In civil engineering applications, alongside inherent resistance, the moisture content in wood is one of the most important factors affecting the durability of wood. The moisture content of wood is significantly influenced by the moisture diffusion coefficient. In this study, the impact of considering humidity fluctuations in a climatic chamber during the sorption process on the computational values of functional coefficients in various models of diffusion coefficient variability with material moisture content was analyzed. The dependence of the moisture diffusion coefficient on moisture concentration in wood was assumed in the form of constant, linear, quadratic, and exponential functions. The coefficients in the sought-after functions were found using the error function minimization method based on the kinetics of moisture sorption measurements in samples of Scots pine in the radial and tangential directions at an average air humidity in the chamber of 32%. As a result of the calculations, it was found that taking into account humidity fluctuations in the climatic chamber improved the fitting of computational curves to experimental curves in the case of constant and exponential models. The values of the diffusion coefficient assumed as constant calculated assuming variable humidity conditions in the chamber, differed on average by about 4% compared to the values of these coefficients obtained without considering air humidity fluctuations. In the case of each analyzed sample, the linear descriptions of the variability of the diffusion coefficient had a less steep course when considering air humidity fluctuations in the climatic chamber during the calculation process.