Metabolomic identification of antibacterial 5‑indanol from Amomum tsaoko fruits and the application of its stem‑leaf by‑products as feed additives for Partridge Shank chickens.

Amomum tsaoko is a medicinal and edible Zingiberaceae herb, while its stems and leaves are largely discarded and underutilized compared with its fruits. This study aimed to characterize the antibacterial active components of A. tsaoko stem and leaf ethyl acetate extract, clarify the antibacterial mechanism of core metabolite 5-indanol, and evaluate the application value of A. tsaoko stem-leaf by-products as green feed additives for Partridge Shank chickens. In vitro antibacterial assays combined with GC-MS metabolomics were performed on four Zingiberaceae species (A. tsaoko, Alpinia officinarum, A. villosum, A. oxyphylla); broth microdilution, time-kill curve, biofilm and cell membrane leakage tests were conducted to verify the bactericidal mode of 5-indanol. For the animal feeding trial, 30-day-old Partridge Shank chickens were randomly divided into five groups: control group (CK, basal diet), 0.05% A. tsaoko stem-leaf group (SL0.05), 0.10% stem-leaf group (SL0.10), 0.15% stem-leaf group (SL0.15), and positive control group supplemented with 0.10% A. tsaoko fruit (F0.10). Growth performance, meat quality, small intestinal morphology and cecal microbiota were systematically detected. The in vitro results revealed that ethyl acetate extract of A. tsaoko stems and leaves exhibited broad-spectrum antibacterial activity against Gram-positive and Gram-negative bacteria, with an MIC of 62.5 μg/mL against Staphylococcus aureus. Metabolomic screening identified 5-indanol as the dominant antibacterial marker compound, whose antibacterial efficacy was 20-fold higher than 2,3-dihydro-1H-indene-4-carbaldehyde. Mechanistically, 5-indanol disrupted bacterial membrane integrity, triggered leakage of intracellular biomacromolecules, suppressed biofilm formation and attenuated hemolytic virulence of S. aureus. In vivo data indicated that 0.10% dietary supplementation of A. tsaoko stems and leaves (SL0.10) achieved the optimal effect: Dietary supplementation with A. tsaoko significantly increased the average daily gain and reduced the feed conversion ratio of Partridge Shank chickens, with the most pronounced effect observed in the 0.10% stem-leaf group. The drip loss of breast and thigh muscle was decreased without adverse impacts on meat pH, color and tenderness. The villus height/crypt depth ratios of duodenum and ileum were elevated by 34.2% and 26.43%, respectively. Meanwhile, the relative abundances of beneficial phyla Firmicutes, Bacteroidetes and Ascomycota in gut microbiota were significantly enriched, and the alpha diversity of bacterial and fungal communities was improved. Collectively, 5-indanol is the key antibacterial component in the ethyl acetate extract of A. tsaoko. Waste stems and leaves of A. tsaoko can be developed as novel antibiotic-free phytogenic feed additives, which realizes full-plant high-value utilization of A. tsaoko agricultural by-products and provides theoretical support for sustainable green livestock breeding.