Long-term Effects Of Perinatal Exposure To Bisphenol A On Dopamine Serotonin And Kynurenine Pathway In Male Mice Offspring -An Metabolomics Approach

Part 1Long-term effects of perinatal exposure to bisphenol A on metabolism in male mice offspring-An Metabolomics ApproachBisphenol A is a widespread environmental endocrine disrupters with proposed/anti-estrogen function. The structure is similar to 17β-estradiol, and it can interfere with normal synthesis and metabolism of hormones. It has been reported that BPA can through the placental barrier and influence the development of neonatal brain. However, the specific mechanism is still unclear.Objective:The main purpose of this part experiment is to discover the metabolites which are significantly changed between the control group and the model group with untargeted metabolomics approach, then enrich them to the related pathway.Methods:C57BL/6J FO pregnant mice were randomly divided in four groups, 10-12 for each group. Mice in exposure group were given subcutaneous injection of Control、2μg/kg/d、10μg/kg/d、/100μg/kg/d, once a day. Exposure time was perinatal period. F1 male offspring were giver free feeding and water until death. Brain tissure and plasma samples were analyzed with UPLC-Orbitrap instrument. The metabolic differences were screened by Principal component analysis, orthogonal partial least squares discriminant analysis and univariate statistical analysis.Results:The control group and the exposure group distinguished well with 9-month male offspring. A total of 17 metabolites had been discovered as the significantly different metabolites, and they are mainly enriched in amino acid metabolism and tricarboxylic acid cycle metabolism.Conclusions:There are Long-term effects of perinatal exposure to bisphenol A on metabolism in male mice offspring. It mainly interferes with the tryptophan metabolism, tyrosine metabolism and energy metabolism.Part 2Simultaneous quantification of neuroactive dopamine serotonin and kynurenine pathway metabolites by ultra performance liquid chromatography tandem high resolution mass spectrometryNeuroactive metabolites in dopamine, serotonin and kynurenine metabolic pathways play key roles in several physiological processes and their imbalances have been implicated in a wide range of disorders, such as Alzheimer’s disease, schizophrenia, anxiety, depression and Parkinson’s disease. The association of these metabolites’ alterations with various pathologies has raised interest in analytical methods for accurate quantification in biological fluids. However, simultaneous measurement of various neuroactive metabolites represents great challenges due to their trace level, high polarity and instability.Objective:To establish and validate a targeted metabolomics approach to simultaneous quantification of neuroactive metabolites spanning dopamine, serotonin and kynurenine metabolic pathways.Methods:3-fold acetonitril was added into the biological sample for protein precipitation. After centrifugation, the supernatant was evaporated to dryness for derivatization. After SPE, the eluate was evaporated to dryness and redissolved with initial mobile phase. The chromatographic column was ACQUITYUPLC(?) BEH-C18 (2.1×100 mm,1.7 μm). The reverse phase chromatographic separation was achieved with a gradient elution program in 20 min. The confirmation and quantification analysis was by Target-MS/MS scan mode.Results:The linearity was good with adjusted R square higher than 0.990. the limit of detection was in the range of 0.04-1 ng/mL, and the limit of quantification ranged from 0.08 to 2 ng/mL. The intra-day precision was less than 9.8%, and the inter-day precision was less than 8.9%. The recovery ranged from 85.2% to 114.0%. the analyte was stable with difference% less than 10%.Conclusions:The targeted metabolomics approach established in this part is sensitive and robust, suitable for monitoring a large panel of metabolites and for discovering new biomarkers in the medical fields.Part 3Long-term effects of perinatal exposure to bisphenol A on dopamine serotonin and kynurenine pathway in male mice offspringObjective:To study the specific effects of perinatal exposure to bisphenol A on dopamine serotonin and kynurenine pathway in male mice offspring.Methods:Apply the targeted metabolomics approach established in part 2 to quantify analysis the biological sample collected in part 1.Results:After exposure to BPA in perinatal period, DA in brain tissue and plasma was significantly decreased in male mice offspring with the increasing exposure dose and time. For tryptophan pathway, male offspring was more prone to metabolize tryptophan to serotonin route instead of kynurenine pathway.Conclusions:The offspring will still have some long-term effects with low-dose exposure to BPA in perinatal period.