A New Technology For The Detection Of Biological Free Radicals

In recent years, Oxidative damage of biological macromolecules, especially DNA and protein, by reactive oxygen species (ROS) have attracted much attention due to their implication in mutagenesis, carcinogenesis, and aging. Thus, it is important to develop biosensors for monitoring the ROS-induced biomacromolecular damage. The general methods for biomacromolecular oxidative damage are sensitive but time-consuming, labor-intensive and costly. Herein, we report a simple, rapid, and label-free colorimetric method for assaying the cleavage of ssDNA by oxidative damage or ssDNA specific nuclease. Moreover, we synthesis of high sensitivity spin traps for the detection of biomacromolecules damged by ROS. The main work of this thesis is summarized as follows:1. A direct visualization method for the cleavage of single-stranded DNA detection was developed by using unmodified gold nanoparticle (AuNP) as indicator. Based on the fact that ssDNA absorbs on AuNPs with a rate that depends on sequence length, we explored a straightforward strategy of controlling the adsorption time of ssDNA on AuNPs for identifying the DNA length change upon the cleavage of ssDNA. Incubation of ssDNA with AuNPs in a controlled time would cause the differential adsorption amount of short ssDNA and long ssDNA on AuNPs and consequently lead to length-dependent stability of ssDNA/AuNPs complex against salt-induced aggregation, thereby causing the corresponding color-change of gold nanoparticles solution in the presence of salt. The method developed here presents a promising technique to monitor ssDNA cleavage process by ssDNA-specific nucleases or free radicals. It also shows potential applications in natural product or drug screening based on inhibition of oxidative damage of DNA.2. In this study, two kinds of cyclic-nitrone spin traps have been successfully synthesized and characterized by ESI/MS,1H-NMR, IR and so on. These two spin traps contain hydroxyl group or carboxyl group, respectively, which improves the water solubility of the spin traps and was beneficial for the application in living organism; Additionally, these spin traps will be further modified by functional molecules(such as fluorescent and biotin) or electroactive molecules(such as ferrocene and methylene blue), which will be used as the novel probes for the detection of biomaromolecule-centered radicals.