The toxicity of ionic liquids (ILs) on soil organisms has aroused wide attention due to their high-solubility. The present investigation focused on the toxicity of 1-octyl-3-methylimidazolium nitrate ([C(8)mim]NO3) on the microbial populations (bacteria, fungi, and actinomycetes), soil enzyme (urease, dehydrogenase, acid phosphatase, and beta-glucosidase) activities, microbial community diversity using terminal restriction fragment length polymorphism (T-RFLP), and abundance of the ammonia monooxygenase (amoA) genes of ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) using quantitative real-time polymerase chain reaction (q-PCR) in brown soil at each trial with doses of 0, 1.0, 5.0, and 10.0 mg/kg on days 10, 20, 30, and 40. The contents of [C(8)mim] NO3 in soil were measured using high performance liquid chromatography with recoveries of 84.3% to 85.2%, and changed less than 10% during the experimental period. A significant decrease was observed from the bacteria, fungi and actinomycetes populations at 10.0 mg/kg, at which the urease activity was inhibited and the beta-glucosidase activity was stimulated on days 20, 30, and 40. In addition, [C(8)mim] NO3 inhibited the dehydrogenase activity at 10 mg/kg on days 30 and 40 and the acid phosphatase activity on day 20. The diversity of the soilmicrobial community and the gene abundance of AOA- and AOB- amoA were also inhibited. Furthermore, the present investigation provided more scientific information for the toxicity evaluation of ILs in soil. (C) 2017 The Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences. Published by Elsevier B.V.