Detection of nitromethane (CH3NO2) has received scientists' attention recently. However, its detection with the direct electrochemical measurement method was reported rarely. In this work, we explore an electrochemical biosensor for CH3NO2 based on immobilization of four heme proteins, sodium carboxymethyl cellulose (CMC), room temperature ionic liquid-1-butyl-3-methylimidazolium tetrafluoroborate ([bmim][BF4]]) and multi-walled carbon nanotubes (MWNTs) biocomposite films. The surface morphologies of the representative heme proteins hybrid films were demonstrated by scanning electron microscopy (SEM). The different parameters, including working potential, pH of supporting electrolyte and the scan rate that governed the performance of the biosensor have been studied in detail and optimized. Electrocatalytic parameters for the determination of CH3NO2 with the four heme proteins film electrodes were evaluated by chronoamperometry. (C) 2012 Elsevier B.V. All rights reserved.