Sharon Idiga, Pradip Mascharak.
University of California, Santa Cruz, Santa Cruz, CA.
Peroxynitrite (PN) is a reactive nitrating and oxidative species that plays a key role in inflammation-mediated disease states, from neurodegenerative disorders to diabetes. PN is formed in the radical terminating reaction between nitric oxide (NO) and superoxide. It has been difficult to observe PN directly within cells due to its transient nature. With a photo-controlled peroxynitrite-generating platform, the Mascharak lab has developed a system that is able to release PN in situ, to better understand the dynamic chemistry of PN. Stable metal nitrosyl, [Mn(PaPy3)(NO)]ClO4 (PaPy3 = N,N-bis-(2-pyridylmethyl)amine-N-ethyl-2-pyridine-2-carboxamide), was used as the source of NO because it rapidly releases NO on illumination with visible light. To match the rapid release of NO from the nitrosyl, the xanthine oxidase/hypoxanthine (XO/HX) system was employed as the source of superoxide. Both sources were encapsulated in a sol-gel matrix to avoid contamination. A multi-well plate was used to study the chemistry of the precursors as well as PN. Construction of the PN-generating platform was completed by layering both NO and XO sol-gels at the bottom of individual wells. Release of the precursor molecules is triggered with exposure to low powered visible light and the addition of HX. This light-activated platform allows the study of PN-mediated chemistry under varied flux ratios and pH conditions. We plan to use the multi-well platform to study the reactions of PN with various biomolecules such as amino acids, peptides, and lipids.