Bio accessibility of tire-associated organic chemicals in fish gut (Oncorhynchus mykiss): insights from an in vitro digestion model

Tire and Road Wear Particles (TRWP) account for an important part of the anthropogenic particles released into the environment. There are scientific knowledge gaps as to the potential bio accessibility of chemicals associated with TRWP to aquatic organisms. This study aimed to investigate the solubilization of five tire-associated chemicals into fish gut using an in vitro digestion model (Oncorhynchus mykiss). Our results show that the targeted compounds were partly and rapidly solubilized into simulated fluids (SF) present in the gastrointestinal tract within a typical gut transit time for fish (3h in SFGASTRIC and 24h in SFINTESTINAL). The effects of food co-ingestion on the solubilization of tire-associated chemicals was compound-specific and either lowered or stimulated their solubilization into the gut fluids. Therefore, the uptake of the tire associated chemicals by the epithelial cells and related toxicity to fish need to be investigated.

Bioaccessibility of organic compounds associated with tire particles using a fish in vitro digestive model: solubilization kinetics and effects of food co-ingestion

Florian Frédéric Vincent Breider, Benoît Jean Dominique Ferrari, Dominique Grandjean, Luke Christopher Harris, Thibault Béranger Masset, Kristin Schirmer

Tire and road wear particles (TRWP) account for an important part of the polymer particles released into the environment. There are scientific knowledge gaps as to the potential bioaccessibility of chemicals associated with TRWP to aquatic organisms. This study investigated the solubilization and bioaccessibility of seven of the most widely used tire-associated organic chemicals and four of their degradation products from cryogenically milled tire tread (CMTT) into fish digestive fluids using an in vitro digestion model based on Oncorhynchus mykiss. Our results showed that 0.06% to 44.1% of the selected compounds were rapidly solubilized into simulated gastric and intestinal fluids within a typical gut transit time for fish (3 h in gastric and 24 h in intestinal fluids). The environmentally realistic scenario of coingestion of CMTT and fish prey was explored using ground Gammarus pulex. Coingestion caused compound-specific changes in solubilization, either increasing or decreasing the compounds’ bioaccessibility in simulated gut fluids compared to CMTT alone. Our results emphasize that tire-associated compounds become accessible in a digestive milieu and should be studied further with respect to their bioaccumulation and toxicological effects upon passage of intestinal epithelial cells.

2022

In-vitro digestion of Tire and Road Wear Particles: Bioavailability of metals and Polycyclic Aromatic Hydrocarbons

Florian Frédéric Vincent Breider, Benoît Jean Dominique Ferrari, Thibault Béranger Masset, Kristin Schirmer

The potential impact of Tire and Road Wear Particles (TRWP) on aquatic organisms has recently gained attention since the occurrence of TRWP in the aquatic environment has been observed in surface water and sediments of numerous regions. Moreover, the ingestion of TRWP by aquatic species, including fish, has recently been demonstrated. However, data regarding the bioavailability and toxicity of contaminants associated with these particles are still lacking. This study aimed (i) to characterize the solubilization potential of metals and PAHs from tire particles and TRWP in Simulated Gastric Fluids (SFG) and Simulated Intestinal Fluids (SIF) designed to mimic Oncorhynchus mykiss (Rainbow Trout) gut conditions and (ii) to assess the impact of influencing factors such as co-ingestion of natural food organic matter on the compounds solubilization. Our results showed that solubilization of all metals and PAHs were enhanced by up to 10-fold in simulated gut fluids compared to solubilization in the control (phosphate buffer saline solution). Zn was highly solubilized after an in-vitro digestion of 24-h with a final concentration in SIF of 3,2 µg.L-1 corresponding to 23 % of the total Zn concentration in TRWP. Five PAHs (Fluoranthene, Phenanthrene, Pyrene, Benzo (e) Pyrene and Benzo (g,h,i) perylene) were quantified in SIF after an incubation time of 24-h and the concentration of individual PAHs in SIF ranged from 42 to 3092 ng.L-1 which represented 0,4 – 1,5 % of their total concentration measured in tire particles. Metals were highly solubilized by the acidic SGF (pH = 2) but only a weak solubilization was observed in neutral SIF (pH = 7,4). Contrastingly, PAHs were not solubilized in SGF and highly solubilized in SIF which was proved to contain high quantities of proteins and surfactants forming micelles. This emphasise the importance of pH in the solubilization of metals and of bile surfactants and organic matter in the solubilization of PAHs from TRWP and brings new insights on the bioavailability of metals and organic pollutants from TRWP. Overall, our results demonstrate the importance of accounting for the ingestion of TRWP as an important route of exposure to metals and PAHs for aquatic organisms. As the solubilization of potential toxic compounds was greatly enhanced by the gut fluids and leads to bioavailability of these compounds, further studies investigating the uptake by the epithelial cells and transfer throughout the whole organism will be performed.

