As a critical strategic hub for opening-up between China and ASEAN countries, the Beibu Gulf is a typical semi-enclosed bay where plastic pollution has drawn considerable concern. However, existing studies have mostly focused on microplastic (MP) pollution, while the vertical distribution and transport processes of nanoplastics (NPs) remain poorly understood. In this study, microplastics and nanoplastics were isolated from water samples collected in the Beibu Gulf region. The mass concentrations of four polymer types—polyethylene terephthalate (PET), polyvinyl chloride (PVC), polystyrene (PS), and polymethyl methacrylate (PMMA)—were quantitatively analyzed using pyrolysis–gas chromatography–mass spectrometry (Py-GC-MS), and their sources were apportioned by the positive matrix factorization (PMF) model. The results showed spatial heterogeneity in micro- and nanoplastic concentrations between nearshore surface waters and offshore as well as bottom waters. Source apportionment indicated that microplastic sources included fishing activities, land-based inputs of plastic waste, and ship emissions. This study presents the first systematic report on the mass concentrations, spatial distribution characteristics, and source apportionment of micro- and nanoplastics in the Beibu Gulf, providing essential baseline data for source-oriented control of plastic pollution and ecological conservation in this region.