Modelling soil organic matter dynamics requires reproducible and accurate data from several methods that follow such evolution based on changes in soil organic matter properties. The objective of this study is to investigate changes in the chemical, thermal and biological properties of soil organic matter after afforestation using emerging methods such as thermal analysis, isothermal calorimetry, and 13C CPMAS NMR. These methods were applied to a chronosequence of soils where large losses of carbon have occurred in the 29 years since afforestation. Results show that over this period the soil organic matter becomes more aromatic, resulting in increased thermal stability and decreased microbial activity. Over longer time frames, between 29 and 40 years after afforestation, soil organic matter increased, mainly in the aliphatic and carbohydrate fractions with enhanced thermal stability and consequent metabolic changes from microbial adaptation to the new organic matter.