Zoutendijk Laura M, Matla Zenzi, Dusza Hanna M, Scholz-Böttcher Barbara M, Weckhuysen Bert M, Mandemaker Laurens D B, Meirer Florian
Inorganic Chemistry and Catalysis, Institute for Sustainable and Circular Chemistry, Utrecht University, Utrecht, 3584 CG The Netherlands.
Division of Toxicology, Institute for Risk Assessment Sciences, Department of Population Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, 3584 CM The Netherlands.
Environ Toxicol Chem. 2025 Jul 11. doi: 10.1093/etojnl/vgaf177.
Concerns are arising about potential health risks of micro- and nanoplastics (MNPs) to human life, as their presence has been reported in human blood, stool, liver, lung tissue, breast milk, and placenta. However, little is known about particle numbers and morphology, which is essential information for developing reliable risk assessment. Therefore, such characterization of MNPs in human tissue is an important yet difficult task, which involves sample digestion as an essential step in the pretreatment of organic matrices. Successful digestion enables accurate characterization of MNPs using micro-spectroscopy. In this study, eight different enzymes or enzyme mixtures commonly used in digestion protocols were tested in four different buffers, to select the best combinations of enzymes and buffers for the preprocessing of human placental tissue for MNP (spectro-)microscopic analysis. Placenta tissue was spiked with fluorescent 200 nm, 500 nm, 1 μm, and 10 μm polystyrene MNPs to analyze morphological stability throughout the digestion and to determine recovery rates (RRs). For the optimal protocol, RRs of 98% ±6 (200 nm), 148% ±8 (500 nm), 147% ±8 (1 μm) and 81% ±8 (10 μm) were determined using confocal fluorescence microscopy (CFM). We explain values >100% by dye leaching and hypothesize that the leached dye can bind to organic residue from tissue with a similar size as the FPS particles, causing false positives when counting MNPs. Morphological changes were not observed for the final digestion protocol both with CFM and scanning electron microscopy. Hence, we demonstrate an optimized enzymatic digestion protocol to digest (placental) tissue and report on the accuracy of the characterization of model MNPs using micro-spectroscopy, which will enable further research with an emphasis on sub-micron (<1µm) sized plastic particles.
微塑料和纳米塑料（MNPs）对人类健康的潜在风险引发了人们的担忧，因为已有报道称它们存在于人体血液、粪便、肝脏、肺组织、母乳和胎盘中。然而，关于颗粒数量和形态的了解却很少，而这些信息对于进行可靠的风险评估至关重要。因此，对人体组织中的MNPs进行此类表征是一项重要但困难的任务，其中涉及样品消化，这是有机基质预处理中的关键步骤。成功的消化能够使用显微光谱法对MNPs进行准确表征。在本研究中，在四种不同的缓冲液中测试了消化方案中常用的八种不同酶或酶混合物，以选择用于人胎盘组织预处理以进行MNP（光谱）显微镜分析的最佳酶和缓冲液组合。向胎盘组织中加入荧光200 nm、500 nm、1 µm和10 µm的聚苯乙烯MNPs，以分析整个消化过程中的形态稳定性并确定回收率（RRs）。对于最佳方案，使用共聚焦荧光显微镜（CFM）测定的回收率分别为98%±6（200 nm）、148%±8（500 nm）、147%±8（1 µm）和81%±8（10 µm）。我们通过染料浸出来解释大于100%的值，并假设浸出的染料可以与组织中与FPS颗粒大小相似的有机残留物结合，在计数MNPs时导致假阳性。使用CFM和扫描电子显微镜均未观察到最终消化方案的形态变化。因此，我们展示了一种优化的酶消化方案来消化（胎盘）组织，并报告了使用显微光谱法对模型MNPs进行表征的准确性，这将有助于进一步研究，重点是亚微米（<1µm）尺寸的塑料颗粒。
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