In vitro human oral bioaccessibility assessment of hazardous chemicals, including N, N′-substituted-p-phenylenediamines, coming from recycled tire crumb rubber

Tires, apart from being formed by rubber and filling materials, contain organic compounds added to make them resistant and durable. The widely use of recycled tire crumb rubber (RTCR), main product of the shredding process of end-of-life tires, can cause human exposure to these chemicals due to its use in synthetic football fields and kid's playgrounds. In 2023, the European Commission banned the use of recycled tire crumb rubber in synthetic fields, giving eight years to replace the used material. This study intends to assess the oral bioaccessibility of antiozonants, crosslinking and vulcanizer agents present in RTCR. With this purpose, the Unified Bioaccessibility Method (BARGE) was used to simulate the material ingestion. RTCR is put into contact with the four simulated biological fluids including saliva, gastric and duodenal juice and bile, attempting to simulate human digestion. Afterwards, the organic compounds present in the fluid need to be extracted and Solid-phase extraction (SPE) was the technique selected after being optimized to obtain the best extraction conditions. Ultrasound assisted extraction was performed to evaluate the total concentration of the target compounds in the crumb rubber matrixes. Liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) was employed to identify and quantify the target compounds. The results showed the bioaccessibility of all the studied analytes, with values ranging from 0.1 % up to 70 %. Benzothiazole was the compound with the highest bioaccessibility with a mean value of 40 % and concentrations reaching 32000 ng g^-1 in the bioaccessible fraction. N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine and its transformation product ((4-Methylpentan-2-yl)amino)-5-(phenylamino)cyclohexa-2,5-diene-1,4-dione) showed an average bioaccessibility of 0.1 % and 1.8 %, respectively, the latter being present in all the analysed samples.