Glycerol is a byproduct generated in large amounts during the process of biodiesel production. The present study reports an alternative method of crude glycerol valorization for the production of citric and malic acid by Yarrowia lipolytica. To enhance the conversion efficiency, we comprehensively investigated optimization of the carbon source, nitrogen source, and pH regulation strategy. Under optimal conditions with the initial concentration of crude glycerol at 40 g/L in combination with 0.3 g/L NH4Cl as the nitrogen source and pH at 4.0 using Na2CO3, we finally obtained 81.11 g/L citric acid and 24.90 g/L malic acid using fed-batch fermentation. In addition, acetate was found to enhance organic acid production and regulate carbon flow among organic acids from crude glycerol. The present study provides an alternative approach for valorization of crude glycerol into value-added biochemicals.