Untargeted metabolomics-based identification of insomnia-ameliorating compounds from five mushrooms

Insomnia, a common sleep disorder in the general population, is defined by difficulties in fall asleep, staying asleep, or nonrefreshing sleep. Five edible Polyporales mushrooms, namely Amauroderma rugosum (AR), Ganoderma lucidum (GL), G. resinaceum (GR), G. sinense (GS) and Trametes versicolor (TV), have been widely used in China for managing insomnia. However, their active components for this application are not fully understood, restricting their universal recognition. To identify sedative-hypnotic compounds shared by the mentioned five Polyporales mushrooms using untargeted metabolomics.

UPLC-Q-TOF-MS/MS was used to establish chemical profiles for the ethanolic extracts of six batches of each mushroom species. The chemoprofile raw data were processed using MS-DIAL software to extract shared compounds of the five mushrooms. Differential content compounds [differential metabolites; VIP (variable importance in the projection) ≥ 2] in the shared compounds were identified using OPLS-DA (orthogonal projection of potential structure discriminant) analysis. A pentobarbital sodium (i.p. 45 mg/kg) induced sleeping mouse model was used to assess the sedative-hypnotic effects of the five mushrooms. The differential metabolites contributing to sedative-hypnotic effects were identified as sedative-hypnotic compounds (VIP ≥ 1.5; Correlation coefficient > 0) by OPLS (orthogonal projections to latent structures) analysis. The sedative-hypnotic effects of the identified compounds were validated in the mentioned mouse model.

Our results showed that a total of 1,860 compounds shared by the five mushrooms were extracted. Ninety-two common differential metabolites were identified. Daily i.g. administration of 0.3 g/kg of the ethanolic extract of AR, GL, GR, GS, or TV for eight consecutive days shortened sleep latency and prolonged sleep duration of model mice, with different potencies. Six triterpenes, four ganoderic acids (B, C1, F and H) and two ganoderenic acids (A and D), were found to be the sedative-hypnotic compounds shared by the five mushrooms. Each of the six triterpenes (i.g. 15, 30 mg/kg; Once) dose-dependently shortened sleep latency and prolonged sleep duration in mice.

To summarize, we for the first time found that the six identified triterpenes contribute to the sedative-hypnotic effects of the five Polyporales mushrooms. Our novel findings provide pharmacological and chemical justifications for the use of the five edible mushrooms in managing insomnia.