Mechanistic understanding of the toxic effects of tri-n-butyl phosphate (TnBP) and tricresyl phosphate (TCP) to Escherichia coli: Evidence from alterations in biomarker expression and perturbations of the metabolic network

Tri-n-butyl phosphate (TnBP) and tricresyl phosphate (TCP), emerging flame retardants and plasticizers, have garnered increasing attention due to their potential risks to ecosystem. A few researches regarding the toxicological mechanisms of TnBP and TCP had been performed, while molecular-level toxic effects of them and metabolic response using microbial models are the lack of relevant investigation. Thus, we investigated the cytotoxicity, oxidative stress response, and metabolic response in E. coli exposed to TnBP and TCP. Exposure to them significantly increased the activities of antioxidant enzymes, indicating activation of the antioxidant defense system. Excessive accumulation of reactive oxygen species (ROS) triggered various biological events, including a reduction in membrane potential (MP), decrease of adenosine triphosphatase (ATPase) activity, and increased malondialdehyde (MDA) content. These findings suggested that oxidative damage compromised membrane proteins function, membrane stability, and intracellular homeostasis. GC–MS and LC-MS-based metabolomics analyses revealed that TnBP and TCP strongly disrupted multiple metabolic pathways, including carbohydrate metabolism, nucleotide metabolism, lipid metabolism, beta-alanine metabolism, pyruvate metabolism and oxidative phosphorylation. These disruptions highlighted the inhibitory effects on molecular functions and metabolic processes. Notably, lipids biomarkers e.g., PC(11:0/16:0), PA(17:1(9Z)/18:2(9Z,12Z)), PE(17:0/14:1(9Z)), and LysoPE(0:0/18:1(11Z)) were significantly altered, verifying that the regulation of lipid-associated metabolite synthesis plays a protective role in maintaining cellular membrane function. In summary, this study enhances our understanding of TnBP and TCP toxicity in E. coli, providing novel insights into their toxicological mechanisms at molecular and network levels. These findings underscore the ecological risks posed by organophosphate flame retardants in aquatic ecosystem.