Effects of rutin on in vitro rumen microbial composition and urea utilization in sheep.

This in vitro study aimed to evaluate rutin as a potential feed additive for improving nitrogen metabolism in the rumen. Two experiments were conducted. In Experiment 1, the effects of rutin (0, 7.5, 10, 12.5, and 15% of substrate dry matter) on fermentation were assessed. The 12.5% rutin dose (RT3) yielded the most favorable outcomes: it reduced ammonia nitrogen (NH₃-N) and pH while increasing microbial crude protein (MCP) after 12 h. RT3 also increased propionate proportion and decreased the acetate-to-propionate ratio, along with the proportions of acetate, isobutyrate, butyrate, valerate, isovalerate, and hexanoate. Microbial community analysis showed a significant increase in Patescibacter, Selenomonas, and Syntrophocococcus, while Spirochaetota and Prevotella decreased.In Experiment 2, a 3 × 4 factorial design explored the interaction between urea (2.5, 5, 7.5%) and rutin (7.5, 10, 12.5, 15%). The combination of 2.5% urea and 12.5% rutin produced the most pronounced synergistic effect: it significantly inhibited urease activity, reduced NH₃-N, and increased MCP. This combination also increased propionate while decreasing butyrate, isobutyrate, valerate, isovalerate, and hexanoate. Microbial analysis revealed enrichment of NK4A214_group and Selenomonas, with inhibition of Prevotella. The results suggest that rutin holds promise as a urease inhibitor to improve urea utilization in ruminants. Further in vivo studies are necessary to confirm these effects and to evaluate the long-term impacts of rutin on animal health and performance.