There is a great need to understand how resource interactions alter the functioning of ecosystems, where the selective elimination of pelagic fishes can lead to changes in food web structure. This work analyzes the trophic niches of three species of commercial importance in the Galapagos Marine Reserve, yellowfin tuna, Thunnus albacares (TA), skipjack tuna, Katsuwonus pelamis (KP), and wahoo, Acanthocybium solandri (AS), via multiple analyses. According to the prey-specific index of relative importance, the most important prey for TA was Dosidicus gigas, while for AS it was unidentified pelagic fish. Interspecific differences were found between the isotopic signatures of AS and those of TA and KP. The isotope mixing model provides evidence for some predominance of D. gigas in the diets of TA and KP, while the fishes Selar crumenophthalmus and Paranthias colonus contribute to the diet of AS. The stable isotope Bayesian ellipses show a high overlap between TA and KP, suggesting a similar use of resources and feeding areas, while the ellipse of AS does not overlap with that of the other species. Both AS and TA were present around the islands more commonly during the day, with a peak in detections in the morning for AS and a greater presence of TA throughout the afternoon; there were only a few detections of KP in the days immediately after tagging. In summary, the results of this study suggest a pelagic foraging strategy with differential consumption of prey between AS and the other species.