Abstract: The dynamic changes of metabolites in H.marmoreus during different culture periods were studied by untargeted metabolomics industrially using cultivated H.marmoreus as experimental materials.The metabolites of H.marmoreus in different culture periods were analyzed throughout Venn analysis, partial least squares discriminant analysis, cluster analysis and pathway enrichment analysis.The results showed that a total of 3 205 metabolites were identified in H.marmoreus during different culture periods.Among them, there were 1 844 metabolites（87.4%） in H.marmoreus during different culture periods.Mainly included carboxylic acids and their derivatives, fatty acyls, organic oxygen compounds, prenol lipids.Herein Metabolic pathway enrichment analysis showed amino acid metabolism, carbohydrate metabolism, cofactor and vitamin metabolism, nucleotide metabolism, lipid metabolism and other pathways dominated.Compared with the control group（without inoculation）,121 differential metabolites were screened, belonging to 15 metabolic pathways.Among them, there were a series of chemicals showed striking features, for instance, lysine biosynthesis, cofactor biosynthesis, riboflavin metabolism, valine, leucine and isoleucine biosynthesis, pantothenic acid and coenzyme A biosynthesis, thiamine metabolism and other 6 metabolic pathways showed significant differences（P＜0.05）,mainly belong to amino acids and B vitamin metabolites, and apart from that, these metabolic pathways display have important effects on the formation of fruiting bodies, nutrients and umami of H.marmoreus.The metabolites of H.marmoreus showed significant differences in different cultivation periods, which tells the specific metabolites in the early stage of culture（Hm2） and fruiting stage（Hm5） changed greatly, while the metabolites in the middle stage of culture（Hm4） changed little compared with those two.This study revealed the dynamic changes of metabolites and the potential mechanism of biosynthesis during the cultivation of H.marmoreus from the perspective of metabolomics, and also provided a scientific basis for elucidating the metabolic regulation mechanism of H.marmoreus and optimizing H.marmoreus.