Abstract: To observe the bactericidal efficacy of Hengjingsuan mainly composed of citric acid, malic acid, sulfamic acid and formic acid and its impact on the intestinal microbiota in mice. Suspension quantitative bactericidal test was used to evaluated the bactericidal efficacy of Hengjingsuan on Escherichia coli 8099, Candida albicans ATCC 10231, Staphylococcus aureus CMCC（B） 26003, Pseudomonas aeruginosa ATCC 15442 and African Swine Fever Virus（ASFV）. The acute toxicity test of Hengjingsuan in mice were was carried out by 14 days maximum dose method and the safe concentration of Hengjingsuan was determined by observing the effect of disinfectant on cell morphology of HEK293T cells and calculating the cell activity. The effect on community structure diversity of intestinal bacteria was analyzed by using high-throughput sequencing technology.The results showed that Hengjingsuan solution at the concentration of 0.1 g·L^-1 could kill E. coli 8099, C. Albicans ATCC 10231, S. aureus CMCC（B） 26003, P. aeruginosa ATCC 15442 effectively within 2 minutes, and it could kill ASFV effectively at 1.56 g·L^-1 within 5 minutes. Hengjingsuan was an actual non-toxic substance, and its safe concentration for the cell was ≤ 1.56 g·L^-1. Compared with normal group, the abundance of bacteroidetes significantly increased in mice treated with Hengjingsuan, meanwhile the abundance ratio of bacteroidetes to Firmicutes also increased. Therefore, Hengjingsuan was a new type of disinfectant, which was safe, non-irritating and had great bactericidal efficacy on target pathogenic microorgan.