Docosahexaenoic acid (DHA) and their CYP-derived metabolites, epoxydocosapentaenoic acids (EDPs), are important fatty acids obtained from dietary sources. While it is known that they have significant biological effects, which can differ between cell type and disease state, our understanding of how they work remains limited. Previously, we demonstrated that DHA and 19,20-EDP triggered pronounced cytotoxicity in H9c2 cells correlating with increased ceramide production. In this study, we examine whether DHA- and 19,20-EDP-induced cell death depends on the type of metabolism (glycolysis or OXPHOS). We cultivated H9c2 cells in distinct conditions that result in either glycolytic or oxidative metabolism. Our major findings suggest that DHA and its epoxy metabolite, 19,20-EDP, trigger cytotoxic effects toward H9c2 cells with a glycolytic metabolic profile. Cell death occurred through a mechanism involving activation of a lysosomal-proteolytic degradation pathway. Importantly, accumulation of ceramide played a critical role in the susceptibility of glycolytic H9c2 cells to cytotoxicity. Furthermore, our data suggest that an alteration in the cellular metabolic profile is a major factor determining the type and magnitude of cellular toxic response. Together, the novelty of this study demonstrates that DHA and 19,20-EDP induce cell death in H9c2 cells with a glycolytic metabolicwct 2 profile through a lysosomal-proteolytic mechanism.