Abstract: Herein, a L-histidine based copper doping carbon dots (Cu/His-CDs) is synthesized via a straightforward one-pot microwave digestion method. By systematically investigating the catalytic mechanism of Cu/His-CDs, it was found that Cu²⁺ initiates Fenton-like reaction-triggered peroxidase (POD)-like catalytic activity, effectively oxidize colorless 3,3′,5,5′-tetramethylbenzidine (TMB) to blue oxTMB by converting hydrogen peroxide (H₂O₂) and O₂ into active oxygen (·OH, ¹O₂, ·O₂⁻). Based on the cascade reaction of glucose oxidase (GOx) and Cu/His-CDs nanozyme, a sensitive colorimetric assaying platform for glucose detection was established with a good linear range of 10 −1000 µmol/L and a calculated detection limit of 1.72 µmol/L. The method was used for the determination of glucose in cigarette samples and exhibited satisfactory spiked recoveries and acceptable relative standard deviation (RSD), and the method is characterized by high selectivity and sensitivity with good accuracy.