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Water-soluble organic nitrogen (WSON) in marine atmospheric aerosols affect the water solubility, acidity, and light-absorbing properties of aerosol particles, which are important parameters in assessing both the climate impact and the biogeochemical cycling of bioelements. Size-segregated aerosol and surface seawater (SSW) samples were simultaneously collected over the subtropical North Pacific to investigate the origin of WSON in the marine atmosphere. The fine-mode WSON concentration (7.5 ± 6.6 ngN m−3) at 200–240∘ E along 23∘ N, defined as the eastern North Pacific (ENP), was significantly higher than that (2.4 ± 1.9 ngN m−3) at 135–200∘ E, defined as the western North Pacific (WNP). Analysis of the stable carbon isotope ratio of water-soluble organic carbon (WSOC; δ13CWSOC) together with backward trajectory indicated that most of the observed WSON in the fine particles in the ENP originated from the ocean surface. We found positive relations among nitrogen-fixation rate, dissolved organic nitrogen (DON) in SSW, and the WSON concentrations. The result suggests that reactive nitrogen (DON and ammonium), produced and exuded by nitrogen-fixing microorganisms in SSW, contributed to the formation of WSON aerosols. This study provides new insights into the role of ocean-derived reactive nitrogen aerosols associated with marine microbial activity.