Polycyclic aromatic hydrocarbons (PAHs) in crude oil have been shown to impair cardiac function. Since mitochondrial function is critical to support aerobic metabolism through the production of ATP, abnormal mitochondrial bioenergetics could be the basis for the impaired cardiac function associated with crude oil exposure. We hypothesized that cardiac mitochondrial function would be reduced after 24h of crude oil exposure. Sub-adult (0.95 ± 0.11 kg, n = 16) Mahi-mahi (Coryphaena hippurus) were kept in 300L static tanks of 35 ppt seawater at 26 °C for 24h in control conditions or 10% crude oil solution. Permeabilized cardiac muscle fibers were isolated from ventricle tissue for study. A portion of the tissue was flash frozen and used in mitochondrial density and enzyme activity analyses. No statistical differences were observed in mitochondrial respiration rate or respiratory flux control ratios. This suggests that 24h of oil exposure has no impact on the electron transport system. In addition, no changes were observed in mitochondrial density, enzymatic activity, or Vmax between treatments. However, oil-exposed mahi had reduced V̇O2 at lower ADP concentrations and the apparent Km ADP was 3-fold higher in oil-exposed mahi. The greater need for ADP in oil-exposed mahi was not explained by changes in respiration rate, mitochondrial density, or enzyme activity. We speculate that the increased need for ADP is potentially due to a weakening mitochondrial membrane or allocation to cellular processes.