This study examined the chemical and structural properties of humic substances and microbiological activity in order to verify differences in carbon dynamics along soil profiles in two Vertisols (Typic Haploxerert and Xeric Epiaquert) and two Alfisols (Mollic Haploxeralf and Ultic Haploxeralf) developed under Mediterranean climate in Italy. Humification parameters, thermal methods, including differential scanning calorimetry (DSC) and thermogravimetry (TG), together with Fourier transform infrared (FT-IR) and fluorescence spectroscopies were used to characterize humic acids (HA). Microbiological activity of soils was assessed by basal respiration, metabolic quotient (qCO2) and Cmic:Corg ratio. FT-IR spectra and thermal analysis DSC/TG of HA extracted from the upper horizons showed a higher aliphatic character, whereas HA extracted from the lower horizons had a higher content of aromatic structures and polysaccharides. Moreover, the fluorescence index of the HA (HIXflu) showed a higher degree of aromatic polycondensation in the subsoils. The Cmic:Corg ratio was negatively correlated with the HIXflu of HA (P