Nine cruises were conducted on a seasonal basis from 2013 to 2015 to investigate the spatial distribution and seasonal variability of δ13C and δ15N in particulate organic matter (POM), and its response to flooding in the Pearl River Estuary (PRE), south China. Our study reveals highly variable isotope ratios between seasons in this subtropical estuary, following seasonal climatic and hydrological cycles. Wet seasons had more isotopically depleted δ13C values, indicating the dominance of terrestrial and freshwater algae POM, whereas the contribution from marine phytoplankton (16%–59%) was higher during the dry seasons. In contrast, δ15N exhibited a sharp increase (up to 17.6‰) at low salinities (0–5) during high flow seasons. This was consistent with high NO3− concentrations, reflecting phytoplankton and bacteria assimilation of δ15N enriched‐ NO3− as well as notable isotope fractionation during microbial mineralization. There was little annual variability in δ13C over the 2 year period; however, particulate nitrogen (PN) exhibited lower concentrations but more enriched isotope values in 2015 than in 2014. This can be best explained by temperature‐modulated biological processing of particulate organic nitrogen, partially due to different biogeochemical responses during normal (2014) and strong El Niño (2015) years. After flooding in June 2015, terrestrial organic matter and freshwater phytoplankton were the major components of POM within the estuary and shelf areas, whereas marine phytoplankton was the dominant component in the adjacent coastal waters with mid‐salinities (10 < S < 20), as revealed by a phytoplankton bloom (>10 μg L−1) and δ13C‐enriched but δ15N‐depleted POM.

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