Unraveling Nanoplastics-Enzyme Interactions: Physicochemical, Structural, Functional, and Cell Biological Characterization of α-Amylase-Nanoplastics Complexes.

The topic of micro- and nanoplastics received significant attention in recent decades due to increasing environmental exposure, strong public perception, and emerging health concerns. While knowledge regarding detection and material characteristics has improved, the understanding of impact on cells remained unclear. As biological effects are initially caused by molecular interactions, consequently direct interactions with biomolecules, such as enzymes, are of particular relevance. In this occasion, effects may vary depending on the plastic type and particle properties. The specific aim of this study was to characterize the direct molecular interactions by means of selected model proteins and a variety of different nanoplastic particles. Therefore, the aim of the study was to exemplarily characterize α-amylase's (as a model enzyme) interactions with different nanoplastics and the resulting effects on enzyme structure and function, as well as cellular responses. The properties of the α-amylase-nanoplastic mixtures were analyzed using dynamic light scattering (DLS), Fourier-transform infrared spectroscopy (FTIR), fluorescence spectroscopy, and Phadebas amylase activity test. Additionally, Caco-2 cells were used as a model system for the human intestinal barrier and exposed to these complexes to evaluate cellular uptake through flow cytometry, microscopy, and viability testing. All applied nanoplastics interacted with α-amylase, forming complexes with adsorption affinities that depended on the particle type (PP ≫ PE > PET ≫ PLA). FTIR and fluorescence analyses showed particle-specific structural changes. Despite these differences in structural response, concentration-dependent enzyme inhibition was measurable, depending on the particle type. Uptake studies on Caco-2 cells indicated no internalization or cytotoxicity. These findings suggest that nanoplastics influence the enzyme structure and function based on their chemical properties, offering new insights into direct enzyme-nanoplastics interactions and their potential impacts on enzymes and cells.