N-ethylmaleimide-sensitive factor attachment proteins (NAP: NAPA and NAPB) play a role in Soluble N-ethylmaleimide-sensitive accessory protein receptor (SNARE) complex dissociation and recycling, associated with neuronal regulation and brain development and various severe early-onset epilepsies. Here, we report two patients from a Chinese family presenting with unexplained early-onset epileptic encephalopathies (EOEE) syndrome characterized by multifocal seizures, profound intellectual disability and global developmental delay. We identified the homozygous c.433-1G>A variant of the NAPB as the causative by trio-based exome sequencing. The novel splicing mutation in NAPB was third variant reported associated with EOEE syndrome. Our results gave further hints on the associations of variants in NAPB with EOEE and indicated that for patients with unexplained EOEE, the NAPB gene should be included into the data analysis from whole exome sequencing, which contributes to uncover more patients affected and rich the phenotypic spectrum.