Objective To investigate the effects of calcium chloride supplementation in drinking water on fat deposition and intestinal flora in mice fed high-fat diet, and provide a theoretical basis for improving carcass characteristics of animals and human health.Method Twenty-seven 4-week-old C57BL male mice were divided into two groups including a high-fat diet (HFD) group and a group with HFD+calcium chloride supplementation in drinking water. The experiment was conducted for 13 weeks. The body weight of mouse was measured weekly. The body fat content of mouse was determined at the end of the experiment. After mice were sacrificed, the subcutaneous fat and epididymal fat of mice were collected and weighed. Mouse feces were collected during 10-12 weeks for high-throughput sequencing of 16S rRNA and the effects on intestinal flora were analyzed.Result Calcium chloride supplementation in drinking water significantly reduced the body weight, body fat content, subcutaneous fat index and epididymal fat index of mice, and compared with HFD group these indexes were reduced by 12.85%, 32.69%, 26.65% and 18.60%, respectively. Meanwhile, calcium chloride supplementation in drinking water increased the community diversity and bacterial abundance in mouse fecal samples. At the phylum level, calcium chloride supplementation had no effect on the abundance of Bacteroidetes, Firmicutes, Proteobacteria, Deferribacteres, Actinobacteria and Tenericumes. At the class level, compared with the HFD group, calcium chloride supplementation in drinking water significantly reduced the relative abundance of Erysipelotrichia and Actinobacteria, whereas significantly increased the relative abundance of Clostridia.Conclusion Calcium chloride supplementation in drinking water reduces the body weight and body fat content of mouse fed HFD, which might be associated with the increase in bacterial diversity and abundance of some specific bacteria in mouse feces.