Metal nanomaterials as mimic enzyme have been widely used for biosensor and catalysis due to their high stability and low cost. Here, [1-methyl-3-ethylimidazole][dicyandiamide] ionic liquid ([EMIM][DCA]) reacted with ruthenium chloride to form [EMIM]₃[Ru(DCA)₆] ionic liquid. Silk was dissolved in the above ionic liquid and then carbonized at high temperature to obtain Ru-biomass carbon composite. The results show that Ru-biomass carbon offers excellent dispersion, small Ru nanoparticles with an average diameter of 7.5 nm which were uniformly dispersed on the surface of carbon sheets, and high oxidase-like activity. Based on the inhibition of chlorpyrifos on catalytic activity of Ru-biomass carbon, a method was established for colorimetric determination of chlorpyrifos. Chlorpyrifos obviously inhibits the catalytic activity of Ru-biochar carbon, leading to a reduced absorbance of blue product. When the concentration of chlorpyrifos was in the range from 10 to 80 ng/mL, the peak absorbance decreases linearly with the increase of chlorpyrifos concentration, giving a detection limit of about 6.5 ng/mL. The as-proposed method provides higher sensitivity and stability compared with present methods, and has successfully applied to detection of chlorpyrifos in peach samples.