One lab-scale biofilter coupling nitrification and anaerobic ammonium oxidation (ANAMMOX) that simultaneously removes Fe, Mn and ammonia from simulated groundwater was adopted to investigate the influence of Mn and ammonia on nitrogen conversion and Mn removal kinetics in this study. The results showed that autotrophic nitrogen removal proportion (ANRP) rose slightly with the raise of Mn concentration and declined along with the raise of ammonia; the average ratios were 49.6%, 51.5%, 51.8%, 52.3%, 52.6%, 48.9%, 47.4% and 38.8%, respectively. Relative constant or slight down trend of accumulated ANRP was detected in filter bed which indicated the superiority of nitrification in relevant areas. After reaching a certain value, Mn could promote ANAMMOX in the upper part of the filter bed and shorten the main ammonia conversion area. As ammonia content rising, the maximum accumulated ANRP reduced and the maximum value acquired height went up. Moreover, the ammonia inhibition threshold for ANAMMOX in the biofilter might be different from waste water treatment. Mn removal could be assessed by first order reaction in all the eight periods and the k values were more comparable than those in abiotic Mn oxidation.