Soil microorganisms have been shown to fix CO₂. Although this process plays only a minor quantitative role on a bulk soil basis, it is important in certain microenvironments of the soil and may change the assessment of data on soil organic matter turnover as determined by stable isotope techniques and on the formation of bound residues from radioactively labelled pollutants. To study this process, we incubated soil in the dark under a ¹³CO₂-enriched atmosphere and found a significant transfer of the label into soil organic matter (1.3 μmol C g⁻¹ soil after 61 days). Enrichment of the label in fatty acids (δ¹³C up to 1200‰) and amino acids (δ¹³C up to 200‰) showed that microbial biomass mediates the process. The data indicate that a wide range of autotrophic and heterotrophic microorganisms are involved and that anaplerotic reactions make a significant contribution. Part of the label appeared to have been already transformed to non-living soil organic matter, after lysis of the microbial cells.