Organic matter extracts from the 01, 02 and Al horizons of a hardwood forest rapidly incorporated [ 35S]sulphate into organic sulphur components larger than 12,000 dallons in molecular weight. In the 02 layer extract, the 35S-label was incorporated into amino acid-S, sulphonate-S and ester sulphate linkages. This process was stimulated by adenosine 5'-triphosphate (ATP), cellulose, cellobiose, glucose, succinate and pyruvate. The involvement of ATP sulphurylase, elaborated by bacteria present in this extract, is suggested by observations that incorporation was also stimulated by Mg 2+ and inhibited by erythromycin, tetracycline, sodium azide, selenate, molybdate and chlorate. 35S-labelled adenosine 5'-phosphosulphate (APS) or 3'-phosphoadenosine-5'-phosphosulphate (PAPS) also served as S donors for organic-S formation but the rate of organic-S formation was much lower than that observed when [ 35S]sulphate was the S donor.