Vinasse, residue from the sugar-energy sector, has been applied as a fertilizer in sugarcane fields due to its nutritional characteristics, which may reduce nitrogen fertilization, especially if associated with diazotrophic bacteria. The objective was to evaluate the use of vinasse as culture medium for Azospirillum brasilense and the chemical changes caused in the residue after bacterial colonization. The experimental design was completely randomized with 9 treatments: T0-control (nutrient broth DYGs inoculated with A. brasilense); T1- 100% vinasse inoculated with A. brasilense; T2- 75% vinasse inoculated with A. brasilense; T3- 25% vinasse inoculated with A. brasilense; T4- 50% vinasse inoculated with A. brasilense; T5- 100% vinasse; T6- 75% vinasse; T7- 25% vinasse and T8- 50% vinasse. Microbial growth was evaluated in a spectrophotometer, by turbidimetry (700nm). Chemical characterization of vinasse was carried out before bacterial inoculation and on days 1, 3, 5 and 7 after inoculation according to the methodologies of the Standard Methods for the Examination of Water and Wastewater 22 ND for the elements nitrogen (4500-Norg), phosphorus (4500-P), potassium (3030-A) and calcium (3500 Ca-B). The determination of organic matter was carried out in accordance with the technical standard NBR 13600. The bacteria A. brasilense survived and multiplied in sugarcane vinasse, causing changes such as the increase in the levels of phosphorus and nitrogen and the reduction of the organic load, mainly in pure vinasse (100%), indicating that the viability of this practice for the application of these organisms in the culture of sugar cane via fertigation.