Long range atmospheric transport is known to facilitate the poleward movement of persistent and semi-volatile chemicals. Their bioaccumulation and biomagnification along Antarctic food chains has been evidenced, including detection in an Antarctic keystone species, Antarctic krill, as well as in krill consumers such as Southern Hemisphere humpback whales. Southern Hemisphere humpback whales spend the austral summer feeding in the Southern Ocean before undertaking one of the longest known mammalian migrations between Antarctic feeding grounds and equatorial breeding grounds. During this time, these mammals undergo voluntary fasting and convert their lipid stores to energy, which inadvertently results in mobilisation of lipophilic chemical burdens from liberated fat stores. Mitochondria play a key role in lipid metabolism, breaking down lipids to power cellular processes. Any disruption to mitochondrial function is expected to influence whole-organism bioenergetics. The toxicological impact of accumulated chemical burdens to humpback whale bioenergetics is unknown. Ethical and logistical constraints prohibit in vivo toxicological investigation on cetaceans. An important focus of in vitro cetacean studies in recent years has involved cell culture. The seahorse analyser is frequently used for measuring mitochondrial function in live cells. By real time measurements of oxygen consumption rate and extracellular acidification rate the assay provides insight into the cause of cellular bioenergetic disruption. This study exposed Southern Hemisphere humpback whale immortalised skin fibroblast cells to four priority chemicals (chlorpyrifos, dieldrin, trifluralin and endosulfan), as determined by their novelty and prevalence in the Antarctic ecosystem. Cells were analysed with the Seahorse assay to quantify chemical effects on mitochondrial function as well as bioenergetics pathways. These findings contribute the first data regarding chemical impacts to bioenergetic function of humpback whale cells, and provide unique insight into contaminant mechanisms of action, as well as taxa-specific chemical effect data in support of evidence-based conservation policy and management.