In this work, preparation and characterization of a novel nanocomposite containing bismuth oxide (Bi(2)O(3)) nanoparticles and multiwalled carbon nanotubes (MWCNTs) was presented. Powder X-ray diffraction (XRD) studies revealed that as-synthesized Bi(2)O(3) nanoparticles are crystalline and belong to α-phase with monoclinic symmetry. Field emission scanning electron microscopy (FESEM) study results showed that the size of Bi(2)O(3) nanoparticles is 50 nm. Energy-dispersive X-ray (EDX) spectra of as-prepared Bi(2)O(3)-MWCNT nanocomposite displayed characteristic Bi and C peaks which confirmed the incorporation of Bi(2)O(3) with MWCNT. The prepared Bi(2)O(3)-MWCNT was also characterized by scanning electron microscopy (SEM) and atomic force microscopy (AFM) studies. The direct electron transfer of horseradish peroxidase (HRP) has been revealed at Bi(2)O(3)-MWCNT modified glassy carbon electrode (GCE). In order to firmly anchor the HRP molecules onto Bi(2)O(3)-MWCNT matrix, a thin layer of 1% nafion (NF) solution was coated as a binder. The fabricated NF/HRP/Bi(2)O(3)-MWCNT/GCE exhibits well defined quasi-reversible redox peaks at a formal potential (E(0)') of -0.326V vs. Ag/AgCl reference electrode in 0.05M phosphate buffer solution (PBS), pH 7. NF/HRP/Bi(2)O(3)-MWCNT film remarkably lowers the over potential for H(2)O(2) reduction than MWCNT, Bi(2)O(3)-MWCNT and unmodified GCEs. The proposed composite film exhibits quick amperometric i-t response (5s) towards H(2)O(2) in the linear range of 8.34-28.88mM with a sensitivity of 26.54 μA μM(-1)cm(-2). The developed NF/HRP/Bi(2)O(3)-MWCNT biosensor has a good operational stability and high selectivity towards H(2)O(2).