Efficient extraction of small microplastic particles from rat feed and feces for quantification.

To date, microplastic is ubiquitously encountered in the environment. Studies analyzing microplastic in terrestrial ecosystems, including animal feces and feed, are few. Microplastic quantification method validation and harmonization are not yet far developed. For the analysis of small microplastic, approximately <0.5 mm, extraction from organic and inorganic materials is fundamental prior to quantitative and qualitative analysis. Method validation, including recovery studies, are necessary throughout the analytical chain. In this study, we developed an optimized, efficient protocol with a duration of 72 h for the digestion of laboratory rat feed and feces. A combination of a mild acidic (H2O2 15%; HNO3 5%) and an alkaline treatment (10% KOH) dissolving the previous filter, followed by enzymatic digestion (Viscozyme®L) proved to be efficient for the extraction and identification of spiked polyamide (15-20 µm) and polyethylene (40-48 µm) from feed and feces samples from rats, showing high recovery rates. Extracted rat feces samples from an in vivo study in which Wistar rats were fed with feed containing microplastic were analyzed with pyrolysis-gas chromatography-Orbitrap™ mass spectrometry, quantifying recovered microplastic in rat feces in environmentally relevant concentrations. The presented three-step protocol provides a suitable, time and cost-effective method to extract microplastic from rat feed and feces.

Intestinal permeability and gene expression after polyethylene and polyamide microplastic ingestion in Wistar rats.

Microplastic particles are ubiquitous in the environment. However, little is known about their uptake and effects in humans or mammalian model organisms. Here, we studied the effects of pristine polyamide (15-20 µm) and polyethylene (40-48 µm) particles after oral ingestion in rats. The animals received feed containing microplastic particles (0.1% polyamide or polyethylene, or a mixture of both polymers) or a control diet without microplastic particles, for 5 weeks. The permeability of the duodenum was investigated in an Ussing chamber, whereas gene expression and concentration of tight junction proteins were measured in gut tissue and plasma. Microplastic particles were quantified by pyrolysis-gas chromatography/mass spectrometry in rats' feces. Rats fed with microplastic particles had higher duodenal permeability. Expression of gene coding for the tight junction protein occludin (OCLN) was higher in PE treated animals compared to control or the PA group. No changes in the expression of the gene coding for zonula occludens protein 1 were detected. Occludin protein concentrations were below the limit of detection of the applied method in both gut and plasma. Zonula occludens protein 1 concentrations in the gut were significantly higher in groups exposed to PA and PE as compared to control, while zonula occludens protein 1 concentrations in plasma did not show significant changes. These results demonstrated that short-term exposure to a dose of 0.1% (w/w) microplastic particles in feed had limited effects on duodenal permeability, expression of pro-inflammatory protein genes and tight junction protein genes in the duodenum.

Micro- and nanoplastic toxicity on aquatic life: Determining factors.

Plastic pollution has become a major environmental concern due to its omnipresence and degradation to smaller particles. The potential toxicological effects of micro- and nanoplastic on biota have been investigated in a growing number of exposure studies. We have performed a comprehensive review of the main determining factors for plastic particle toxicity in the relevant exposure systems, from publications until including the year 2018. For a focused scope, effects of additives or other pollutants accumulated by the plastic particles are not included. In summary, current literature suggests that plastic particle toxicity depends on concentration, particle size, exposure time, particle condition, shape and polymer type. Furthermore, contaminant background, food availability, species, developmental stage and sex have major influence on the outcome of plastic particles exposures. Frequently reported effects were on body and population growth, energy metabolism, feeding, movement activity, physiological stress, oxidative stress, inflammation, the immune system, hormonal regulation, aberrant development, cell death, general toxicity and altered lipid metabolism. Several times reported were increased growth and food consumption, neuro-, liver- or kidney pathology and intestinal damage. Photosynthesis disruption was reported in studies investigating effects on phytoplankton. For the currently unquantified plastic particles below 10 µm, more toxic effects were reported in all aquatic life, as compared to plastic particles of larger size.