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Objective·To study the effect of rhubarb administration on the balance between intestinal flora and the body.Methods·Wistar rats were randomly divided into 4 groups (n=8), which were given extractions of rhubarb 0.1 g/kg (low dose group), 2.5 g/kg (medium dose group), 4.5 g/kg (high dose group) and normal saline (control group) by intragastric administration for 5 d, and the daily change of fecal water content of rats was observed. Gas chromatography/time of flight mass spectrometry (GC/TOFMS) approach was used to detect the metabolites in serum, colon tissue and fecal of rats on the 5th day of administration. Principal component analysis (PCA) and partial least squares discrimination analysis (PLS-DA) were used to analyze the differences of metabolites between different dose groups and the control group. The metabolites with statistical significance were obtained by t-test.Results·The water content of rat feces in the dose group gradually increased with the time and dose after rhubarb administration. Compared with the control group, 28, 18 and 20 differential metabolites were obtained in serum, colon tissue and fecal samples from different dose groups, which showed significant changes (P