Abstract: An experiment was conducted to study the effects of original zymolytic conditions on nitrogen loss of brewer's spent grains-based bio-organic fertilizer by using the quadratic uniform-precision rotatable central composite design including four factors and five levels. The result shows that the order of each factor's influence to nitrogen loss is: pH value > inoculation fungal biomass > ratio of straw to lee > water content. There are positive interactions between the four factors and the interactions between pH value and water content and between inoculation fungal biomass and ratio of straw to lee are not significantly different. In this experiment, the suitable ranges of pH value, inoculation fungal biomass, water content and ratio of straw to lee are: 6.398-6.4186, 0.3991%-0.4031%, 64.7014%-64.8285% and 40.9326%-41.0956%, respectively.