Toxic Effects And Mechanisms Of Polystyrene Microplastics And Aryl Phosphate Flame Retardants On Marine Microalgae

Both microplastics and organophosphate flame retardants(OPFRs)are emerging pollutants and widely distributed in the ocean.With the increase in the consumption of plastics and OPFRs,the pollution of the two pollutants is getting worse.Microplastics and OPFRs have toxic effects on marine organisms,causing a huge threat to the marine ecosystem.In addition,microplastics can carry and transport OPFRs in seawater,making the effects of the two pollutants on marine life more complex.As primary producers,marine microalgae are of great significance for maintaining the stability of the marine ecosystem,and they are typical test species for toxicological study.There have been some studies on the toxicity of microplastics to microalgae,however,most of them are limited to the detection of physiological indicators.While there are very few studies on the toxicity of OPFRs to marine microalgae.Based on the research status,the single and combined toxic effects and mechanism of microplastics and aryl OPFRs on marine microalgae were investigated.The purpose of this study is to provide basic data and theory for evaluating the impacts of microplastics and OPFRs on marine environment.The main findings are as following:(1)The effects of microplastics on the physiology and metabolic pathways of Dunaliella salina(D.salina)were investigated and clarified.Based on the results of toxic experiment,both polystyrene(PS)and aged polystyrene(A-PS)inhibited the growth of D.salina,but promoted the pigment synthesis in algal cells.The oxidative stress analysis indicated that PS and A-PS with high concentration(≥ 50 mg/L)induced high production of reactive oxygen species(ROS),and caused oxidative damage to algal cells.Untargeted metabolomics analysis indicated that PS and A-PS significantly increased the differential metabolites.KEGG pathway enrichment analysis showed that PS and A-PS perturbed the amino acid-related metabolic pathways.In particular,the amino acid biosynthesis and ATP-binding cassette(ABC)transporter pathways were significantly up-regulated,thus promoting nitrogen storage and transmembrane transport in D.salina.Transmembrane transport requires a large amount of ATP;as a result,algal cell division is inhibited.These results explained the toxicity of microplastics to microalgae at the physiological and metabolic levels more comprehensively.In addition,comparing the results,it was found that the interaction between A-PS and algal cells was stronger,inducing higher ROS level,which led to more serious oxidative damage and stronger oxidative stress response.(2)The toxic effects of 2-ethylhexyl diphenyl phosphate(EHDPP),tricresyl phosphate(TCP),and triphenyl phosphate(TPhP)on Chaetoceros meulleri(C.meülleri)at the physiological and transcriptomic levels were systematically studied to illustrate the toxicity mechanisms.Within the experimental concentration range,the OPFRs caused a positive dose effect on the cell growth inhibition and algal cell malformation.Detection results also showed that OPFRs induced a decrease of chlorophyll a content and rETR,resulting in a significant increase of ROS.The excessive ROS induced membrane lipid peroxidation,thus causing irreversible damage to the algal cell membrane structure and function.In addition,affected by OPFRs,the results of transcriptome sequencing showed that the expression level of several genes related to photosynthetic proteins were significantly down-regulated,and gene expression involved in glutathione peroxidase,superoxide dismutase,and glycerophospholipase were significantly up-regulated.Besides,EHDPP,TCP and TPhP induced significantly down-regulation of gene expression in ribosome,resulting in the destruction of its structure and function.The expression of gene involved in proline biosynthesis was also down-regulated,as a result,the resistance ability of algal cells to OPFRs stress was reduced.The above results illustrated that the toxicity of EHDPP,TCP and TPhP to C.meülleri was attributed to photosynthesis inhibition,oxidative damage,and protein synthesis blocking.Comparing the structural properties of the three OPFRs and their toxicity to the microalgae,it was found that OPFRs with higher hydrophobicity had stronger interaction with algal cells,resulting in higher toxicity.(3)Here,the effects of the single and combined toxicity of PS microplastics and TPhP on the cell growth,photosynthesis,and oxidative stress of D.salina and C.meulleri were investigated.PS microplastics inhibited the growth of algae cells and caused a dose-dependent effect on oxidative stress and severe cell membrane damage.However,there was no obvious decrease in chlorophyll a content,and PS at high concentration significantly promoted the synthesis of chlorophyll a.The TPhP(≥1.6 mg/L)also inhibited cell growth and caused cell membrane damage,except at low concentrations(≤ 0.8 mg/L),which stimulated algae growth in a short time.Moreover,no obvious decrease in chlorophyll a and maximal photochemical efficiency of PSII was found in the TPhP experimental groups except for TPhP at the highest concentration.In the combined toxicity experiment,it was found that PS microplastics could adsorb TPhP,which reduced the concentration of TPhP in the culture medium and the interaction between PS microplastics and algal cells,thus reducing the toxicity of TPhP to microalgae.Using the additive index and independent action methods,the combined toxic effects of PS and TPhP on the algae were evaluated as antagonistic.