Acid mine drainage (AMD) generated from mining activities has caused serious environmental concerns such as soil and water contamination and ecosystem destruction in the center karst region of Southwest China. The environmental pollution problems especially for soil ecosystem destruction caused by AMD have been extensively studies in recent years. The previous studies in this field have concentrated on the red soil system, however, have failed to consider the karst soil system. Especially for the study about the AMD contamination of paddy soil system in the vulnerable karst area has not yet been reported. Thus, the objective of this study was to evaluate environmental impacts of AMD irrigation on karst paddy soil. The findings in this study should have certain theoretical and practical significance in preventing the environmental pollution from mining activities. In this study, we chose the karst regions in Southwest China as the study area, where paddy ecosystem and AMD from coal gangue dumps were selected to be the research object. Through the addition of different levels of AMD to simulate continuous sewage irrigation. The physicochemical and biochemical parameters such as pH, EC, Eh, Fe, Mn, Cu, Zn, SO₄^2-, urease, phosphatase, and catalase activity were investigated to assess the effects of long-term irrigation with AMD on paddy soil. The results showed that the paddy soil have a good buffer performance for acidity under the lower pollution level of AMD. However, long-term high-intensity continuous pollution of AMD could lead to significant acidification of overlying water and paddy soil, and significantly increase the salinity of overlying water and paddy soil, and significantly change the redox environment conditions of the original paddy soil system. The continues pollution of AMD could significantly increase the concentration of contaminant such as iron, manganese, sulfate, and heavy metals in the overlying water. These contaminants in the overlying water could be reduced by the adsorption effect and buffer system of paddy soils under lower level of AMD, however, the adsorbed contaminants (Mn, Cu, and Zn) by paddy soil could be released into overlying water under higher level of AMD. The microbial biomass, urease, and phosphatase activity could also be significantly reduced under higher level of AMD. A large amount of H⁺ and rich iron ions from AMD addition could significantly promote the decomposition of H₂O₂ causing false positive trend of hydrogen peroxide enzyme activity continue to increase. In conclusion, the physicochemical and microbial characteristics of paddy soil in the karst region will be significantly changed under the long-term irrigation and pollution of AMD and ultimately severely threaten to the environmental quality of paddy soil ecosystem.