Influence of Fermentation by Four Fungi on Chemical Constituents and Pharmacological Activities of Panax notoginseng Leaves

In this study, Panax notoginseng leaves (PNL) were separately fermented by Rhizopus oryzae, Neurospora crassa, Monascus and Mucor rouxianus, and changes in the contents of total saponins, total polysaccharides, total phenolics, and total flavonoids, as well as in vitro antioxidant activity and α-glucosidase inhibitory activity of PNL during the fermentation process were assessed. Additionally, the metabolite profiles of raw and fermented PNL were analyzed by ultrahigh performance liquid chromatography-mass spectrometry (UPLC-MS/MS). The results indicated that the contents of total saponins and total polysaccharides in PNL were significantly reduced by fermentation with each of these four fungi. Interestingly, R. oryzae, N. crassa and Monascus were found to increase the content of total phenols, while Rhizopus oryzae and Mucor rouxianus increased the total flavonoid content. Fermentation for 3–5 days significantly enhanced the antioxidant activity of PNL, and the most pronounced effect was achieved with Monascus fermentation for three days, which increased the 2,2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid (ABTS) radical cation scavenging capacity and ferric reducing capacity by 14.30%, 5.13%, and 18.40%, respectively. Moreover, the α-glucosidase inhibitory activity of PNL initially decreased and then increased during fungal fermentation, which increased by 16.03% after Monascus fermentation for six days. By untargeted metabolomics analysis, 573 metabolites were identified from PNL. After three days of Monascus fermentation, the up-regulated metabolites were significantly less than the down-regulated ones, and the predominant up-regulated metabolites were amino acids, alkaloids, carbohydrates, lipids, phenols, flavonoids, and terpenoids. Furthermore, analysis of the 21 saponins showed that only ginsenoside F2 was significantly up-regulated, while eight other saponins were significantly down-regulated. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated that co-factor biosynthesis, flavonoid biosynthesis, purine metabolism, and pyrimidine metabolism were the most probable metabolic pathways during the fermentation process. In conclusion, fungal fermentation, especially Monascus fermentation, can effectively improve the functional activity of P. notoginseng by metabolic alterations through various pathways. Therefore, this study provides a scientific reference for the green processing of PNL.