Study of antioxidant properties of metal aspartates.

The effects of Zn, Mg, Cr, Cu, and Mn aspartates, their commercial formulation Inzolen, and the individual commercial medicine Unizinc, on oxygen radical production by enzymes [xanthine oxidase, horseradish peroxidase, and reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase] and phagocytic cells (human blood leukocytes) have been studied. The formation of oxygen radicals was measured by luminol- and lucigenin-amplified chemiluminescence and by the reduction of cytochrome c. All these compounds (excluding Cr aspartate) turn out to be inhibitors of oxygen radical formation in the systems studied (excluding horseradish peroxidase). Their inhibitory activities were a consequence of both the scavenging of free radicals and the inhibition of xanthine oxidase and NADPH oxidase activities. As expected, the most active free-radical scavengers were transition metal Cu and Mn aspartates, which mimicked the activities of copper-zinc and manganese dismutases. However, surprisingly non-transition metal Zn and Mg aspartates were also able to scavenge oxygen radicals. It was suggested that the scavenging activities of Zn and Mg aspartates may be explained by affecting the rate of spontaneous dismutation of the superoxide ion. In addition, it was found that Zn aspartate is an efficient inhibitor of the formation of the most reactive hydroxyl radicals. These antioxidant properties of Zn aspartate make it important in medicine for the prevention and treatment of free radical pathologies.

Free radical generation during interaction of chrysotile asbestos with natural compounds.

Free radical generation was found after the addition of some natural biological substances (adrenaline, ascorbate, ubiquinone Q9, etc.) to a chrysotile asbestos suspension. This was detected by the chemiluminescent method with lucigenin as an indicator. The detailed study of the chemiluminescent reaction in the adrenaline-chrysotile system indicated that the reaction required hydroxyl ions, which arose in the chrysotile suspension, and was accompanied by a superoxide radical formation. At the same time, the radical production was very low in suspensions of amphybole asbestos, talc, and quartz, which could not alkalinize the water medium. On the basis of these results, it may be concluded that chrysotile has a unique ability to generate free radicals upon interaction with some biological molecules in a water medium. This fact may explain the great carcinogenicity of chrysotile. The injection of cigarette smoke solution into chrysotile (but not into amphibole asbestos or talc) suspension induced intensive chemiluminescence. This suggests that smoke aggravates the effect of chrysotile on human health by increasing free radical generation on the surface of the fibers.

[Chemiluminescent determination of superoxide dismutase activity]. / Khemiliuminestsentnoe opredelenie aktivnosti superoksiddismutazy.

A specific chemiluminescent method for the assessment of superoxide dismutase (SOD) activity in human blood has been developed. Intensive lucinogen-dependent chemiluminescence was observed during adrenalin oxidation in alkaline medium, and the level of this chemiluminescence essentially decreased in the presence of SOD. 50% inhibition of chemiluminescence was observed in the presence of 25 ng/ml of the enzyme, this indicating a high sensitivity of the method as against a similar spectrophotometric technique. SOD activities in donor blood samples were measured. The described method is offered for clinical application.

Release of active oxygen radicals by leukocytes of Fanconi anemia patients.

The release of oxygen radicals by blood and bone marrow leukocytes of patients with Fanconi anemia (FA) has been studied. It was found that the nonstimulated FA leukocytes and those stimulated by concanavalin A, SiO2, latex, and opsonized zymosan produced enhanced levels of luminol- and lucigenin-dependent chemiluminescence (CL) in comparison with normal leukocytes. At the same time, the ratio of the intensity of lucigenin-dependent CL to that of luminol-dependent CL was significantly smaller for FA leukocytes than for normal leukocytes. From these findings and from the effects of antioxidative enzymes and free radical scavengers on CL, it was concluded that FA leukocytes release enhanced amounts of oxygen radicals and that these free radicals contain enhanced amounts of hydroxyl or hydroxyl-like radicals more active than superoxide ion. It was proposed that elevated reactivity of the oxygen radicals released by FA leukocytes may be a major factor in the development of Fanconi anemia; this proposal is supported by the first positive results of treatment of FA patients with rutin (a nontoxic natural free radical scavenger and chelator).

Oxygen radical-mediated mutagenic effect of asbestos on human lymphocytes: suppression by oxygen radical scavengers.

The mutagenic effect of chrysotile asbestos fibers and zeolite and latex particles on human lymphocytes in whole blood has been studied. It was concluded that their mutagenic activities were mediated by oxygen radicals because they were inhibited by antioxidant enzymes (SOD and catalase) and oxygen radical scavengers (rutin, ascorbic acid, and bemitil). It was proposed that oxygen radicals were released by phagocytes activated upon exposure to mineral dusts and fibers. The study of lucigenin- and luminol-amplified chemiluminescence of peritoneal macrophages stimulated by chrysotile fibers and zeolite and latex particles has shown that their mutagenic action is probably mediated by different oxygen species, namely, by the iron-oxygen complexes (perferryl ions) plus hydrogen peroxide, hydrogen peroxide, and superoxide ion, respectively. From the oxygen radical scavengers studied, rutin was the most effective inhibitor of the mutagenic effect of mineral fibers and dusts.

