Biodegradation Mechanism Of Tricresyl Phosphate By Brevibacillus Brevis And Toxicity Of Its Metabolites

Organophosphate flame retardants?OPFRs?are a kind of phosphorous flame retardants widely used.In recent years,OPFRs have attracted widespread attention because they are easily released into the environment and have toxic effects,posing a potential threat to the environment and human health.Up to date,only limited studies have focused on the biodegradability and degradation mechanism of OPFRs.In the present study,tricresyl phosphate isomers?TCPs?were used as representative contaminant of OPFRs,and Brevibacillus brevis was selected as the experimental strain.The objective of this work was to investigate the biodegradation characteristics and transformation mechanism of TCPs by B.brevis.In addition,the cytotoxic effects and mechanism of TCPs and its metabolites were also analyzed.The detailed conclusions are as follows:?1?B.brevis could degrade TCPs effectively and the degradation efficiency of 1 mg/L TCPs by B.brevis?2 g/L?reached 86.52%at pH 7 and 30°C after 5 days of incubation.During the degradation process,cell morphology changes,cell functional group hydroxyl?-OH?,fatty acids,organic phosphates,etc.will affect the degradation of TCPs by B.brevis.Moreover,the addition of cytochrome P450 inhibitor markedly decreased the degradation of TCPs by B.brevis.Meanwhile,high concentration of TCPs induced oxidative stress on the cells of B.brevis,which caused the increase of ROS content.Bioaugmentation with B.brevis could stimulate the TCPs degradation in river water microcosms and TCPs decreased by approximately 91.3%after 120h of incubation.?2?B.brevis could degrade TCP isomers effectively and the degradation efficiency of 1mg/L TpCP,TmCP,ToCP reached 93.91%,82.91%and 53.92%,respectively,within five days.Moreover,the intracellular enzymes functioned critically during degradation.The major metabolites of TCPs were identified using high-resolution liquid chromatography-qualitative quadrupole time-of-flight mass spectrometry?LC-Q-TOF-MS?within 5 days of degradation.The degradation pathways of TCPs by B.brevis were proposed based on the detected metabolites,which mainly involved hydrolysis and hydroxylation processes.In the process of biodegradation,B.brevis metabolism caused the release of Na⁺and Cl^-as well as the absorption of some nutrient ions including K⁺,PO₄^3-,Mg²⁺,and SO₄^2-;the presence of oxalic acid,citric acid,acetic acid,and malonic acid was also detected.?3?TmCP and ToCP had different degrees of inhibition on the viability of A549 cells.Meanwhile,TmCP and ToCP destroyed the integrity of mitochondrial membrane and further induced oxidative stress in cells,resulting in decreased mitochondrial membrane potential and inducing apoptosis of A549 cells.High concentration of TmCP and ToCP induced cell cycle arrest at G0/G1 phase in A549 cells,and decreased the proportion of cells at S phase.However,TpCP and the metabolites of TCPs had no significant toxic effects on A549 cells.The results in this study revealed the mechanism of microbial degradation and transformation of TCPs,and analyzed the potential toxic effects of TCPs and its metabolites on the environment and human health.This study also provided the data support for developing bioremediation technology for environments contaminated with OPFRs.