Current deterministic water quality modelling methods are not sufficient to calculate the transport of estrogenic endocrine disrupting chemicals (eEDC) concentrations in a river. This is because eEDCs are present in very low concentrations and are very costly to analyse, its present and acts in a mixture of chemicals and little are understood regarding the interactions and transformation processes between eEDCs and its environment. During this study, eEDC activity and its transport along the Zenne river was investigated. The estrogenic equivalent concentrations (BEQ) were determined with Estrogen responsive elements - Chemical Activated Luciferase Gene Expression (ERE-CALUX) analysis. A water mass budget and BEQ load budget was calculated by using the sampling sites as flux boxes boundaries in the study area of the Zenne River. The BEQ measured along the Zenne River ranged from 0.7 - 16.5[ng E2-Eq/L]. The spatial BEQ along the Zenne shows a definite trend in the summer which can suggest a seasonal dependence in the summer (July and August). The application of the water budget to the Zenne confirmed that WWTP effluent is a major contributor to the flow of the Zenne. The WWTPs contributes between 42% -100% of the total flow of the Zenne River. The nearly closed BEQ load budgets [g E2-Eq/day] is a strong indication that BEQ may be conservative as it is transported along the Zenne River. The total contribution of the hospitals in Brussels, assuming that the effluent is treated at one of the WWTPs, is not so significant compared to the load contribution of the WWTP effluent. PCA was applied to establish whether the observations of in-situ water quality monitoring and BEQ relate to each other. It was noted that the PCA model with two components explained more than 70% of the observed variability in TEM, DO, CON, SPM and BEQ variables. PCA indicated that there is an upstream (Z3) – downstream (Z11) gradient for the TEMP, COND and BEQ variables. A more detailed analysis showed that these data also segregate along the PC2 axis following a seasonal pattern. This study clearly highlighted that urban wastewater is a major contributor to eEDC concentrations in the Zenne river. This in turn highlights the importance of wastewater treatment in reducing the BEQ loadings in water resources. This study also highlighted possible correlations with easy measurable water quality parameters namely: TEMP, COND and DO.