Abstract: In order to optimize the fermentation process conditions of mulberry leaf tea and explore the changes in antioxidant activity of mulberry leaf tea after fermentation. This experiment used response surface analysis to determine the optimal process conditions for fermenting mulberry leaf tea under three variable conditions: Fermentation temperature, fermentation time, and inoculation amount of Aspergillus niger. The content of bioactive substances and antioxidant properties in fermented mulberry leaf tea were determinated. The findings indicated that 5 days and 16 hours of fermentation duration, 30 ℃ of fermentation temperature, and 16.55% of inoculation amount were the ideal process parameters, and that the fermented mulberry leaf tea scored as high as 90 on the sensory scale under these conditions. In addition, the content of total flavonoids, total polysaccharides, polyphenols, free amino acids, rutin, and quercetin in fermented mulberry leaf tea increased by 14.39%, 12.67%, 2.90%, 3.46%, 81.13%, and 19.35%, respectively, compared to mulberry leaf green tea. Tea brown pigment with a content of 7.55%±0.37% was also detected in fermented mulberry leaf tea. In addition, the scavenging rate of hydroxyl radicals, DPPH radicals, superoxide radicals and the chelating rate of divalent iron ions of fermented mulberry leaf tea at the concentration of 1/1000 (g/mL) were 1.16, 1.07, 1.06 and 0.82 times higher than that of mulberry leaf green tea, respectively. The results of this study provide theoretical basis and references for further development and application of mulberry leaf in the future.