As concerns about sugar-sweetened beverages are increasing, interest in natural sweeteners with zero calories is also growing. This study explores the health impacts of rebaudioside D and M, emerging sugar substitutes, compared to traditional alternatives and stevioside. In mouse models, we investigated the effects of rebaudioside D and M on obesity, liver health, thermogenic adipocytes, skeletal muscle, and gut microbiota. Results show no weight gain or exacerbation of obesity, no promotion of hepatic steatosis, no impairment of brown adipose tissue function and no changes in skeletal muscle metabolic proteins with either rebaudioside. Importantly, both of these sugar substitutes positively influence gut microbiota without adverse metabolic effects, suggesting their potential as safe sugar alternatives.