Chemical optimization and immunomodulatory activity of an optimized polyherbal functional beverage in Salmonella typhi-induced mice model

Polyherbal functional beverages enriched with diverse bioactive constituents are increasingly recognized as promising immunomodulatory nutraceutical candidates. This study aimed to optimize a polyherbal drink formulated from single clove garlic, red ginger, lemon, honey, and apple cider vinegar using Response Surface Methodology (RSM), and to evaluate its immunomodulatory efficacy through an in vivo experiment in mice induced with Salmonella typhi. Optimization targeted the maximization of total phenolic content, flavonoid concentration, and antioxidant activity. The optimized formulation showed enhanced flavonoid content and antioxidant activity compared with the commercial herbal drink. To validate its biological performance, the optimized beverage was administered to S. typhi–infected mice. Flow cytometry analysis revealed that infection induced significant alterations in T-cell subsets, characterized by increased CD4⁺ and CD8⁺ populations and elevated Th1 cytokines. Mice receiving the optimized polyherbal drink exhibited further enhancement of CD4⁺ T cells, increased production of IFN-γ and TNF-α, and markedly higher IL-10 expression, indicating improved immune activation accompanied by more balanced inflammatory regulation. These immunological improvements surpassed those observed in mice receiving the commercial formulation. The enhanced responses were likely attributable to the phytochemical composition of the optimized drink, including phenolics, flavonoids, and antioxidant constituents known to modulate T-cell activity and attenuate infection-induced inflammation. These findings highlight its potential as an immune-supportive nutraceutical and underscore the need for further research involving mechanistic elucidation, long-term safety evaluation, and clinical validation.