Dynamical changes of tea metabolites fermented by Aspergillus cristatus, Aspergillus neoniger and mixed fungi: A temporal clustering strategy for untargeted metabolomics

Dark tea fermentation involves various fungi, but studies focusing on the mixed fermentation in tea remain limited. This study investigated the influences of single and mixed fermentation on the dynamical alterations of tea metabolites. The differential metabolites between unfermented and fermented teas were determined using untargeted metabolomics. Dynamical changes in metabolites were explored by temporal clustering analysis. Results indicated that Aspergillus cristatus (AC) at 15 days, Aspergillus neoniger (AN) at 15 days, and mixed fungi (MF) at 15 days had respectively 68, 128 and 135 differential metabolites, compared with unfermentation (UF) at 15 days. Most of metabolites in the AN or MF group showed a down-regulated trend in cluster 1 and 2, whereas most of metabolites in the AC group showed an up-regulated trend in cluster 3 to 6. The three key metabolic pathways mainly composed of flavonoids and lipids included flavone and flavonol biosynthesis, glycerophospholipid metabolism and flavonoid biosynthesis. Based on the dynamical changes and metabolic pathways of the differential metabolites, AN showed a predominant status in MF compared with AC. Together, this study will advance the understanding of dynamic changes in tea fermentation and provide valuable insights into the processing and quality control of dark tea.