Protease is used in protein digestion. Trypsin, one of the most common proteases, cleaves exclusively at C-terminus of amino acid Lysine and Arginine in proteins. Moreover, organic solvents are often added for the trypsin digestion to modify native proteins to denatured proteins, and this tendency makes it effective to digest proteins. In this study, we investigate the digestion efficiency of trypsin for the digestion of bovine serum albumin and horse skeletal muscle myoglobin. Sample solutions were prepared with different amounts (0 %, 10 %, and 20 %) of acetonitrile and digested using trypsin with a help of microwave irradiation. Digested peptides were analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and the sequence coverage and the intensity of the protein peak were used as indicators of trypsin activity.