Autochthonous organic carbon (AOC) formed by biological carbon pump (BCP) in surface waters may serve as a significant carbon sink. The locations, magnitudes, variations and mechanisms responsible for the terrestrial missing carbon sink by BCP are uncertain, especially in large river systems. In this study, hydrochemical characteristics, carbon isotope compositions of dissolved inorganic carbon (DIC) and organic carbon (OC), n-alkane homologues and C/N ratios of organic matter along the Yangtze River and the Yellow River were investigated to constrain the OC source and the significance of BCP effect. It was found that (1) DIC concentrations in the Yellow River were much higher than those in the Yangtze River, which was controlled primarily by the temperature effect; (2) AOC in the both rivers was characterized by lower C/N ratios and 813CPOC values. Based on calculation of n-alkanes compounds, the AOC proportions ranged from 29 to 88% (49% on average, with a higher proportion (55%) in the rainy season than in the dry season (46%)) and 19-68% (41% on average; with a lower proportion in the rainy season (31%) than in the dry season (51%)) in the Yangtze River and the Yellow River, respectively, indicating intense aquatic production. Low dissolved CO2 concentration (6.17 mu mol/L on average) of the Yangtze River limited the aquatic production and decreased the BCP effect in the dry season, indicated by lower AOC proportion. However, the BCP effect increased in the Yellow River in the dry season mainly due to the increased light penetration; (3) even in high turbidity riverine systems such as the Yellow River, the aquatic photosynthetic uptake of DIC could produce considerable AOC. These findings clearly show the formation of AOC by BCP in both the clear and high turbidity riverine systems, suggesting a potential direction for finding the terrestrial missing carbon sink.