Ganirelix, a peptide-based drug used to treat female infertility, has been in high market demand, which attracted generic formulation. A hitherto unknown impurity of ganirelix was observed in our formulation process, which reached ~0.3% in 6 months and led to a detailed investigation of its structure. In-depth analysis of ESI-MS/MS data of this impurity coupled with an artificial intelligence prediction tool led to a highly unusual putative structure, that is, N-(2-carboxyethyl)-ganirelix ( N CE-GA), which was authenticated by chemical synthesis from ganirelix and NMR analysis and via corroborated HPLC and MS/MS data with the formulation-derived impurity., (© 2023 European Peptide Society and John Wiley & Sons Ltd.)