The feasibility of using pyrene as a fluorescent probe in complexes cyclodextrin-pyrene-analyte in aqueous solutions was studied for several analytes (A) of different polarity. Bands that prove the formation of the complex Py₂βCD were found for the first time in the UV-VIS spectrum of an aqueous solution of pyrene in the presence of β-cyclodextrin. A characteristic of a fluorescent probe is the ratio I ₃/I ₁ in the fluorescence spectrum, where I ₃ is the intensity of the third vibronic line at ∼800 cm⁻¹, and I ₁ is the intensity of the 0-0 transition. The value of I ₃/I ₁ for Py₂βCD is highly sensitive to the addition of an analyte with a concentration of several μM/L to the aqueous solution. Dependences of (I ₃/I ₁)/C on the function of the dielectric constant ɛ of the analyte fɛ= (ɛ − 1)/(2ɛ + 1) are divided into two straight lines with different slopes for nonpolar and polar solvents. Energies of insertion of analytes into Py₂βCD were calculated by the quantumchemical PM3 method. An increase in (I ₃/I ₁)/C corresponds to the more thermodynamically stable complex 2APy₂βCD.