In this study, Biceps femoris (BF) and Triceps brachii (TB) of Aberdeen Angus steers, from the hindquarter and forequarter, respectively, were compared from their fatty acid composition, lipid health, and lipid enzyme activity indices points of view. For this, ten animals were produced in a pasture system and ten were finished in a concentrate-based system. TB presented a significantly higher intramuscular fat, saturated C14:0, C14:1, C16:1, CLA, and MUFA content. BF presented a significantly higher LA, ARA, EPA, DPA, DHA, PUFA, total n-6, and n-3 content, and a significantly higher PUFA/SFA ratio. Significant differences between muscles were found regarding lipid enzyme activity indices, but not concerning atherogenic and thrombogenic indices. Also, meat from different feeding systems was compared, where meat from pasture presented a better fatty acid composition regarding cardiovascular health aspects. In conclusion, BF presented a better composition in the most nutritionally relevant fatty acids, with exception of CLA.