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• Insects are proposed in Europe as protein sources for feed and food application. • Methods for insect-containing ingredient authentication needed. • Marker peptides were identified for Lesser Mealworm and Black Soldier Fly species. • Three species- specific markers were identified as most suitable for authentication by untargeted and targeted LC/MS methods. • The species-specific marker peptides were detected in insect-enriched (down to 1%) feed formulations. Insects have been proposed as new source of proteins to meet the growing demand connected to the increasing world population. In the EU the inclusion of insect proteins in feed and food is strictly regulated. Hence, analytical methods able to discriminate and identify different insect species in food and feed are a necessity. In this work, a peptidomic approach was applied to determine peptide biomarkers for two edible insect species: lesser mealworm and black soldier fly. Three species specific peptide biomarkers were identified using LC-MS/MS. The two insects were mixed with fish standard feed at different concentrations, to evaluate the feasibility of their use as markers in complex matrices. The detection of marker peptides was confirmed down to 1% insect amount. The data here reported constitutes the first proof of concept for the potential application of the peptide marker approach for the identification and quantification of insect ingredients. [ABSTRACT FROM AUTHOR]