Hesperetin-7--glucoside (Hes-7-G) is a typical flavonoid monoglucoside isolated from Citri Reticulatae Pericarpium (CRP), which is commonly used as a food adjuvant and exhibits potential biological activities. To explore the interaction between Hes-7-G ingestion and microbiome and host metabolism, here, 16S rRNA gene sequencing was first used to analyze the alteration of fecal microbiome in mice after Hes-7-G intake. Metabolic homeostasis in mice was subsequently investigated using untargeted H NMR-based metabolomics and targeted metabolite profiling. We found that dietary Hes-7-G significantly regulated fecal microbiota and its derived metabolites, including short-chain fatty acids (SCFAs) and tryptophan metabolites (indole and its derivatives), in feces of mice. Regulation of microbiota was further confirmed by the significantly changed urinary hippurate and trimethylamine -oxide (TMAO), co-metabolites of the microbe and host. We also found that dietary Hes-7-G modulated the host tricarboxylic acid cycle (TCA) involved in energy metabolism. These findings suggested that Hes-7-G exhibits potential beneficial effects for human health.