Are quinones producers or scavengers of superoxide ion in cells?

The effects of quinones (benzoquinone, menadione, and doxorubicin) on the superoxide production in cell free systems (xanthine oxidase and rat liver microsomes) and of polycationic electrolyte- and latex-stimulated rat peritoneal macrophages have been studied. Contradictory results were obtained in cell free systems when two traditional assays for detection of superoxide ion, the cytochrome c reduction and the lucigenin-dependent chemiluminescence (CL), were used: all quinones inhibited the lucigenin-dependent CL at sufficiently large concentrations, but they did not inhibit at all the reduction of cytochrome c. It was proposed that the cytochrome c assay gave erroneous results due to the reversibility of the interaction of semiquinones with dioxygen. The effect of quinones on the superoxide production by peritoneal macrophages was biphasic: all quinones stimulated the O2-. formation at low concentrations and inhibited it at elevated concentrations. It was concluded that among the quinones studied, only menadione was capable of stimulating the superoxide production via a one-electron transfer mechanism in cell free systems, while the stimulatory effect of small concentrations of quinones on the O2-. production in macrophages was possibly due to their action on the activation of NADPH oxidase.

[Chemoluminescence analysis of the functional state of blood phagocytes in silicosis and chronic dust-induced bronchitis]. / Khemoliuminestsentnyi analiz funktsional'nogo sostoianiia fagotsitov krovi pri silikoze i khronicheskom pylevom bronkhite.

Blood leukocyte function in patients with silicosis and chronic dust-induced bronchitis was examined by cellular chemiluminescence method for registration of free oxygen radical production. Significant differences in oxidation metabolism of blood neutrophils in the patients with these two nosologic entities were revealed. In silicosis blood phagocyte activation with silica particles in vitro activated production of active oxygen forms and chemiluminescent cells in all cases by 3 to 30 times. In chronic dust-induced bronchitis the production of active oxygen forms under the same conditions was either unchanged or enhanced but negligibly, the activation coefficient not surpassing 3. These data point to a new approach to the differential diagnosis between the above-mentioned conditions.

[The mutagenic action of the dust of natural zeolites and chrysotile asbestos]. / Issledovanie mutagennogo deistviia pyli prirodnykh tseolitov i khrizotil-asbesta.

The cell chemiluminescence method was used to demonstrate the ability of asbest and zeolite dusts from 8 deposits of the USSR to induce generation of free oxygen radicals in the phagocytosing cells suspension. It has been found that asbest and zeolite (0.01 and 0.05 mg/ml) increase levels of cells with chromosome aberrations in human cell cultures. The cytogenetic effect of asbest was inhibited by superoxide dismutase (50 mg/ml). The damaging effect of zeolite was decreased by the pharmacological drug bemithyl (0.007-0.07 mM) and completely eliminated by catalase (20 mg/ml). The results obtained indicate that mutagenic effect of dust particles of asbest and zeolite is mediated by oxygen radicals.

[The role of free oxygen radicals in the pathogenesis of Fanconi's anemia]. / Rol' svobodnykh radikalov kisloroda v patogeneze anemii Fankoni.

The authors describe the results of a study into antioxidants (superoxide dismutase and catalase) and production of the active forms of oxygen (superoxide anion radical, hydrogen peroxide, hydroxyl radical, and singlet oxygen) by blood and bone marrow leukocytes from patients with Fanconi's anemia. The generation of the active forms of oxygen was induced by silica particles while the formation of radical products was recorded by means of luminol-dependent chemiluminescence. Unbalanced excessive production of oxygen radicals such as hydroxyl radicals combined with a decrease in the activity of antioxidant defence enzymes was discovered. It is believed that it is this circumstance that causes the derangement of the membranous structures and multiple breakdowns in the hereditary apparatus. It is suggested that antiradical preparations may be applied to correcting therapy of patients with Fanconi's anemia.

[Mechanism of the cytotoxic action of the natural zeolite clinoptilolite]. / Mekhanizm tsitotoksicheskogo deistviia prirodnogo tseolita klinoptilolita.

Interaction between the natural ceolite clinoptilolite and cell suspensions has been investigated using rat peritoneal macrophages and erythrocytes. The ceolite under study has been demonstrated to exhibit a high hemolytic activity and cytotoxicity. The viability of macrophages was evaluated from the incorporation of trypane blue. The ability of macrophages to phagocytosis was measured by chemiluminescence with luminol. The modification of clinoptilolite surface by ammonia ions led to a decrease in its cytotoxic properties. Ethanol, mannit and sodium azide did not affect whereas catalase appreciably reduced the ability of CPT to damage the membranes of macrophages and red cells. The role of hydrogen peroxide in the mechanism of cell membrane damage is discussed.