The objective of this study was to evaluate the effect of virginiamycin (VM) combined with crude glycerin (CG) on dry matter intake (DMI, % BW), average daily gain (ADG), and fatty acid (FA) profile (g/100 of FA) of Longissimus dorsi muscle of Nellore bulls. Forty-eight bulls with initial BW (408.4 ± 22.2 kg) were individually fed with feedlot finishing diets without virginiamycin (VM-) or with 25 mg/kg of VM in DM (VM+) combined with diets without crude glycerin (CG-) or with 100 g/kg of CG (80% glycerol) in DM (CG+). The sugar cane bagasse was used as the exclusive roughage in the proportion of 20% in DM of diet and crude glycerin replace corn in the diet formulation. After 81 d of feeding bulls were weighed and slaughtered. After 24 h of cooling carcasses, samples of longissimus muscle (LM) between the 12th and 13th ribs were taken and freeze-dried. The fatty acids were quantified by gas chromatography. The data of animal performance and fatty acid profile were analyzed as a completely block randomized design arranged in a 2 × 2 factorial, virginiamycin and crude glycerin. Statistical model included the fixed effect of virginiamycin (1 degree of freedom, DF), crude glycerin (1DF) and all interactions. Random effects were block (1DF) and residual error. The data were analyzed using PROC MIXED of SAS. There were no interactions for the variables evaluated (P > 0.05). The DMI, % BW was greater for CG+ diets than CG-, independent of VM addition (1.95 v 2.06%; P < 0.05). ADG over 81 d was similar (1.30 kg/d) for CG or VM diets (P < 0.05). The G: F were statistically similar (0.12) between diets (P > 0.05). Total saturated fatty acids (SFA) in LM decreased in CG+ than CG- diets (45.64 vs. 42.64%; P < 0.05. Total monounsaturated fatty acids (MUFA) was greater in CG+ than CG- (42.57 vs. 40.33%; P < 0.05). Total unsaturated fatty acids (UFA) was greater in VM+ than VM- diets (50.28 vs. 50.38%; P < 0.05) and n-6/n-3 ratio did not change with CG (P > 0.05) or VM inclusion (P > 0.05). CG+ diets had greater UFA/SFA than CG- (1.05 vs. 1.15; P < 0.05). CG inclusion improves animal performance and it can be used to replace VM in finishing diets of Nellore bulls.