Abstract: In order to improve the physicochemical properties and antibacterial activity of rhein, its structure was modified. Using O-benzotriazole-N, N, N′, N′-tetramethylurea(HBTU) as condensing agent, and N, N-diisopropylethylamine(DIPEA) as catalyst, six target compounds (4a-4f) were synthesized by the reaction of rhein with L-proline methyl ester hydrochloride, L-tryptophan methyl ester hydrochloride, L-phenylalanine methyl ester hydrochloride and substituted benzylamine. The structures of the target compounds were confirmed and characterized by(¹H, ¹³C, ¹⁹F) NMR and high-resolution mass spectrometry. The antibacterial activities of these compounds against Escherichia coli, Staphylococcus aureus and Proteus vulgaris were tested for the first time. The results indicated that some of the target compounds had good antibacterial activities, and the minimum inhibitory concentration(MIC) value of compound 4a against S. aureus was 0.78 mg/L, and the MIC value of compound 4e against P. vulgaris was 0.39 mg/L. Their antibacterial activities were better than that of rhein and even better than that of the control drug ampicillin.