Measurements of enzymes involved in alginate biosynthesis were straightforward in mucoid (alginate-positive) Azotobacter chroococcum ATCC 4412 crude extracts. At the stationary growth phase, where the production of the exopolysaccharide was greatest, the enzymes phosphomannose isomerase and GDP-mannose pyrophosphorylase increased markedly, whereas phosphomannomutase and GDP-mannose dehydrogenase kept the high activity levels measured in the acceleration growth phase. In nonmucoid (alginate-negative) A. chroococcum and A. vinelandii strains, the activities of phosphomannose isomerase and GDP-mannose pyrophosphorylase were rather low or, in some cases, undetectables. Except in A. chroococcum MCD1, which exhibited a low activity, phosphomanomutase was high in the nonmucoid Azotobacter strains, and GDP-mannose dehydrogenase reached a significant activity level in two out of four nonmucoid strains tested. The results suggest that derepression of phophomannose isomerase and GDP-mannose pyrophosphorylase is a sine qua non condition for alginate formation by A. chroococcum.