In current research the optimization of culture conditions Azotobacter vinelandii D-08 on nutrient mediums with different content of molasses – 20, 30, 40 and 50% (by weight) was conducted. It is established that the maximum accumulation of polysaccharides was 13,78 g/l and it was observed at growth of the strain on the medium containing 50% molasses for 72 hours. The results of the method of gel permeation chromatography showed that the obtained polysaccharide has a molecular weight of 0.93532 kDa and 312.5014 kDa.