Significant benefit of soil organic matter (SOM) to crop productivity is scientifically well documented. The main constituents and active fractions of SOM are humic substances (HS) and phenolic compounds. Since both these two components strongly impact plant–soil relationship, it is importantly from an ecological point of view to discriminate their biological effects and relating them to their composition. In this study we compared the biological effects of HS, and the soil water soluble phenols (SWSP) on growth, antioxidant activities, carbohydrates, proteins, phenols, and vitamins of Pinus laricio callus. Each extract was assessed for the content of low molecular weight organic acids, soluble carbohydrates, fatty acids, and phenolic acids. Moreover, Fourier transform infrared (FT-IR) and surface-enhanced Raman scattering (SERS) spectroscopies were applied to study their molecular structure. The results showed that HS produced better callus growth compared to the control and SWSP. Carbohydrates decreased in presence of HS while proteins, vitamin C and E increased. In contrast, in callus treated with SWSP the amount of glucose and fructose increased as well as all the antioxidant activities. The data evidenced that HS rich in tartaric and fatty acids had beneficial effects on callus growth contrary to soil water-soluble phenols rich in aldehydes, and syringic, ferulic, and benzoic acids.