Pretreatment of straw separated from cattle and horse manure using N-methylmorpholine oxide (NMMO) was investigated. The pretreatment conditions were for 5 h and 15 h at 120 C, and the effects were evaluated by batch digestion assays. Untreated cattle and horse manure, both mixed with straw, resulted in 0.250 and 0.279 Nm3 CH4/kgVS (volatile solids), respectively. Pretreatment with NMMO improved both the methane yield and the degradation rate of these substrates, and the effects were further amplified with more pretreatment time. Pretreatment for 15 h resulted in an increase of methane yield by 53% and 51% for cattle and horse manure, respectively. The specific rate constant, k0, was increased from 0.041 to 0.072 (d-1) for the cattle and from 0.071 to 0.086 (d-1) for the horse manure. Analysis of the pretreated straw shows that the structural lignin content decreased by approximately 10% for both samples and the carbohydrate content increased by 13% for the straw separated from the cattle and by 9% for that separated from the horse manure. The crystallinity of straw samples analyzed by FTIR show a decrease with increased time of NMMO pretreatment.