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Abstract Plant protection products (PPP) contain active substance(s) and co-formulants. As complex mixtures, PPPs are susceptible to toxicokinetic or toxicodynamic interactions, enhancing toxicity. Considering these interactions is crucial to avoid underestimating risk. In vitro testing identifies toxicokinetic interactions among PPP components and establishes toxicological threshold values for these products. In vitro-in vivo extrapolation (IVIVE) using physiologically based kinetic (PBK) modeling helps derive organism-based threshold values for PPPs. Currently, PBK modeling is primarily applied to single substances. We present a proof-of-concept for generating data for whole-mixture PBK modeling for PPP operator risk assessment. A PPP containing difenoconazole and mandipropamid was selected based on our previous study, which identified their metabolic interactions. Since PPP operators are predominantly dermally exposed, traditional rapid equilibrium dialysis (RED) for determining fraction unbound and pooled human liver microsomes (HLM) for hepatic clearance are insufficient. We integrated RED and HLM with EpiDerm Full Thickness 400 (EFT-400), a skin-penetration model. EFT-400 was spiked with concentrated and dilutions of formulated product and pure difenoconazole for 24 h before transferring receptor contents to RED or HLM. Difenoconazole concentration was measured using LC-MS/MS, and fraction unbound and hepatic clearance were determined. Hepatic clearance of difenoconazole decreased significantly in concentrated formulated product and dilutions compared to pure difenoconazole, indicating CYP enzyme inhibition by mandipropamid. Difenoconazole fraction unbound remained unchanged in concentrated formulated product but increased with product dilution. This strategy provides a framework for PBK modeling of mixtures, acknowledging potential combination effects and toxicokinetic interactions within complex mixtures with clear risk drivers such as PPPs.