There is no much information on the rate and dynamic of carbon accumulation in the soil under eucalypt plantations in Brazil. Conditions which favor eucalypt fast growth may, also, contribute to rapid decomposition of the soil organic matter (SOM) and plant residues left in the area. This study aimed to: determine SOM stock and quality under natural savanna vegetation, pasture, and eucalypt plantations; and evaluate changes in SOM as affected by the number of eucalypt rotations. Soil samples were collected under Eucalyptus grandis plantations and in nearby areas of natural savanna, and cultivated pasture, in Itamarandiba county, Minas Gerais state, Brazil. The content of soil total organic carbon (TOC) under eucalypt plantation, 20 year old, was similar to those under natural savanna and pasture. Eucalypt cultivation did not affected C and N stocks in the microbial biomass but contributed with larger quantities of light fractions of SOM, leading to an increase of SOM as compared with the other two vegetation types. However, there was a change in the carbon type, as 20 years of eucalypt cultivation lead to a reduction in C-C4 at a rate of 3 ‰ per year. SOM from the savanna area showed a higher stage of humification as compared with eucalypts and pasture, as indicated by FTIR analyses. Therefore, replacing the natural savanna vegetation by eucalypt plantations did not impair the stocks and fractions of SOM. The humic acid fraction (HA) from the pasture, and the fulvic acid fraction (FA) from eucalypt plantation were more resistant to thermodegradation as compared with the other two vegetation types. The hydrophobicity index of the AH fraction from the pasture was 0.83, from the savanna 0.38, and from eucalypts 0.29, indicating the pasture AH higher recalcitrancy in comparison with savanna and eucalypts. The same order was observed for the FA fraction which showed indexes of 1.43, 0.92, and 0.50, respectively. Successive rotations of eucalypts lead to an increase in the stock of TOC, specially in areas previously used with pasture. However, soil total nitrogen stock reduced as the time of eucalypt cultivation increased. The amount of humic substances increased with the amount of litterfall. Fine roots growing in the forest floor contributed to increase the amount of SOM, light fractions of the SOM, and carbon in the microbial biomass. Therefore, eucalypt plantations, which replaced natural savanna vegetation, increased SOM, being the carbon stock dependent on the productivity of the eucalypt stands.