2021

In vitro acute toxicity assessment of tire tread particles using rainbow trout cell lines of gill and intestine

Florian Frédéric Vincent Breider, Benoît Jean Dominique Ferrari, Thibault Béranger Masset, Kristin Schirmer

Tire and road wear particles (TRWP) are generated by the abrasion of tires while driving, and recent questions were raised in regards to their potential contribution to microplastics released into the aquatic environment and their potential toxicological impacts. Our study aimed to determine the toxicity of TRWP and associated chemicals to fish using two rainbow trout (Oncorhynchus mykiss) cell lines representing the gill (RTgill-W1) and the intestinal (RTgut-GC) epithelium. Acute exposure to cryogenically milled tire tread (CMTT) particles and CMTT digestate following in vitro digestion was investigated. To obtain the effective concentrations (EC50) causing a 50% cell viability loss, tests were performed in accordance with OECD TG249: cell viability was assessed after 24 hours exposure using a multiple-endpoint assay indicative of cell metabolic activity, membrane integrity and lysosome integrity. Finally, chemical composition of the exposure medium was analyzed to assess which chemicals could be responsible for the observed acute effects. Our results show that metabolic activity was the most sensitive indicator of CMTT and CMTT digestate toxicity. EC50 values after direct exposure to RTgill-W1 and RTgutGC cells were estimated at 2.01 g/L and 3.80 g/L, respectively. 2-Mercaptobenzothiazole was found at the highest concentration among organic chemicals leaching out from the particles. It could be a major contributor to the loss in metabolic activity based on its toxicity to rainbow trout in vivo, although additional particle or mixture effects cannot be ruled out at this point. For digestate toxicity, EC50 was determined at 12.15 g of tire particle/L of digestive fluid. The observed toxicity could be due to high amounts of Zn, N-(1,3-dimethylbutyl)-N′-phenyl-p-phenylenediamine (6PPD), 1,3-Diphenylguanidine (DPG) or 2-Mercaptobenzothiazole leaching out during in vitro digestion. Traces of 6PPD-quinone were also detected in the exposure medium, but additional cytotoxicity assays using the RTgill-W1 cell line showed no significant toxicity of this compound up to 3 mg/L. Overall, our in vitro toxicity data agree with reports of acute fish (in vivo) toxicity and show that acutely toxic concentrations of CMTT are well above concentrations measured in river water (0.3 to 4 mg/L). However, we determined the highest no-observed-effect concentration at 10 mg/L, while up to 100 mg/L were measured in road runoffs.

2021

In-vitro digestion of Tire and Road Wear Particles: Bioavailability of metals and Polycyclic Aromatic Hydrocarbons

Florian Frédéric Vincent Breider, Benoît Jean Dominique Ferrari, Thibault Béranger Masset, Kristin Schirmer

2021

In vitro acute toxicity assessment of tire tread particles using rainbow trout cell lines of gill and intestine

Florian Frédéric Vincent Breider, Benoît Jean Dominique Ferrari, Thibault Béranger Masset, Kristin Schirmer

2021