The fluorescence spectra of coupled bioluminescent enzyme system (NAD(P)H:FMN-oxidoreductase and luciferase) in the different viscosity and temperature above the optimum was investigated. It was shown that the increasing in viscosity leads to concentration quenching of the fluorescence intensities. The largest contribution to the fluorescence spectrum includes the «internal» tryptophan residues. The fluorescence spectra obtained for the luciferase and NAD(P)H:FMN-oxidoreductase was showed lack of conformational changes in the structure of proteins in the viscous medium simulated various concentrations of glycerol and sucrose at the extreme temperature.