Metabolism and Export of Lipid Peroxidation Products as Assayed in Human Monocytic THP-1 Cells Using Electrospray LC-MS/MS

摘要

We have developed LC-MS/MS methods to analyze the effect of Asc supplementation on the production and export of phase I and phase II metabolites of HNE in the cell lysate and medium. Identity confirmation were made based on comparison of EPI scans of biological metabolites to those of the appropriate synthetic standards prepared in our laboratory. Exposing Asc pretreated Human monocytic THP-1 cells to HNE(Ac)_(3) revealed increased formation of the glutathione conjugates. Adducts of Asc with HNE were also detected in medium and lysate samples. Use of HNE[Ac]_(3) requires hydrolysis to restore HNE reactivity strongly indicating the endogenous formation of these ascorbyl-adducts. This approach represents a comprehensive method for metabolic profiling of HNE in human cells using a prodrug-like substrate, and electrospray LC-MS/MS in order to further understand the detoxification and elimination affects of ascorbic acid in this system.