Oxygen Assays Articles
Hydrogen Peroxide Detection: An Overview
Hydrogen peroxide, like superoxide, can react with a variety of targets in the cell, and has been associated with a number of diseases. Aerobic organisms express catalase and peroxidase enzymes to prevent damage by H2O2. Peroxidases—most of which contain a heme at their active site—undergo an enzymatic cycle in which a molecule of peroxide oxidizes the enzyme, releasing one of the peroxide oxygen atoms as water, and leaving the other complexed to the enzyme-bound iron. This configuration is known as Compound I. The second part of the cycle involves a transfer of electrons from a reducing substrate to the enzyme, releasing the bound oxygen as water, and regenerating the starting configuration of the active site. The net reaction is:
where AH is the reducing substrate. Catalase is a specific sub-form of peroxidase, in which the reducing substrate is a second molecule of hydrogen peroxide. In this case the reaction is:
Perhaps the prototypical example of a peroxidase is horseradish peroxidase (HRP), which is frequently used as a label in immunochemistry. Coupled to an antibody, HRP can be sensitively detected by its catalytic oxidation of luminol in the presence of hydrogen peroxide. The oxidized luminol is metastable, rearranging to give off a photon of light.
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