The brain is the body's most oxidatively active organ, consuming over 20% of the oxygen we breathe in every day. On the other hand, many diseases associated with aging and the brain, including cancer and neurodegenerative diseases such as Alzheimer's and Parkinson's, have a strong oxidative stress component stemming from cellular oxygen mismanagement. Oxidative stress is the result of unregulated production of reactive oxygen species (ROS), and accumulation of oxidative damage leads to the functional decline of organ systems. We are developing new fluorescent probes for oxygen metabolites, redox status, and enzyme activity to study the molecular mechanisms of oxidative processes in living cells, tissue, and ultimately in vivo.

Fluorogenic Reagents for Probing Hydrogen Peroxide Signaling and Stress. Hydrogen peroxide is a major reactive oxygen species in living systems and a common small-molecule marker for oxidative stress. However, the chemical biology of peroxide is much more complex, as recent studies suggest that this oxygen metabolite, in certain situations, can also serve as a messenger in signal transduction by reacting with redox-active sulfur. This signal/stress dichotomy is reminescent of nitric oxide and offers a brand new view on the potential roles of peroxide in biology. To study the chemistry and chemical biology of hydrogen peroxide in living systems, we are creating new fluorescent probes to map its generation, translocation, and function. A key challenge for probe design is achieving specificity for hydrogen peroxide over a host of very similar reactive oxygen species, including superoxide, nitric oxide, and alkyl peroxides. We have recently discovered a new tactic for selective peroxide detection through the peroxide-mediated deprotection of boronic esters to phenols. Peroxyresofurin-1 (PR1), Peroxyfluor-1 (PF1), and Peroxyxanthone-1 (PX1) represent a first-generation series of peroxide-selective fluorescent probes that respond by increases in red, green, or blue fluorescence, respectively. All three probes are cell-permeable, non-toxic, and can be used to image changes in the levels and distributions of hydrogen peroxide in living cells, including primary hippocampal neurons. Current efforts are directed at using these first-generation Peroxysensors for studies of peroxide oxidation biology, as well as the synthesis of new probes that have attenuated sensitivity and/or the ability to be targeted to specific subcellular locations.

We are also exploring new organic reaction mechanisms for detecting various other reactive oxygen species. Particular species of interest are superoxide, alkyl peroxides, hydroxy radical, hypochlorite, ozone, and singlet oxygen. In addition, optical sensors of redox status are also being pursued.