Fatty acids were analyzed by gas chromatography using a DB-FFAP column of 30 m length and 0.25 mm inner diameter and a programmable temperature vaporizer injector. Following the trophic biomarker concept, the fatty acids 16:0, 20:5(n-3) and 22:6(n-3) were classified as typical components of biomembranes. High levels of 16:1(n−7) as well as 16:4(n−1) and 18:1(n−7) were used as diatom markers and 18:4(n−3) as fatty acid marker for dinoflagellates. The fatty acid 18:1(n-9) indicates carnivorous feeding. A carnivory index was applied based on the fatty acid ratio 18:1(n−9) / [16:1(n−7) + 16:4(n−1) + 18:1(n−7) + 18:4(n−3) + 18:1(n−9)] to reflect the proportion of carnivorous compared to herbivorous feeding in an organism. Fatty acid compositions of zooplankton and fish were taxon-specific and did not depend on sampling area or depth. Most species showed a dominance of typical membrane fatty acids, e.g., 16:0, 20:5(n-3) and 22:6(n-3). The dominant copepod Calanus chilensis had a low carnivory index and elevated amounts of diatom fatty acid markers which point to a predominantly herbivorous feeding. Among the krill species, Euphausia mucronata had the lowest carnivory index compared to the other euphausiids indicating a more herbivorous feeding. The squat lobster Pleuroncodes monodon had a significantly lower carnivory ratio compared to the deep-sea decapods Gennadas sp. and Acanthephyra sp. emphasizing its different trophic role compared to other decapods.