Natural and human-induced hydrological changes can influence organic carbon (OC) composition in fluvial systems, with biogeochemical consequences in both terrestrial and marine environments. Here, we use bulk and molecular carbon isotopes (13C and 14C) to examine spatiotemporal variations in particulate OC (POC) composition and age from two locations along the course of the Yellow River during 2015 and 2016. Dual carbon isotopes enable deconvolution of modern, pre-aged (millennial age) soil, and fossil inputs, revealing heterogeneous OC sources at both sites. Pre-aged OC predominated at the upstream site (Huayuankou) throughout the study period, mostly reflecting the upper riverine OC. Strong downstream (Kenli) intra-annual variations in modern and pre-aged OC were caused by increased contributions from modern aquatic OC production under the drier and less turbid conditions during this El Niño year. The month of July, which included the human-induced water and sediment regulation (WSR) event at Kenli, accounted for 82% of annual POC flux, with lower modern OC contribution compared with periods of natural seasonal variability. Both natural and human-induced hydrological events clearly exert strong influence on both fluxes and composition of Yellow River POC which, in turn, affect the balance between OC remineralization and burial for this major fluvial system.