The conducted laboratory tests allowed determination of the efficiency of removing ammonium nitrogen, iron, and manganese in the biofiltration process on chalcedonite beds. The process of water purification was carried out by a single- and two-stage biofiltration method with gravitational and anti-gravitational flow. The study examined the extent to which chemical activation of the bed with potassium manganese (VII) affects the course of the nitrification process and the rate of biofilm formation. The obtained test results indicate that two-stage biofiltration, with initial chemical activation at the first stage of biofiltration, is an effective method for purifying waters with an abnormal content of ammonium nitrogen with simultaneous removal of iron and manganese. Activation of the bed had an effect on, among other things: biofilm formation time, efficiency of removing manganese (II) ions, and oxygen consumption in the biofiltration process. Due to the longer maturation time of the activated bed, the normative value of ammonium nitrogen (< 0.39 N-NH4+) was obtained on the 23rd day of the operation of the filters, and in the non-activated bed on the 14th day. The method of bed preparation did not affect the efficiency of removal of iron compounds.