Carbon-supported Pt–Ru alloys with a Pt/Ru ratio of 1:1 were prepared by NaBH4 reduction at room temperature. X-ray diffraction (XRD) measurements indicate that the as-prepared Pt–Ru nanoparticles had a face-centered cubic (fcc) structure. X-ray photoelectron spectroscopy (XPS) analyses demonstrate that alloying with Ru can decrease the 4f electron density of Pt, which results in a positive binding energy shift of 0.2 eV for the Pt 4f peaks. The catalytic properties of the synthesized Pt–Ru alloy catalysts were compared with those of commercial Pt/C catalysts by linear sweep voltammetry (LSV). The results show that the mass activity of the oxygen reduction reaction (ORR) is enhanced by 2.3 times as much mass activity of Pt relative to the commercial Pt/C catalyst. Single-chambered microbial fuel cell tests also confirm that the Pt–Ru alloys as cathode catalysts have better performance than that of commercial Pt/C catalysts.