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Access Type

WSU Access

Date of Award

January 2011

Degree Type

Dissertation

Degree Name

Ph.D.

Department

Physiology

First Advisor

Husam M. Abu-soud

Abstract

Hypochlorous acid (HOCl) is a potent oxidant generated by the hemoprotein myeloperoxidase. Although HOCl plays an important role in the innate immune response,sustained high levels of HOCl has been implicated to play a harmful role. In several pathological conditions such as atherosclerosis, endometriosis and sickle cell disease where HOCl is elevated there are reports of significant free iron accumulation. Free iron is toxic since it can lead to the generation of other secondary free radicals such as hydroxyl radical by Fenton reaction. The exact source and mechanism by which the free iron is generated is not clearly understood. This work establishes a mechanistic link between high HOCl and elevated free iron. Our results show that HOCl can mediate heme destruction from hemoglobin and red blood cells to liberate free iron. HOCl was also shown to aggregate the protein moiety of hemoglobin. To understand the chemistry of the process to a greater extent, the reaction products obtained when hematin and protoporphyrin was reacted with HOCl, was compared to those obtained when purified hemoglobin or red blood cells were treated with HOCl. Detailed HPLC and mass spectrometric analysis of the reaction products revealed that the presence of the metal center does not play any role in the pattern of cleavage of the tetrapyrrole ring. Subsequent studies showed that treatment of cyanocobalamin with HOCl, also led to generation of proinflammatory reaction products such as cyanogens chloride. Given the role of HOCl in generating free iron and cyanogen chloride it would be beneficial to scavenge HOCl. In this respect, we report that lycopene a common dietary carotenoid displays excellent HOCl scavenging property. Owing to the extensive hyperconjugation in its structure, one molecule of lycopene theoretically has the

capability to scavenge 39 molecules of HOCl. Currently the only known biomarker of HOCl is 3-chlorotyrosine and 3, 5-dichloro tyrosine, both of these two are hydrophilic. But the reactivity of HOCl extends to the lipophilic compartments as well. Lycopene and its oxidation products are lipophilic in nature. Therefore lycopene oxidative fragments have the potential to be used as a biomarker for HOCl mediated damage to the lipophilic compartments in the cell.