Abstract
Crude oil is a chemically complex mixture of compounds that can disrupt endocrine and metabolic pathways in marine organisms. While polycyclic aromatic hydrocarbons (PAHs) are well-characterized drivers of toxicity, the contributions of polar fractions remain poorly understood. The identification of toxicologically relevant drivers within complex mixtures can be achieved by combining mechanistic assays focused on ligand-activated transcription factors with effect-directed analysis. This study aimed to determine the relative contribution of distinct chemical groups to overall toxicity by assessing the toxicity of a crude oil water-accommodated fraction (WAF) freshly prepared in culture medium, its extract, and chemically defined fractions. The WAF extract was fractionated into saturates, monocyclic aromatics (MAHs), naphthalenes (NAPs), PAHs, and resins by normal-phase HPLC, and characterized by comprehensive two-dimensional gas chromatography-mass spectrometry (GC×GC-MS).
Key receptor-mediated pathways were assessed including the aryl hydrocarbon receptor 2a (Ahr2a), estrogen receptor alpha (Era), peroxisome proliferator-activated receptor 1a (Ppar1a), and pregnane X receptor (Pxr), mediating xenobiotic metabolism, endocrine signaling, lipid metabolism, and detoxification. Transcriptional responses of selected biomarker genes were evaluated by exposure of Atlantic cod precision-cut liver slice (PCLS) cultures followed by qPCR assays, and cell-based in vitro luciferase reporter gene assays using cod and zebrafish receptors were used for transactivation studies. Both WAF extract and resin fraction induced cytochrome P450 1a (cyp1a) in PCLS and activated cod Ahr2a in vitro, while only WAF extract activated zebrafish Pxr. Era and Ppar1a were unresponsive to WAF extract and its fractions. Ongoing testing of selected single compounds from both PAH and resin fractions will further clarify their individual toxicity and contributions to mixture-driven effects.