Display omitted •Co-vacancy rich Co3O4 is synthesized by using layered cobalt glycerate as a precursor.•Vco-Co3O4 exhibits outstanding catalytic activity for the hydrolysis of ammonia borane.•Vco-Co3O4 yields a high hydrogen production rate of 934 mL min−1 gCat-1.•Vco-Co3O4 shows a low activation energy of 32.65 kJ mol−1 for catalytic hydrolysis. Development of efficient and economical non-precious metal catalysts for dehydrogenation of ammonia borane is an important way to store and release hydrogen. Vacancy-engineered defective catalysts with modulated electronic structure allow ideal adsorption and dissociation of reactant molecules for the optimized catalytic activity. In this study, we report a facile synthesis of cobalt-defective Co3O4 (VCo-Co3O4) nanocatalyst by the calcination of layered cobalt glycerate as a key precursor. The stacked cobalt-oxygen sheets and interlayer bound glycerate anions in a layered structure are favorable for introducing cation vacancy during calcination process. The obtained Vco-Co3O4 exhibits outstanding catalytic activity for the hydrolysis of ammonia borane, along with a high hydrogen production rate of 934 mL min−1 gCat-1 and a low activation energy of 32.65 kJ mol−1. The catalytic performance of the VCo-Co3O4 catalyst can be well maintained for consecutive five cycles. The current study establishes a new example of cation vacancy strategy to facilitate the hydrolysis of ammonia borane for hydrogen generation.