Recent approaches developed in metabolomics using liquid chromatography–tandem mass spectrometry (LC-MS/MS) enabled us to assign a part of specialized metabolites in plants. However, the approaches are not good enough for the rest of the metabolites, which are still unknown. To characterize the unknown metabolites, more appropriate and precise approaches need to be developed. Here, a procedure to analyze15N-labeled and nonlabeled LC-MS/MS data for identification of monoterpene indole alkaloids was developed. © 2022, The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.