Absence of muscarine in the Entoloma rhodopolium complex in northern Japan

Entoloma rhodopolium (Fr.) P. Kumm. has long been recognized as a major cause of mushroom poisoning in Japan and has traditionally been regarded as a muscarine-containing species. However, direct chemical evidence for muscarine in this taxon remains limited, and recent molecular studies indicate that specimens identified as E. rhodopolium in Japan comprise multiple species within a complex, leaving the distribution of muscarine unclear. In this study, we examined muscarine occurrence in phylogenetically characterized specimens of the E. rhodopolium complex from northern Japan. Internal transcribed spacer (ITS) sequences from 52 specimens were analyzed with reference data to determine phylogenetic placement, resolving 14 lineages corresponding to ten described species and four unidentified lineages. Muscarine was quantified using a validated LC-MS/MS method with a detection limit of 0.05 μg/g. Muscarine was not detected in any specimen across all lineages. These results indicate that muscarine is unlikely to be widely distributed within the E. rhodopolium complex in Japan and suggest that other compounds may be responsible for the reported toxicity, warranting further integrative chemical and taxonomic studies.