Study On Toxic Effects And Molecular Response Mechanisms Of Typical Non-steroidal Anti-inflammatory Drugs On Scenedesmus Obliquus And Oryza Stiva.L

Due to the massive use and improper disposal of pharmaceuticals and personal care products（PPCPs）,their eco-toxic effects have attracted widespread attention.This article study four non-steroidal anti-inflammatory drugs（NSAIDs）with different structures:Dex-troibuprofen（S-（+）-IBU）,racemic ibuprofen（rac-IBU）,aspirin（ASA）and ketoprofen（KEP）on the toxic effects on Scenedesmus obliquus and rice（Oryza sativa L.）,and to explore the effects of NSAIDs on plants.The toxic effects and internal mechanisms are expected to pro-vide a theoretical basis for the evaluation of the ecological effects of NSAIDs.The main con-tents include:（1）The growth inhibition of NSAIDs on S.obliquus is positively correlated with the concentration.Low-concentration NSAIDs can promote the growth of algae and the growth inhibition rate continues to increase and causes irreversible damage under high-concentration.The relative toxicity of the four NSAIDs is:KEP>ASA>rac-IBU>S-（+）-IBU.Transmission electron microscopy showed that cell ultrastructure was damaged under NSAIDs stress,and there were plasmolysis and swelling of chloroplasts.The content of photosynthetic pigment（Chla,Chlb and Caro）is negatively correlated with the concentration of NSAIDs;the chloro-phyll fluorescence parameters（basic fluorescence F₀,maximum fluorescence F_m,maximum photochemical quantum yield F_v/F₀ and maximum photosynthetic efficiency F_v/F_m）and fast light response curve（actual quantum yield Y（II）,photochemical quenching coefficient（q P）and relative electron transport rate（ETR））are in a concentration-effect relationship with NSAIDs;photosynthesis and respiration rates show a downward trend with the increase of NSAIDs concentration,and the expression of genes related to the thylakoid electron transport chain is down-regulated,indicating NSAIDs cause significant obstacles to electron transfer and CO₂ fixation.There is a positive correlation between the content of reactive oxygen radi-cals（ROS）and the concentration of NSAIDs.The activity of superoxide dismutase（SOD）,catalase（CAT）and the content of antioxidant glutathione（GSH）showed a trend of first in-creasing and then decreasing.The results of the integrated biomarker index（IBR）analysis are consistent with the conclusions of acute toxicity and photosynthesis.（2）0.5 mg L^-1 KEP stimulated the growth of rice seedlings slightly.With the increase of KEP concentration,the growth of rice seedlings was inhibited,the ultrastructure of the seed-lings was severely damaged,the starch grains increased significantly and a lot of vacuoles appeared;and the dead cells of the roots increased which forming a jelly-like film,and in-duced damage to plasma membrane proteins and changed membrane performance.Chloro-phyll fluorescence parameters including F_v/F_m,F_v/F₀,Y（Ⅱ）,ETR,adjusted energy dissipation quantum yield（Y（NPQ））and non-adjusted energy dissipation quantum yield（Y（NO））all changed,the key genes（HEMA,HEMG,CHLD,CHLG,CHLM and CAO）of photosynthetic pigment synthesis are regulated,and the content of photosynthetic pigments is reduced;the expression genes in the photosynthetic electron transport chain are all regulated,and electron transfer is hindered,photochemical energy conversion and self-protection regulation mecha-nisms are weakened;and it is speculated that when KEP is exposed,the activation of the PGRL/PGR5 pathway will help the cyclic electron transfer（CET）to produce ATP required and maintain physiological activities under stress.The ATP content decreased;H⁺-ATPase enzyme was activated first and then decreased;OSA gene IV family（OSA6,OSA10）and III family（OSA9）encoding H⁺-ATPase enzymes are significantly up-regulated in shoots and roots,which can be better response to KEP stress.KEP causes a large amount of O₂^-·to ac-cumulate in rice seedlings,along with the increase in the content of peroxides and osmotic substances;SOD,CAT,ascorbate peroxidase（APX）activity and GSH content change ac-cordingly to maintain normal ROS levels to protect cells from oxidative stress.Antioxidant enzyme genes（CSD1,CSD2,FSD1,MSD1,CAT1,CAT2,CAT3,APX1 and APX2）coordinate with each other under KEP stress and may help fine-tune the defense system to maintain bal-ance.IBR analysis showed the increase of KEP concentration led to a gradual increase in stress pressure on rice seedlings.（3）The content of nitrate nitrogen（NO₃^--N）in roots continued to decrease with the in-crease of KEP concentration,while the NO₃^--N in shoots increased first and then decreased,which may be a positive response to photosynthesis.The nitrate reductase（NR）activity in the roots showed a trend of increasing and then decreasing.The NR activity in shoots continued to increase;the NO₃^--N content in shoots was inconsistent with the NR activity,which may be due to tissue specificity and assimilation regionalization;Ammonium nitrogen（NH₄⁺-N）con-tent first increased and then decreased;nitrite reductase（Ni R）activity in shoots gradually de-creased,while Ni R activity in roots first increased and then decreased;the trend of glutamine synthetase（GS）is similar to Ni R;the activity trend of glutamine synthetase（GS）and gluta-mate synthase（GOGAT）is similar to Ni R;the response kinase-glutamate dehydrogenase（GDH）continues to decrease in roots,and shows a trend of rising first and decreasing in shoots,the total amount of free amino acid presents a gradual accumulation trend.PCA showed KEP can significantly change the correlation of photosynthetic pigments,energy synthesis and nitrogen assimilation in rice seedlings,and peroxide production is one of the important signs of stress.Root microbiome showed KEP resulted in significant changes in the root microbial community structure,and increased community diversity and richness.The abundance of the original dominant and probiotic bacteria of the root system such as Proteus,Bacillus and Sphingomonas sharply decreased and the abundance of Methylophilus and Xan-thobacter increased significantly.This may be due to the change of the root exudates by KEP.The PICRUSt indicates the richness and enrichment of the KEGG pathway of photosynthesis,antioxidant stress,nitrogen assimilation and energy metabolism decreased after KEP treat-ment,and the predicted enzyme content decreased,indicating KEP may exert selective pres-sure on root microorganisms and change the plant-microbe interaction and thereby affecting the growth of rice seedlings.