Equisetum palustre L. is known for its toxicity for livestock. Several studies in the past addressed the isolation and identification of the responsible alkaloids. So far, palustrine (1) and N-5-formylpalustrine (2) are known alkaloids of E. palustre. A HPLC-ESI-MS/MS method in combination with simple sample work-up was developed to identify and quantitate Equisetum alkaloids. Besides the two known alkaloids six related alkaloids were detected in different Equisetum samples. The structure of the alkaloid palustridiene (3) was derived by comprehensive 1D and 2D NMR experiments. N-5-Acetylpalustrine (4) was also thoroughly characterized by NMR for the first time. The structure of N-5-formylpalustridiene (5) is proposed based on mass spectrometry results.Twenty-two E. palustre samples were screened by a HPLC-ESI-MS/MS method after development of a simple sample work-up and in most cases the set of all eight alkaloids were detected in all parts of the plant. A high variability of the alkaloid content and distribution was found depending on plant organ, plant origin and season ranging from 88 to 597 mg/kg dried weight. However, palustrine (1) and the alkaloid palustridiene (3) always represented the main alkaloids. For the first time, a comprehensive identification, quantitation and distribution of Equisetum alkaloids was achieved. (C) 2015 Elsevier Ltd. All rights reserved.