The broken efficiency of cell wall and the release amount of Pichia pastoris intracellular protein under different cell breaking conditions were investigated in this paper. The results showed that broken efficiency using hot alkali combined with high-pressure homogenizing method was higher than that of enzyme hydrolysis, hot alkali treatment and high-pressure homogenation, respectively. Suspended medium had little effect on the broken efficiency of yeast cell, but had significant effect on the protein release yield. The results indicated that optimal condition for intracellular proteins extraction was 30% (wet weight, w/v) of yeast cells suspend in 50 mM phosphate buffer (pH 10.0), water bathed at 60?C for 2 hours, homogenized twice at 100 MPa pressure. The broken efficiency of Pichia pastoris cell could reach 87.6% and the protein yield was 35.48 g per 100 g cells.