The ensiling of sugarcane induces loss of energy by alcoholic fermentation, and the addition of crude glycerin may improve the nutritional value of this forage. The objective of this work was to evaluate sugarcane silage with different concentrations of glycerin and the response of different starter cultures to the changes in microbial diversity, products of fermentation, and aerobic stability. The glycerin treatments increased dry matter (DM) and water-soluble carbohydrates (WSC) and reduced the neutral detergent fiber (NDF) concentration. The presence of glycerin resulted in a linear effect (p < 0.01) for DM, WSC, NDF, and DM loss. The silage inoculated with Lactobacillus hilgardii UFLA SIL52 showed a decrease in DM loss (p < 0.01). The glycerin treatments reduced the production of lactic acid (p = 0.05) and tended to decrease ethanol (p = 0.06) concentration. The dosage of glycerin did not affect aerobic stability (p = 0.28). Addition of glycerin to sugarcane silage improved chemical characteristics, decreased the ethanol content, and changed the microbial diversity.