ANALYSIS OF LIGHT EMITTED DURING OXIDATIVE STRESS

University Of South Alabama, Mobile AL

Investigators

Abstract

DESCRIPTION: (Adapted from the applicant's abstract) Oxidative stress is a common denominator in many pathological scenarios. During oxidative stress, reactive oxygen species (ROS) start and/or propagate chain reactions which result in the oxidation of most biological molecules leading to cell damage and death. Thus, it is important to have non-invasive techniques that will estimate the magnitude of an oxidative insult in real time. Yet, none of the available methods can do this without interfering with the process. We propose that the ultraweak light emitted during the oxidation of proteins and lipids, which is proportional to the steady state concentration of ROS, may be a tool to monitor the kinetics of oxidative stress without interfering with the course of the reaction or having to wait for the accumulation of oxidation products. Light emission results from the decay of electronically excited species produced during protein and lipid oxidation. In order to amplify these signals, it is important to know which are the primary emission wavelengths. Preliminary studies indicate that most of the emitted light is in the range between 500 and 700 nm, but the available photon counters are not sensitive enough to be used for more detailed spectral analysis. The sensitivity of this technique would be increased substantially using diode arrays (CCDs) with higher quantum efficiency, but the lack of information regarding peak emission wavelengths limits their application to single-photon counting. The aim of this study is to re-design our photon counter to characterize the spectra of light emitted during the oxidation of model systems including free amino acids, proteins, liposomes, lipid/protein mixtures and, finally, intact cells. A phototube with large surface area and broad spectral range will be used as a detector. The proposed changes will increase the current sensitivity by about one order of magnitude. The detailed spectral analysis obtained in this study will also provide important information for future development of highly sensitive CCD-based instrumentation to monitor oxidative stress in a variety of scenarios. Most of the studies proposed in this application will be carried out by undergraduate students under the direction of the PI.