Glycosides were extracted from green tomatoes with ethanol as the solvent. The extracted glycosides were purified by an HPD macroporous resin and a silica gel column, crystallised and then recrystallised to yield white powder. The minimal inhibitory concentration (MIC) of the extracted tomato glycosides against Escherichia coli was determined to be 3.54 mg/mL through agar diffusion test. The effects of pH, temperature and reaction time on the anti-bacterial activity of the tomato glycosides were investigated. The bacteriostatic effect of the tomato glycosides was stronger than that of 1.5% sodium benzoate. The tomato glycosides exhibited the strongest anti-bacterial effect at pH 7.