Well-defined porous PtAu alloyed nanoflowers (PtAu ANFs) were fabricated by a one-pot surfactant-free successive co-reduction approach. Methotrexate, an antimetabolite and antifolate drug, was employed here as the shape-regulator and stabilizer for the first time, while no any seed, polymer, or surfactant was involved during the synthetic process. The as-prepared architectures were characterized by transmission electron microscopy, energy-dispersive X-ray spectroscopy, X-ray photoelectron spectroscopy, and X-ray diffraction in details. The as-synthesized hybrid nanocrystals exhibited improved catalytic performances for ethanol oxidation and oxygen reduction in contrast with PtAu-10 NCs, PtAu-50 NPs, and commercial Pt black catalysts, demonstrating their promising applications in fuel cells. Copyright (c) 2015, Hydrogen Energy Publications, LLC. Published by Elsevier Ltd. All rights reserved.