Degradation Mechanism Of Tris-(2-chloroisopropyl) Phosphate By Ultraviolet Driving Aops And Toxicity Evaluation Of Its Degradation Products

Organophosphate flame retardants?OPFRs?have been raising concern as a class of emerging organic contaminants?EOCs?due to persistence,bioaccumulation and bio-toxicity.However,only limited researches regarding potential human health risk evaluation,the degradation mechanism of OPFRs using UV-AOPs and their degradation products toxicity were published.In current study,TCPP?tris-?2-chloroisopropyl?phosphate?was selected as the target contaminant and A549 cell was used as the common model cell lines.The objectives of present work were to evaluate the cytotoxicity and toxicity effects of TCPP compared with other different flame retardants and explore the degradation mechanism and transformation patterns using three different UV-AOPs.Additionally,the toxicological evaluation on cellular levels and molecular levels by flow cytometry and proteomics was performed.The detailed conclusions are as follows:?1?Different inhibitory effects on cell viability of A549 cell were induced by TCPP,TCEP,TCP,TPHP,TBBPA,BDE-47 in both dose-and time-dependent manner,among which BDE-47 exhibited the powerful inhibition and TCEP was the weakest.High dosage OPFRs and BFRs??100?mol/L?significantly induced the increase of the reactive oxygen species?ROS?,the reduction of mitochondrial membrane potential?MMP?to cause the MMP dysfunction,which induced the oxidative stress to result in cell apoptosis and the elevation of intracellular free Ca²⁺levels.The cytotoxicity induced by different flame retardants were mediated by oxidative stress.Simultaneously,the cell apoptosis induced by OPFRs and BFRs was caspase-3 dependent mitochondrial pathway.?2?The degradation reaction of TCPP using UV/TiO₂,UV/PS,UV/H₂O₂ followed a pseudo-first order kinetics with an apparent rate constant(k_obs)were 0.3146/min,0.1653/min,0.1328/min,respectively.The removal efficiency were separately 99.97%?UV/TiO₂?,99.91%?UV/PS?and 96.1%?UV/H₂O₂?with a relatively lower mineralization rates.A rapid removal efficiency was observed in a neutral condition,the removal efficiency of TCPP was conspicuously inhibited under alkaline condition.The removal efficiency of TCPP was significantly declined with the increase of Cl-,NO₃-and HA concentrations,moreover HA exhibited powerful inhibitory impact on TCPP removal than that of Cl-,NO₃-.Additionally,six degradation products?UV/TiO₂?,twelve degradation products?UV/PS?,nine degradation products?UV/H₂O₂?were identified based on the HRMS analysis,respectively,including the hydroxylated,dechlorinated and keto derivatives.The major reaction mechanism involved oxidation,hydroxylation,dechlorination and dealkylation.The degradation products included C₆H₁₃Cl₂O₄P?m/z 251.0002?,C₃H₈ClO₄P?m/z 174.9922?,C₉H₁₇Cl₂O₅P?m/z 307.0266?,C₉H₁₇Cl₂O₆P?m/z 323.0217?,C₆H₁₂ClO₆P?m/z 247.0134?,C₉H₁₈Cl₃O₅P?m/z 343.0033?,C₉H₁₆Cl₃O₅P?m/z 340.9876?,C₆H₁₃Cl₂O₅P?m/z 266.9954?,C₃H₈ClO₅P?m/z 190.9877?,C₃H₄ClO₃P?m/z 154.9669?,C₉H₁₈ClO₆P?m/z 289.0587?,C₉H₁₈ClO₄P?m/z 257.0708?.Based on EE/O analysis,the pretreatment of HA will facilitate to high-efficiently remove TCPP using UV-AOPs.?3?The toxicity evaluation of TCPP and its degradation products on E.coli using flow cytometer suggested TCPP can be transformed to other less toxic and non-toxic small molecule metabolite,which reduced oxidative stress on bacteria,declined the intracellular ROS levels,altered the intracellular and extracellular distribution of Na⁺/K⁺,improved the intracellular ion homeostasis dysregulation and elevated the membrane potential.Meanwhile,cell apoptosis exhibited a decrease tendency and inhibitory effect on phase C?biosynthesis phase of DNA,amino acid biosynthesis and metabolism?was weakened,all of which promoted the normal metabolism of microorganism.?4?Based on the analyses of differential proteins using iTRAQ,144 proteins and 154proteins were significantly differential expressed after exposure to 10 min degradation products and 60 min degradation products,respectively.40 up-regulated proteins synthesis and 104 down-regulated proteins synthesis in E.coli were induced by intermediate mixture produced in the early stage?10 min?,whilst 64 up-regulated proteins and 90 down-regulated proteins expression were triggered by the further degradation products?60 min?.These differential proteins mainly involved in biosynthesis,cellular nitrogen metabolism,cellular stress response,amino acid metabolism,catabolic processes,membrane transport processes and DNA transcriptional translation processes,and precursor metabolite synthesis and energy production processes,etc.The degradation products stimulated the polysaccharide and pentose phosphate metabolism,resulting in the up-regulation expression of metabolic pathway.Simultaneously,they induced the up-regulation expression of proteins correlative with DNA,RNA and ATP biosynthesis.The up-regulation expression of fabA,fabD,fabF and fabZ promoted the biosynthesis of unsaturated fatty acids and phosphatidic acid.Besides,degradation products also induced the accumulation of fumarate,succinate,malate and oxaloacetate in the TCA cycle,which facilitated up-regulation expression of the metabolic pathways of oxidative phosphorylation and amino acid/lipid biosynthesis.Meanwhile,universal stress proteins such as uspA,uspC,uspE,uspF and uspG exhibited down-regulated expression proved that degradation intermediate mixture exhibited lower toxicity than original TCPP.The results in current study revealed the cytotoxicity of TCPP,TCEP,TCP,TPHP,TBBPA,BDE-47 and the toxicity mechanism.Additionally,the reaction mechanisms of TCPP using different UV-AOPs were separately clarified,and the interference of TCPP and its degradation products on cellular metabolism,molecular networks and phylogenetic level was revealed using flow cytometer and iTRAQ.This study also provided a novel toxicological evaluation manner for the safety of the UV-AOPs technology.