Abstract: To improve the quality of orah fruit wine, Saccharomyces cerevisiae, Pichia manshurica and Lactobacillus plantarum were used in the single and mixed microbial fermentation of orah fruit wine. The physical and chemical indexes in the fermentation process were detected, and the flavor substances produced in the fermentation process were analyzed and compared. The results showed that the mixed fermentation group of S. cerevisiae, P. manshurica and L. plantarum (Zp group) had the highest sugar utilization rate and the obvious advantage of reducing the organic acids, in which the reducing sugar content was 2.54±0.02 g/L, and the total acid was 5.34±0.28 g/L. Compared with other fermentation groups, Zp group had the strongest antioxidant activity, in which the total phenol content was 279.82±1.36 mg/L, the superoxide anion content was 71.23±0.65 μmol/mL, and the scavenging rate of hydroxyl radical was 70.75%±0.17%. Liquid chromatography-mass spectrometry (LC-MS) was used to determine the organic acids of the fermentation groups. The results showed that lactic acid and acetic acid were the main organic acids produced during the fermentation groups. In particular, the relative content of total organic acids in Zp group was 7.9% lower than that in other fermentation groups, indicating that L. plantarum had good biological deacidification. Gas chromatography-mass spectrometry (GC-MS) was used to determine the aroma components of the fermentation groups, and a total of 68 volatile components were detected. Compared with the single bacteria fermentation group, the mixed microbial fermentation group of S. cerevisiae and P. manshurica (Sk group) and Zp group increased the types and contents of esters, and higher alcohols, which increased the aroma complexity of orah fruit wine. Through sensory evaluation, the results revealed that Zp group scored the highest in taste, aroma, and color, with a total score of 88.55±1.2. Zp group had moderate sweet and sour, and bright colors, which indicated that mixed microbial fermentation was helpful to improve the taste, enrich the aroma complexity, and enhance the quality of orah fruit wine. In this study, S. cerevisiae, P. manshurica, and L. plantarum were used for the first time to ferment orah fruit wine, which would provide a theoretical basis for quality control and product development of orah fruit wine.