Stereoselective Effects Of Ibuprofen On Adult Zebrafish Based On Omics

Ibuprofen(IBU),as an over-the-counter drug is commonly used to relieve symptoms of pain and fever,and it is also a new type of environmental pollutant that is widely present in the water environment—pharmaceutical and personal care products(PPCPs).Studies have shown that ibuprofen can cause oxidative damage,immune dysfunction and nerve damage.Ibuprofen is a chiral drug and both R-(-)-ibuprofen and S-(+)-ibuprofen are detected in the environment,and there are differences in the nature and toxicity.Therefore,it is of great practical significance to study the effects of ibuprofen on the stereoselective toxicity of aquatic organisms.At present,there are few studies on the stereoselective toxicity of ibuprofen enantiomers.In this dissertation,zebrafish was used as a model organism to explore the stereoselective toxicity mechanism of ibuprofen racemates and enantiomers on adult zebrafish.The following conclusions were drawn:In this study,ibuprofen was separated by high performance liquid chromatography to prepare the enantiomer needed for the exposure experiment.And the stability of R-(-)-Ibuprofen and S-(+)were examined within 24 hours under different conditions and in different solvents.The results show that the enantiomers of IBU are stable,and they can be used for exposure tests.The UPLC-Qtof-MS was used for non-targeted metabolomic analysis of polar small molecule metabolites in zebrafish brain tissue after exposure.A total of 45 biomarkers and 26 related metabolic pathways were screened,mainly involving energy metabolism,oxidative stress,gene regulation,neural development,and cell growth.In addition,22 amino acids were quantified by LC-MS/MS.As a result,it has been found that tyrosine,tryptophan,and phenylalanine involved in the development of the nervous system,cysteine,glycine,and methionine,which are related to oxidative stress,and immune functions such as arginine,and glutamic acid,glutamine and aspartic acid associated with energy metabolism,all changed.And they all showed obvious stereoselective differences after exposure to different enantiomers of ibuprofen.The UPLC-Qtof-MS was used for lipidomic analysis of lipid molecules in zebrafish brain tissue samples after the end of exposure to ibuprofen.46 biomarkers,mainly glycerophospholipids,cholesterol esters,pregnenolone,glycerolipids,and sphingolipids,were identified.At the same time,metabolic pathway analysis was performed.In addition,46 sphingolipid quantification methods were established by means of Qtrap to quantify 46 sphingolipids in the brain of zebrafish after exposure.The results showed that there were significant differences between groups in 16 sphingolipids.The results showed that long-term ibuprofen exposure can lead to lipid metabolism disorders in adult brain tissue,and there are obvious stereoselective differences in the effects of different ennatiomers of ibuprofen on the type and content of lipid metabolites in the brain of zebrafish.Zebrafish brain tissues were harvested at 7 days,14 days and 28 days after exposure,and the changes of superoxide dismutase(SOD),catalase(CAT)and glutathione thioltransferase(GST)were measured.Results showed that no significant changes in SOD were found on 7th and 14th day,but significant upregulation was occurred on 28th day with no obvious stereoselective effects among rac-/R-(-)-/S-(+)-IBU treated samples.No significant changes in CAT were found on 7th and 14th day,but significant upregulation was occurred on 28th day with significant stereoselective effects among rac-/R-(-)-/S-(+)-IBU treated samples.GST was affected on 7th with no obvious stereoselective effects,but significant stereoselective effects were found on 14th and 28th day.It can be concluded that long-term ibuprofen exposure may lead to a certain degree of oxidative damage in the brain of adult zebrafish,and enantiomers of IBU exhibited significant stereoselective effects.In conclusion,our study proved that chiral IBU at environmental level can induce metabolic disorders and oxidative stress based on metabolomics and lipidomics.And the application of metabolomics and lipidomics also provided a new insight into toxicologic study of other PPCPs.