Production Of Hydroxyl Free Radicals Research Articles

Mutation spectrum of copper-induced DNA damage.

The ability of metal ions to damage DNA and cause mutagenesis has been analyzed with reversion and forward mutation assays using single-stranded DNA templates. We previously reported that incubation of phi X174 am3 DNA with Fe2+ in vitro results in mutagenesis when the treated DNA is transfected into Escherichia coli spheroplasts (Loeb, L. A., James, E. A., Waltersdorph, A. M., and Klebanoff, S. J. (1988) Proc. Natl. Acad. Sci. U.S.A. 85, 3918-3922, 1988). We now extend these studies to other metal ions. Of the metal ions tested, copper ions were the most mutagenic; the frequency of mutants produced was equal to or greater than that produced by Fe2+. Mutagenesis by Cu+ was diminished by catalase, mannitol, and superoxide dismutase suggesting the involvement of H2O2, hydroxyl ions, and superoxide, respectively. However, the findings that Cu+ and Cu2+ are nearly equally mutagenic and that the mutagenic activities are not completely inhibited by oxygen free radical scavengers make it unlikely that the mechanism for mutagenesis is simply the production of hydroxyl free radicals. The spectra of mutations produced by either copper ion using the lacZ gene as a target are very similar and differ from those reported with other agents. The predominant mutagenic sequence changes are single-base substitutions, the most frequent being replacement of a template C by a T. This transition presumably results from mispairing of an altered C with deoxyadenosine. Copper-induced mutations are not randomly distributed. Instead, they are found predominantly in clusters suggesting direct interaction of copper ions with specific nucleotide sequences in DNA. Evidence is considered that the high frequency of C----T transitions may be a common manifestation of DNA damage by oxygen radicals.

Porphyrin photosensitization and phototherapy.

Porphyrin photosensitization and phototherapy.

Hydrogen peroxide and hydroxyl free radical production by hematoporphyrin derivative, ascorbate and light

Photofrin II, a commercial preparation of a hematoporphyrin derivative containing a high proportion of the active di-hematoporphyrin ether, was found to convert oxygen to hydrogen peroxide and hydroxyl free radicals in the presence of ascorbate and light. In addition, a light-dependent increase in the ascorbate free radical concentration was observed in hematoporphyrin derivative/ascorbate solutions with subsequent oxidation of ascorbate.

27] High-pressure liquid chromatography-electrochemical detection of oxygen free radicals

The presence of superoxide, hydrogen peroxide, and a catalytic amount of iron complexed to an appropriate liquid, such as di- or triphosphate nucleotides, will lead to the production of hydroxyl free radicals (OH·). Hydroxyl free radicals are extremely reactive and thus would be present at any specific time in at most very low amounts in biological systems even under conditions of severe oxidative stress. Detection of hydroxyl free radicals per se by electron paramagnetic resonance (EPR) in biological systems is virtually impossible; however, the use of spin-trapping techniques offers possibilities. The results presented illustrate that the hydroxyl-free-radical adduct of 5,5-dimethyl-1-pyrroline N-oxide (DMPO) (DMPO-OH·) and its one-electron reduction product (DMPO-OH 2 ) can be separated out and detected by high-pressure liquid chromatography (HPLC)-electrochemical detection. It is clear, because biological systems tend to reduce DMPO-OH· to the diamagnetic DMPO-OH 2 and because HPLC-electrochemical detection offers extreme sensitivity as well as selective separation that this method has excellent potential of being very useful as a tool for studying oxygen free radicals in biological systems.

Hydroxyl free radical production in iron-cysteine solutions and protection by zinc

Hydroxyl radicals (OH −) can be formed on incubation of an oxygenated solution of ferrous sulphate and cysteine. This has been demonstrated by esr using the spin trap DMPO (5,5-dimethyl-1-pyrroline-1-oxide), catalase, and the radical scavengers ethanol and propan-2-ol. Hydroxyl radicals are not formed when excess zinc sulphate is present. These results provide support for the pro-oxidant action of iron and cysteine and a possible protective role for zinc.