Toxicokinetics and metabolomics analysis to evaluate the effect of DEHP exposure on sex and age in rats

Abstract

Di-(2-Ethylhexyl) phthalate (DEHP) is a ubiquitous environmental endocrine disruptor that adversely affects homeostasis, reproduction, and developmental processes. DEHP can be inevitably penetrated through the air, food, cosmetics, and plastic devices. After absorption, it shortly converts to Mono-2-ethylhexyl phthalate (MEHP) which has higher toxicity. The previous studies to date clarify the damage derived from exposure to DEHP and its metabolites have been shown to differ by sex and age, emphasizing the importance of understanding the toxicokinetic and metabolomic activity of these chemicals. However, the underlying mechanisms are still unclear. In this study, we treated 5mg/kg of DEHP by intravenous injection or oral administration to adult male and female rats (9 weeks) and young female rats (5 weeks) and obtained plasma, urine, and 6 tissues (lung, kidney, liver, testicle, womb, and ovary) for observing toxicokinetic processes of the major metabolites of DEHP: MEHP, mono-(2-ethyl-5-hydroxy-hexyl) phthalate (MEHHP), mono-(2-ethyl-5-oxo-hexyl) phthalate (MEOHP). In addition, an untargeted metabolomics analysis was performed using urine collected at five different times (0, 4h, 8h, 12h, and 24h) to present differentiated metabolic pathways by exposure to DEHP. For the determination of the significant variables, multivariate statistical analysis using ANOVA-simultaneous component analysis (ASCA) and partial least squares-discriminant analysis (PLS-DA) were used, and we found that metabolic profiles were significantly different in sex and age. From the important metabolites that induced the difference on sex and age over time, it can be suggested that steroid hormone biosynthesis, taurine and hypotaurine metabolism, linoleic acid metabolism and primary bile acid biosynthesis were affected by the absorption of DEHP. Overall, this study systematically describes the comprehensive mechanism of DEHP exposure to sex and age and highlights the power of toxicokinetic model design and appropriate multivariate statistical analysis using ASCA combined with PLS-DA in the discovery of key metabolomics pathway. This study can provide the scientific reference for understanding sex and age-dependent toxicokinetics and metabolic pathways following exposure to DEHP.