Quantifying micro- and nanoplastics in blood, urine, and soft tissues using optimized cascaded microfiltration and pyrolysis-GC/MS for exposomic investigations.

Microplastics (1 μm-5 mm) and nanoplastics (<1 μm) arise from the breakdown of larger plastic materials through thermal, photochemical, and mechanical degradation. Due to the wide use and haphazard disposal of plastic wastes, micro- and nano-plastics (MNPs) are ubiquitously detected in air, water, soil, and food, which leads to growing concern about chronic exposure and potential health impacts. MNPs have been detected across trophic levels and in more than 1000 species, indicating their extensive bioaccumulation. Recent studies have also detected MNPs in multiple human tissues and bodily fluids, suggesting that MNPs can be absorbed, circulated, and excreted. While various spectroscopic and microscopic techniques such as Raman, μ-FTIR, and LD-IR have advanced MNPs characterization, their applicability in environmental exposomic studies is limited by polymer-specific constraints, false-positive risks, and reduced sensitivity in complex biological matrices, particularly for submicron particles. In this study, we validated and applied cascaded microfiltration (1.2 μm then 0.7 μm pore sizes) followed by pyrolysis-gas chromatography mass spectrometry (Py-GC/MS) method enable concurrent quantification of MNPs (PS, PC, PMMA, PET, PVC, PP) in various biospecimen, facilitating exposure assessment. The method validation is focused on reducing background contamination and false-positive signals via matrix-matched calibration and rigorous quality control (MQLs 0.04-1.42 μg/mL). Applied to real samples (n = 20 each human/shark blood; dolphin fluids/tissues), we quantified total MNPs at 0.80 μg/mL median in human blood (PET 100% detection, PS/PVC dominant) and at 2.74 μg/mL in shark blood (PVC predominant, >3x humans). From size-range comparisons, we observed MNPs (transitional size range in 0.7-1.2 μm) prevail in all matrices showing 74-99% in different types of bodily fluids, suggesting possible partitioning of smaller MNPs into cellular fractions in blood. This work establishes a validated, and cross-species applicable method for sensitive and contamination-controlled quantification of MNPs in both fluid and solid biological matrices.