Composite material based on room temperature ionic liquid (RTIL) N-butylpyridinium hexafluorophosphate (BPPF6), sodium alginate (SA), and graphite was used to construct a novel horseradish peroxidase (HRP) biosensor for the determination of H2O2. The morphology of the as-prepared biosensor was characterized by scanning electron microscopy (SEM). Cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) were used to characterize the process of the performance of the biosensor.. Parameters affecting the performance of the biosensor, including the concentrations of o-aminophenol (OAP) and HRP and the pH value of substrate solution, were optimized. Under the optimized experimental conditions, H2O2 could be detected in a linear calibration range of 1.0 to 6.0 mu M with a correlation coefficient of 0.9847 (n = 7) and a detection limit of 0.5 mu M at a signal/noise ratio of 3. The prepared biosensor not only had economic and disposable property but also showed good detection precision, bioactivity, storage stability, and reproducibility. (c) 2008 Elsevier Inc. All rights reserved.