Summary Catalyst support alters the electronic structure of metal catalysts, resulting in an enhanced activity. In this study, nitrogen‐doped defective graphene nanosheet‐supported Pt nanoparticles (2.8 nm) (Pt/N‐dGN) were prepared and successfully used as electrocatalysts in the sequential and transient catalysis of the neutral d‐glucose oxidation reaction (GOR). The Pt nanoparticles were determined to be single crystals enclosed with the (110) plane. The spherical‐aberration‐corrected field transmission electron microscopy image revealed a Pt/N‐dGN bearing with point vacancies, 5‐8‐5, 5‐9, and 5‐7‐7‐5 defects. N‐dGN was prepared by fast exfoliation and sequential nitrogen doping with urea using a sonoelectrochemical and hydrothermal method. The Pt/N‐dGN with a Pt binding energy of 71.4 eV showed significant mass activity (7.83 × 1019 molecules g−1Pt) toward the sequential catalysis of neutral GOR. The electron transfer number was 2.71 and approximately two‐electron transfer mechanisms occurred. The repeated currents of GORs were observed in the catalysis from the 2nd cycle to the 25th cycle.