Introduction: Population growth and increasing water demand have exacerbated water resource scarcity. Treated wastewater (TWW) is a valuable alternative resource, but its complex composition, including micropollutants, raises concerns about reuse risks. Reactive barriers (RB) made with natural materials offer a solution to remove TWW contaminants. Evaluating RB process effectiveness and comprehending contaminant fate remain challenging. Recent advances in High-Resolution Mass Spectrometry (HRMS) and non-target analysis (NTA) provide an opportunity to identify contaminants and evaluate RB processes. The study aims to develop an NTA approach to assess RB effectiveness and understand the NTA contribution for broadening perspectives on the impact of reactive barrier treatment. Methods: In a serie of batch experiments, TWW was mixed with a RB under two conditions: TWW in contact with RB (TWW+RB) and TWW alone (TWW). After 1 h of contact, samples were extracted and analyzed using HPLC-HRMS. NTA workflow and statistical comparison between the two sample conditions were applied for data treatment. Additionally, a quantitative method was applied to 28 pharmaceutical residues. Sorption (%) was calculated by comparing concentrations in TWW+RB and TWW experiments. NTA’s applicability for sorption estimation was investigated using intensities from differential analysis. Results: Differential analysis shows significant differences in compounds that decrease or increase after contact with BR. A prioritization workflow identified and annotated 115 compounds. Natural products predominantly increased after RB contact, while compounds exhibited a significant 75% decrease in peak area are mainly pharmaceuticals. Notably, 61% of these molecules were sorbed at more than 50%. The sorption of emerging contaminants (ECs) estimated by NTA correlated satisfactorily with sorption quantified through target analysis, although caution regarding the matrix effect is necessary. Discussion: NTA provides a comprehensive view of RB treatment performance, giving information on the compounds released by the barrier and its sorption capacity. NTA emphasizes the benefits of RB treatment while highlighting the need to study compounds reloaded by RB. It is a powerful tool for understanding the advantages of nature-based reactive barrier treatment in improving the quality of TWW.