Pollution of urbanized rivers with excess nutrients due to groundwater discharge is an increasing environmental concern worldwide. Dan'ao river, a typical urbanized river in the Guangdong-Hong Kong-Macao Greater Bay Area, is experiencing heavy water pollution. However, the groundwater-derived nutrient loads had not yet been thoroughly quantified. In order to quantify the contribution of groundwater-derived nutrient inputs, we combined the methods of hydrological investigations and radium isotopes. Groundwater and river water samples were collected from the river upstream to the estuary for the analyses of radium quartets and nutrients including DIN, DIP and DSi. The results showed that the radium activities in both surface water and groundwater decreased from the estuary to the upstream. The groundwater discharge rate was estimated by the radium mass balance model using short-lived radium isotopes (223Ra and 224Ra). The estimated groundwater discharge rate ranged from 1.99 × 105 to 6.67 × 105 m3 d−1, comparable to the upstream river discharge rate of 4.23 × 105 m3 d−1. The groundwater-derived nutrient fluxes were 165.66–554.98 mmol m−2 d−1 for DIN, 2.47–8.26 mmol m−2 d−1 for DIP and 63.73–213.49 mmol m−2 d−1 for DSi, respectively. They contributed 19%~44% DIN, 16%~39% DIP, and 31%~60% DSi of all the nutrient inputs into the Dan'ao River, respectively. In addition, the nutrient inputs by groundwater discharge has an average DIN:DIP ratio of as high as 190, which is able to potentially affect the riverine and marine nutrient structures. These findings may provide useful information for designing control strategies for reducing massive nutrient inputs to Dan'ao River in the future.