Abstract: The azo-based chromogenic reagent 3,5-dicarboxylic acid-4'-hydroxyazobenzene was synthesized and its color reaction with copper(II)was also investigated. The amount of 3,5-dicarboxylic acid-4'-hydroxyazobenzene and the effect of sodium tetraborate buffer solution on the formation of complexes were determined by spectrophotometer. A new measured method for the micro copper(II)in tricholoma was established.The results showed that the mass concentration of copper(II)obeyed Bill's law in the range of 0~2.5 μg·mL^-1. The regression equation was Y=0.1696x+0.0237,R²=0.9921,and the molar absorptivity was 4.3×10⁴ L·mol^-1·cm^-1. Copper(II)complexes with 3,5-dicarboxylic acid-4'-hydroxyazobenzene formed a 1:1 complex. The recovery of standard addition was 97.0%~101.3% and RSD was 0.93%~1.50%. A new ultraviolet spectrophotometric method for the determination of copper ion in Tricholoma was developed. The method was simple and selective. The result was(5.95±0.49) mg·100 g^-1,which was almost the same as that measured by ICP spectrometer(5.89±0.09) mg·100 g^-1.