This experiment was conducted to investigate the effects of dietary tea seed polysaccharide and Macleaya cordata alkaloids on intestinal flora of yellow-feathered broilers and antimicrobial activity of Macleaya cordata sanguinarine. A total of 750 1-day-old healthy yellow-feathered broilers were randomly allocated to 5 groups with 6 replicates per group and 25 broilers per replicate. Broilers in control group were fed a basal diet, and the others in experimental groups were fed the basal diets supplemented with antibiotic (10 mg/kg colistin at 1 to 28 days of age and 5 mg/kg flavomycin at 29 to 56 days of age, antibiotic group), tea seed polysaccharide (0.04%, polysaccharide group), Macleaya cordata alkaloids (10 mg/kg at 1 to 28 days of age and 20 mg/kg at 29 to 56 days of age, Macleaya cordata group), tea seed polysaccharide (0.04%) and Macleaya cordata alkaloids (10 mg/kg at 1 to 28 days of age and 20 mg/kg at 29 to 56 days of age, polysaccharide+Macleaya cordata group).The experiment lasted for 56 days. Antibacterial effects and minimum inhibitory concentration (MIC) of Macleaya cordata sanguinarine on Staphylococcus aureus, Escherichia coil, Salmonella and Pasteurella were determined by agar diffusion paper method and double dilution method of test tube liquid, respectively. The results showed that:the ratio of villus height to crypt depth (V/C) of jejunum of yellow-feathered broilers in polysaccharide group, Macleaya cordata group and polysaccharide+Macleaya cordata group was significantly higher than that in control group (P<0.05). Compared with control group and antibiotic group, the number of Lactobacillus in jejunal contents of yellow-feathered broilers in polysaccharide group, Macleaya cordata group and polysaccharide+Macleaya cordata group was significantly increased (P<0.01). Macleaya cordata sanguinarine was medium sensitive to Staphylococcus aureus and Pasteurella, and was low sensitive to Escherichia coli and highly sensitive to Salmonella. MIC of Macleaya cordata sanguinarine on Staphylococcus aureus, Escherichia coli, Pasteurella and Salmonella were 25.00, 25.00, 12.50 and 1.56 μg/mL, respectively. In conclusion, dietary tea seed polysaccharide and Macleaya cordata alkaloids can both replace antibiotic, can effectively improve jejunal morphological structure, increase the number of Lactobacillus in jejunum of yellow-feathered broilers, and combined effects is equivalent to individual addition effects. Macleaya cordata sanguinarine has strong antibacterial effects on Salmonella, and the effects are superior to that of penicillin sodium.