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Laser Ablation as a Versatile Tool To Mimic Polyethylene Terephthalate Nanoplastic Pollutants: Characterization and Toxicology Assessment Davide Magrì,†,‡,¶ Paola Sánchez-Moreno,§,¶ Gianvito Caputo,† Francesca Gatto,§,∥ Marina Veronesi,⊥ Giuseppe Bardi,§ Tiziano Catelani,# Daniela Guarnieri,§ Athanassia Athanassiou,† Pier Paolo Pompa,*,§ and Despina Fragouli*,† †
Smart Materials, Istituto Italiano di Tecnologia, Via Morego, 30, 16163 Genova, Italy Department of Informatics, Bioengineering, Robotics and Systems Engineering, University of Genova, Via All’Opera Pia, 13, 16145 Genova, Italy § Nanobiointeractions & Nanodiagnostics, Istituto Italiano di Tecnologia, Via Morego, 30, 16163 Genova, Italy ∥ Department of Engineering for Innovation, University of Salento, Via per Monteroni, 73100 Lecce, Italy ⊥ D3-PharmaChemistry, Istituto Italiano di Tecnologia, Via Morego 30, 16163 Genova, Italy # Electron Microscopy Facility, Istituto Italiano di Tecnologia, Via Morego 30, 16163 Genova, Italy ‡
S Supporting Information *
ABSTRACT: The presence of micro- and nanoplastics in the marine environment is raising strong concerns since they can possibly have a negative impact on human health. In particular, the lack of appropriate methodologies to collect the nanoplastics from water systems imposes the use of engineered model nanoparticles to explore their interactions with biological systems, with results not easily correlated with the real case conditions. In this work, we propose a reliable topdown approach based on laser ablation of polymers to form polyethylene terephthalate (PET) nanoplastics, which mimic real environmental nanopollutants, unlike synthetic samples obtained by colloidal chemistry. PET nanoparticles were carefully characterized in terms of chemical/physical properties and stability in different media. The nanoplastics have a ca. 100 nm average dimension, with significant size and shape heterogeneity, and they present weak acid groups on their surface, similarly to photodegraded PET plastics. Despite no toxic effects emerging by in vitro studies on human Caco-2 intestinal epithelial cells, the formed nanoplastics were largely internalized in endolysosomes, showing intracellular biopersistence and long-term stability in a simulated lysosomal environment. Interestingly, when tested on a model of intestinal epithelium, nano-PET showed high propensity to cross the gut barrier, with unpredictable long-term effects on health and potential transport of dispersed chemicals mediated by the nanopollutants. KEYWORDS: nanoplastics, polyethylene terephthalate, laser ablation, Caco-2, intestinal epithelium, nanotoxicology
T
he huge plastic islands floating in the oceans, known as the five gyres, are the symbol of human impact on the planet and the emblem of the “Anthropocene”.1,2 © XXXX American Chemical Society
Received: February 19, 2018 Accepted: June 19, 2018
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DOI: 10.1021/acsnano.8b01331 ACS Nano XXXX, XXX, XXX−XXX
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Cite This: ACS Nano XXXX, XXX, XXX−XXX
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ACS Nano
Figure 1. PNP synthesis. (a) Schematic representation of the laser ablation procedure applied to produce PET PNPs. (b) Representative TEM micrograph and size distribution of PET PNPs.
impact.23−25 Several toxic effects of PNPs are described in the literature; nevertheless uptake and clearance seem to be the factors mainly influenced by the nanometric dimension. Interacting with lipids, PNPs could perturb cell membranes, penetrate by different endocytosis pathways, and remain in the organism longer than microsized plastic particles.26−28 As already observed for diverse types of organic and inorganic particles, the smaller size favors their passage through the intestinal barrier.29,30 Furthermore, the high surface/volume ratio of the smallest plastic debris make them ideal candidates to interact and transfer many hazardous contaminants (polychlorinated biphenyls, organochlorine pesticides, polyaromatic hydrocarbons, etc.) to living organisms, increasing their impact and toxicity.31−33 However, due to the difficulties in performing a correct sampling, separation, concentration, and identification of real PNP samples, the investigation of their interactions with biological systems and other compounds requires the use of model engineered systems.9 In particular, until now, the studies related to this class of pollutants have been mainly based on the use of monodispersed polystyrene nanospheres, synthesized by colloidal chemistry, following bottom-up approaches.29,34−38 Such protocols typically produce plastic nanostructures with few defects and homogeneous chemical composition, substantially different from the PNPs dispersed in the environment, which have irregular shapes and complex surface chemistry, due to the degradation experienced.9 Moreover, the PNPs synthesized by bottom-up approaches could contain solvent residuals, surfactants, and other substances used during their synthesis process, which may affect their behavior in aqueous media and their interactions with biological systems.39 Little is known about the effects PNPs on humans; nevertheless a handful of studies using polystyrene nanospheres have been carried out showing their potential to cause sublethal cellular injury to in vitro liver tissue and to disrupt iron transport using an in vitro model of intestinal epithelium after acute oral exposure.29,40 On the other hand, literature reports the lack of toxicity upon interaction with carboxylated polystyrene nanoparticles to various human cell types including HeLa epithelial cells, lung epithelial and astrocytoma cell lines, and an in vitro model of the blood−brain barrier.41,42 The strategy proposed in this study offers the possibility to fabricate diverse types of PNPs that can be used for the realistic investigation of their interactions with biological
Since the beginning of the twentieth century, the development of the polymer science and the launch of plastic production have led to one of the greatest revolutions for human society, although, at the same time, to plastics waste accumulation in the marine environment, which is estimated to reach 250 million tonnes in 2025.3,4 Although plastic is considered to be a long-lasting and stable material in the marine environment, weathering processes, such as biotic and abiotic degradation,5,6 induce the breakdown of plastic debris, resulting in the formation of fragments with dimensions ranging from a few centimeters to nanometers.3,5 Specifically, UV-light-induced oxidation is the most effective abiotic process for the degradation of floating plastics in open waters, while on beaches this phenomenon is enhanced by the contribution of temperature and mechanical abrasion.6−8 While macroscopic plastics can be isolated from a variety of matrices and environments (water, sediments, stomach contents, tissues), the sampling and quantification of the smallest fraction are very challenging, resulting in very heterogeneous data. Those data are mainly focused on plastic microparticles (PMPs), while there is poor knowledge about the fraction having a size of
