Abstract: High-concentration anaerobic fermentation of cow manure was investigated using an anaerobic batch reactors under laboratory conditions at 20℃. In this present study, acetic acid content was used as response function, and three factors, namely, temperature, liquor concentration and retention time, were selected as input variables in the process of hydrolysis and acidogenesis of cow manure using a central composite rotatable orthogonal experimental design. The ranges of temperature, liquor concentration and retention time were 25～35℃, 6%～10% and 2～12 days, respectively. In the quadratic nonlinear model, the principal and secondary order of every parameter was analyzed with single-parameter method. The optimal experimental extent of every parameter was found by frequency analysis. Experimental results show that acetic acid content is greatly influenced by retention time, temperature was second on it, and liquor concentration was the least. Acetic acid content was more than 66% under the condition of 26～30℃, concentration 7%～9% and retention time 2～5 days. And acetic acid content was less than 60% under the condition of 26～30℃, concentration 10% and retention time 10～12 days.