Taking synthetic wastewaters as materials, the effects of micro-aerobic pretreatment on anaerobic digestion of organic waste for methane production were investigated in this paper by experiments. The results indicated that micro-aerobic pretreatment before anaerobic digestion could stimulate the activity of the hydrolysis bacteria, thus enhance the acidification of organic substrate and methane fermentation. Methane production was increased by 28% when the wastewater was pretreated for 4 hours under micro-aerobic condition and the maximal methane production rate was also increased by 57.5%. However when the pretreated time was 10 hours, methane production bacteria (MPB) could be poisoned and methane production was greatly decreased. Methane production could not also be increased evidently if the pretreated time was too short. The optimal pretreatment time was 4–6 h. So the micro-aerobic pretreatment process can stimulate the acidification of organic substrate and it may be applied in the anaerobic digestion of complex wastes such as kitchen wastes if the pretreatment time is well controlled.