This paper describes thermal denaturation and trypsin digestion of protein on a microchip as an alternative application of a temperature controllable microchip. Analysis of the protein of small volume and low concentration, which is impossible in macro scale, could be possible with the fairly reduced process time using the proposed temperature controllable microchip. We optimized the parameters concerning thermal denaturation on a microchip such as thermal denaturation temperature, thermal denaturation time, digestion time and concentration of protein using BSA(bovine serum albumin) as a reference sample. Then we applied the optimized parameters to the other proteins (ovalbumin, myoglobin, hemoglobin, cytochrome C, Ubiquitin). The proposed method on a microchip in this paper needed an even shorter reaction time, smaller volume of sample and smaller concentration of sample compared to the previously presented marco scale thermal denaturation and trypsin digestion method. We could successfully acquire the thermally denatured protein in 1 minute at 85°C and the digested peptides in 10 minutes at 37°C with 3 μl/0.2 μM protein. The acquired average sequence coverages are range from 24 to 57% for the test proteins, which are sufficient for the protein identification in practical use.