As living standards rise, the overconsumption of sugary and calorific foods has led to a rise in obesity, diabetes, and other diseases. In response to the increasing demand for healthier diets, the food industry is actively seeking sugar alternatives. Among these alternatives, d-allulose as a functional sweetener has garnered significant attention for its low-calorie content, low glycemic index, and health benefits. This review summarizes recent advancements in d-allulose research, including its physiological functions, potential applications, and bioproduction methods. This review consolidates the known physiological functions of d-allulose and assesses its potential applications in the food and medical industries. Furthermore, the review explores recent progress in biotechnological production technologies, such as enzymatic conversion and microbial fermentation, which are key to producing d-allulose. d-Allulose is a standout natural sweetener with low calories and a low glycemic index, providing health benefits like lowering blood sugar and lipids, antioxidants, preventing obesity, and regulating metabolism. In the food industry, d-allulose is suitable for use in a variety of products, including baked goods, beverages, confectionery, and yogurt. The primary methods for its production are enzymatic conversion and microbial fermentation, both of which offer scalable and sustainable approaches. Recent research has advanced the production of d-allulose using low-cost raw materials, including agricultural and forestry waste, and even CO