The status and kinetics of monocyte activation during acute P. knowlesi infection was investigated by latex‐induced, luminol‐dependent chemiluminescence (CL) response. The contribution of various reactive oxygen species (ROS) to CL response was estimated before infection and at peak parasitaemia (day 7 post infection) by using scavengers of ROS (benzoate, catalase and superoxide dismutase). The chemiluminescence index (CLI) was not found to be significantly different from controls on day 2 postinfection, but was significantly higher on days 5 and 7 postinfection. Hydroxyl radical (OH·) production was considerably elevated, whereas superoxide anion(O2−·) and hydrogen peroxide (H2O2) production dropped following infection. These changes in generation of ROS are discussed in relation to the progression of parasitaemia to high levels, immunopathology and immunosuppression during acute P. knowlesi infection.