From Transient to Metastable: Generation, Characterization, and Biomimetic Reactivity Studies of Well-Defined Heme Peroxynitrite Model Systems

Mechanistic intricacies of peroxynitrite (PN)-mediated transformations remain fundamentally unresolved, despite their proposed roles in cancer, cardiovascular and autoimmune diseases, neurodegeneration, and inflammatory reaction cascades among others. Central to these biological phenomena are heme proteins, giving credence to the pivotal nature of heme-bound PN adducts. Nonetheless, the transient nature of heme-PN intermediates has significantly convoluted their capture and characterization, leaving an impactful deficit in contemporary literature. We herein report the generation of three tetraarylporphyrin-based heme-PN adducts that are metastable at -80 °C, via the reactivity of heme peroxo adducts with nitrosonium cations. Notably, this method allows the generation of heme-PN using only stoichiometric equivalents of nitrosonium, which presumably plays a key role in escalating their stability in solution. These new heme-PN adducts have been characterized with a medley of spectroscopic (UV-vis, NMR, EPR, and resonance Raman) and theoretical (DFT) methods, as well as mass spectrometric tools including isotope-labeling studies. Moreover, a broad variety of organic substrate reactivities of heme-PN is demonstrated herein, making close parallels to long-standing proposed biological implications of those adducts. Kinetic investigations into heme-PN-mediated phenol nitration reactivities are also reported. Spectroscopic and theoretical investigations unambiguously support O-O bond homolysis as the predominant pathway of heme-PN decay, leading to the formation of ^•NO₂ radicals. This work, therefore, shines light on important aspects of heme-PN chemistries pertaining to its formation and reactivity toward various biomimetic substrates.