The multitude of structural and physicochemical properties of toxicants pose a major challenge for the concurrent analysis of many different chemical classes. For applicability to future, high-throughput exposome-scale studies, the sample extraction procedure must be comprehensive, yet still rapid. To deplete matrix components from 200 µL urine and serum, a one-phase extraction followed by protein precipitation was used. Conversely, a two-phase extraction was utilized for breast milk. Due to the diverse chemical properties of the extracted compounds, the chromatographic column had to retain molecules with a broad range of polarities, and also effectively separate several isomers with identical mass to charge (m/z) ratios. In addition to implementing a state-of-the-art UHPLC system, careful optimization of the chromatographic conditions including gradient, eluents, and the addition of NH4F as an organic modifier was essential. To maximize sensitivity, the ion optic parameters of the triple quadrupole mass spectrometer were individually optimized for each compound. For details refer to the Online Methods section.