Esterification of inorganic phosphate by mitochondria-free extracts from pea cotyledons was shown to be stimulated by D-fructose-1: 6-diphosphate, and in the presence of NaF, though not in its absence, to be dependent on the addition of acetaldehyde. The rate of this glycolytic phosphorylation was determined by the level of both diphosphopyridine nucleotide and adenosine triphosphate (or adeno-sine diphosphate) in the extracts. Adenosine triphosphate added alone (2 mM.) inhibited phosphorylation, but this inhibition was reversed by diphosphopyridine nucleotide though not by reduced glutathione or cysteine. Adenosine triphosphate inhibited glycolytic phosphorylation in extracts from pentan-1-ol treated seeds to a greater extent than in extracts from normal seeds; this effect has been shown to be due to the lower content of diphosphopyridine nucleotide in these seeds compared with normal seeds.