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Natural polysaccharide is a type of functional carbohydrate polymer under heated investigation. The structure of natural polysaccharides is in close relationship with their actual biological functions such as antioxidant, immunoregulation and antitumor effects. Herein, we have identified numerous factors that significantly affect the biological functions of polysaccharides, including molecular weight, degree of polymerization, hydrophilicity and metabolic capability. Mitochondria, as a key polysaccharide-targeting organelle, undergo a number of alterations during the treatment, which includes morphological changes, ROS production, membrane potential and permeability, ETC activity and ATP production. Intriguingly, polysaccharides exhibit protective effects and maintain mitochondrial function on normal cells, while in cancer cells, polysaccharides tend to accelerate apoptosis and exacerbate mitochondrial dysfunction. Numerous signaling pathways, including antioxidant signaling pathway, anti-inflammatory signaling pathway and apoptotic signaling pathway are activated correspondingly. This review underlines the significance of the structural features of the polysaccharide, and thereafter indicates the modes of physiological function.