In this study, a new simulation-optimization approach is proposed to solve the multi-pollutant waste load allocation (WLA) problems in surface water systems. The proposed approach simulates the fate and transport of multiple pollutants utilizing the AQUATOX model. Since AQUATOX is an independent model and its integration into the optimization process is computationally not efficient, a multi-pollutant concentration-response matrix (mpCRM) is developed by using the results of the AQUATOX model. This mpCRM is then integrated into an optimization model where a nonlinear generalized reduced gradient (GRG) optimization method is used. Unlike in previously conducted studies, a new optimization formulation is proposed wherein the multiple pollutant loads are allocated among the source locations depending on their pre-assigned load allocation weights. This formulation allows for an equal or variable pollutant load allocation plan among source locations depending on the water management strategy for the watershed. The applicability of the proposed approach is evaluated on a sub-watershed of the Kucuk Menderes River Basin (KMRB) in Turkey by considering different load allocation scenarios. Furthermore, a detailed sensitivity analysis is conducted to evaluate the model results for different problem parameters. Identified results suggest that the proposed simulation-optimization approach is an effective way to solve the multi-pollutant WLA problem.