological changes, viral RNA levels, lung index, and inflammatory cytokine levels were evaluated. Untargeted metabolomics and 16S rRNA gene amplicon sequencing were performed to characterize JYH-mediated alterations in pulmonary metabolites and the gut microbiota. Spearman correlation analysis was conducted to assess associations between differentially abundant bacterial genera and significantly altered metabolites. Results: JYH alleviated RSV-induced pulmonary histopathological injury, reduced viral RNA levels, decreased lung index and interleukin-6 (IL-6) levels, and increased interferon-γ (IFN-γ) levels. Metabolomic profiling identified 46 differential metabolites, among which 26 showed a reversal trend following JYH administration. These metabolites were mainly enriched in pathways associated with the synaptic vesicle cycle, lysosomal function, and Forkhead box O (FoxO) signaling. Gut microbiota analysis showed that JYH increased microbial richness and diversity, whereas KEGG-based functional prediction indicated that the differentially abundant taxa were primarily involved in amino acid, carbohydrate, and nucleotide metabolism. Moreover, correlation analysis revealed significant associations between key bacterial genera, including Gemella, Sutterella, and CC_115, and differential metabolites such as pyridoxamine, uridine monophosphate (UMP), and argininosuccinic acid. Conclusions: JYH may protect against RSV-induced pneumonia by restoring pulmonary metabolic homeostasis and modulating gut microbiota composition. These findings provide new insights into metabolite–microbiota interactions underlying the anti-RSV activity of JYH.