With the urgent need for antiviral agents, antiviral materials with high biocompatibility and antiviral effects have attracted a lot of attention. In this study, gallic acid, a natural polyphenolic compound, was transformed into biocompatible graphene quantum dots (GAGQDs) which exhibit enhanced antiviral activity against pseudorabies virus (PRV). The as-prepared GAGQDs inhibit PRV proliferation with a 104-fold reduction in viral titers. Investigation of the antiviral mechanism revealed that GAGQDs inhibit the adsorption, invasion and replication of PRV infection. Treatment with GAGQDs regulates the expression levels of interferon-related antiviral proteins, including mitochondrial antiviral-signaling protein (MAVS), signal transducer and activator of transcription 1 (STAT1) and 2',5'-oligoadenylate synthetase 1 (OAS1), suggesting that GAGQDs can stimulate innate antiviral immune responses, resulting in enhanced antiviral effects. More importantly, GAGQD treatments alleviate clinical symptoms and reduce mortality in PRV-infected mice. Our results reveal the enhanced therapeutic effects of GAGQDs against PRV infection in vitro and in vivo, suggesting the potential of GAGQDs as a promising novel antiviral agent.