This dataset contains the values of natural abundance of stable carbon and nitrogen isotopes in bulk tissues, carbon values in total lipid extracts and lipid fractions, nitrogen values in amino acids, as well as the fatty acid profiles from the total lipid extracts and lipid fractions measured in 33 samples of 4 small pelagic fishes sampled during the PELACUS0322 cruise (NE Atlantic, April 2022). Six capture sites were distributed over 4 sampling zones in Galicia and in adjacent waters of the southern Bay of Biscay. Length, weight, sex and maturity stage data for each individual analyzed are also provided. Fishes were collected by midwater trawls of ca. 30 m² pelagic net equipped with a graded-mesh netting (starting with 30 mm and ending with 4 mm). The duration of each trawl was between 25 and 40 min. Different species occurred in the same trawls. Samples of muscle tissue were preserved freeze-dried. No treatments for lipid removal were used prior to carbon analyses. Lipids were extracted from freeze‐dried aliquots using a modified version of the Bligh & Dyer (1959) method. Neutral and polar lipid fractions from the total lipid extracts were separated by adsorption chromatography on a silica gel micro-column (100–200 mesh, 60 Å, pre-combusted silica gel) using solvents of different polarities. Stable isotope determinations (also carbon and nitrogen contents) were made by isotope-ratio mass spectrometry using an elemental analyzer (bulk samples and lipid extracts; Close et al. 2014; Bode et al. 2018) or a gas-chromatograph (derivatized amino acids; Bode et al. 2021) as the inlet device. Identification and quantification of fatty acids were made by gas-chromatography mass-spectrometry.