Realgar is a naturally occurring arsenic sulfide that is also called red arsenic (or Xionghuang, in Chinese) due to its deep red color. It contains over 90％ tetra-arsenic tetra-sulfide (AS4S4). However, disadvantages such as low solubility in water, poor gastrointestinal absorption, high toxicity, and poor bioavailability have seriously limited clinical applications of realgar. By taking advantage of an arsenic-resistant strain of indigenous Acidithiobacillus ferrooxidans (A. Ferrooxidans), we succeeded in the development of a novel method for realgar preparation by bioleaching processes. However, little is known about the bioleaching of realgar using A. Ferrooxidans under variousconditions due to lack of thorough studies. The parameter optimization of biological processes that is significant novelty and scientific impact for realgars clinical use. The fundamental knowledge derived from this study should provide new insights into and novel applications for preparation of realgar. We are interested to obtain optimal conditions in order to enhance realgar dissolution. These factors are worthy of study include temperature, Ph, bacteria population, concentration of ferrous iron, and pulp density. The primary aims of this paper were investigated to evaluate environmental parameters for bioleaching of arsenic from realgar using A. Ferrooxidans. The behavior of A. Ferrooxidans in response to different leaching conditions was investigated in a series of batch experiments. The bacterial leaching of realgar has been investigated using A. Ferrooxidans. The effects of a range of crucial operational parameters, including temperature, Ph values,bacteria population, ferrous iron concentration, and pulp density on the efficiency of the bioleaching process were investigated. Conversion algorithms were applied to test the experimental data from the bioleaching of realgar. The experimental data on realgar conversion were analyzed and simulated by mathematical models.