Towards the validation of an in vitro method to predict the bioavailability of soil organic contaminants

For several decades, the presence of organic contaminants such as polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs) or pesticides such as chlordecone (CLD) in ecosystems has raised significant concerns regarding environment and human health. Humans may be exposed to these substances through unintentional ingestion of soil (hand-to-mouth contact particularly common in children [1]), potentially leading to health hazards. Considering the total contaminant concentration in soil may lead to an overestimation of the risk, as only the bioaccessible fraction released into the digestive tract can be absorbed. Predicting the bioavailable fraction by estimating oral bioaccessibility is relevant for accurate risk assessment [2]. Bioaccessibility can be estimated by in vitro methods simulating human digestion, already available for trace metal elements (for ex. Unified Bioaccessibility Method). The PBET (Physiology Based Extraction Test) method simulates a stomachal digestion following by an intestinal digestion. Recently, an adaptation for hydrophobic organic contaminants has been suggested by adding Tenax (absorbent sink) in the intestinal phase, improving the extraction of organics liberated from soils (for ex. DDT pesticide [3], PCBs [4], PAHs [5]). The Ti-PBET (Tenax-improved PBET) method shows promising results, but its use may be limited by the high cost of Tenax. The general objective of our work was therefore to improve the Ti-PBET test in terms of both practicability and cost. We compared different adsorption supports for extracting PAHs and PCBs, and found XAD-2 performed comparably to Tenax while being more cost-effective; XAD-2 was therefore selected for subsequent test improvements. To simplify resin recovery after intestinal digestion, we tested two ways of incorporating XAD-2 into the in vitro assay: (1) enclosed in a tea bag; (2) freely dispersed within the flask. No significant differences were observed between the two types of control samples (p > 0.05), although certain compounds such as PCB 28 and fluorene showed higher chromatogram signals with the tea bag method. Although more convenient, this option was abandoned as it increased the risk of control contamination and compromised the accuracy of bioaccessibility estimates. We finally tested the Xi-PBET (XAD-2–improved PBET) method by spiking PCBs at 1, 5, 10, 50, and 200 ng/mL (n = 3) to assess the recovery rates of six congeners (PCB 28, 52, 101, 138, 153, 180) and GC–MS/MS quantification accuracy across this concentration range. Recovery rates ranged from 87–132% (CV ≤ 12%) between 5 and 200 ng/mL, while the 1 ng/mL concentration showed higher overestimation and variability (120–188%; CV ≤ 69%) likely due to low spiking volumes and background noise. These results confirm that 500 mg of XAD-2 effectively captures the entire free PCB fraction between 5 and 200 ng/mL without saturation of the resin, and that GC–MS/MS is reliable for this concentration range. This work has demonstrated that XAD-2 resin is a good alternative to Tenax for reducing the cost of bioaccessibility testing without compromising its effectiveness, making such testing more accessible to the contaminated sites and soils sector. However, in vitro bioaccessibility methods still require improvement to provide reliable estimates. Therefore, our future work will compare Xi-PBET against other in vitro assays using a wide range of soils contaminated with different pollutant families, and validate results against bioavailability measurements obtained from in vivo experiments on piglets (the best model to simulate children’s physiology). The in vitro methodologies are expected to provide a valuable tool for site and soil managers in health risk assessment