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This research aimed at optimizing the combination of sugar addition (%w/v) and yeast concentration (%v/v) on alcohol fermentation stage in the production of red dragon fruit vinegar. In this study, 8%(w/v) was used as the lower limit and 12%(w/v) was used as the upper limit for sugar addition. While 0.2%(v/v) used as the lower limit and 0.8%(v/v) was used as the upper limit for yeast concentration. The results from alcohol fermentation process, in which the alcohol concentration ranged from 4 to 14%(v/v) was then used as the response and added to Response Surface Methodology (RSM) for obtaining the optimal solution. The results showed that the optimal solution for sugar addition was 12%(w/v) and yeast concentration was 0,8%(v/v) producing alcohol with concentration of 7.956%(v/v) and the accuracy of 94.51% based on RSM program. The characteristics of red dragon fruit vinegar from the optimal solution was as follows: final alcohol concentration of 1.12%(v/v); acetic acid concentration of 3.43%; pH of 3.5; total soluble solid (TSS) of 3.4ºBrix; IC50 of 20.2665 mg/ml; and diameter of inhibition zone (antimicrobial) of 0.67 cm, respectively. However, the acetic acid of red dragon fruit vinegar was still lower than that of the standard and other commercial vinegar products. Yet, utilization of subgrade red dragon fruit as vinegar was found to increase the antioxidant activity by 20.34%.