As a protein resource, only a little of cottonseed meal was used in feed industry due to the presence of toxin, gossypol. To obtain strains for gossypol detoxification, 16 soil samples were collected from China and 144 strains were isolated. Among them, a strain (Y-2) possess gossypol detoxification markedly. The strain Y-2 was identified as Pichia guilliermondii by traditional and molecular genetic identification. This strain was non-pathogenic yeast, and was first reported used on degradation of gossypol. Mutant YUV-51 with the highest detoxification was obtained by UV mutation. The detoxification rate of gossypol was up to 58% under optimized culture conditions: inoculation of 0.025g wet cell/g cottonseed meal, 30 °C, initial moisture content of solid substrate 50%, 48 h. Moreover, to avoid degradation of a large part of free gossypol before the fermentation, a great lot of energy could be saved with no heat-moisture treatment.