Citric acid is a key biochemical in the emerging bioeconomy, to date mostly produced through fermentation by A. niger of molasses. One main issue is represented by the replacement of the molasses-based medium with less expensive and more sustainable feedstocks. Crude glycerol, a secondary product from the biodiesel industry was used without any pretreatment as substrate for fermentation by Yarrowia lipolytica DSM 8218 to produce it. The raw glycerol was retrieved from a local biodiesel production plant. The fermentation set-up was first optimized in flasks by using the Response Surface Methodology (RSM). A 3-level Full Factorial Design of Experiment was employed to investigate the impact of crude glycerol concentration and C/N ratio on citric acid yield and two N sources (ammonium sulphate and yeast extract) were alternatively tested. The optimized conditions were subsequently applied to a 2L bioreactor. The best conditions to maximize YCA were found to be: crude glycerol 33 g/L and C/N 141, which resulted in the production of 8 g/L of citric acid and in a metabolic yield of 0,27 g/g. Scale up in bioreactor indicated that the citric acid production like can be controlled and improved by adjusting the concentration of dissolved oxygen.