Irina P. Edimecheva

Quinones as free-radical fragmentation inhibitors in biologically important molecules

Effects of a number of quinones and diphenols of various structures on free-radical fragmentation processes taking place in f -diols, glycerol, 2-aminoethanol, glycero-1-phosphate, ethylene glycol monobutyrate, maltose, and some lipids were investigated. Quinone additions have been found to change the direction of free-radical transformations of the compounds cited above by inhibiting formation of the respective fragmentation products owing to oxidation of radicals of the starting compounds. The results obtained and literature data available allow a suggestion to be made that the system quinone/diphenol is able to not only deactivate or generate such active species as O 2 ” m but also control the realization probability of free-radical processes of peroxidation and fragmentation in biologically important molecules.

Effects of Phenolic Compounds on Reactions Involving Various Organic Radicals

Investigation of effects produced by 26 various phenol and diphenol derivatives, including industrial and natural antioxidants (ionol, bis-phenol 2246, α-tocopherol), on final product yields of radiation-induced free-radical processes involving peroxyl, alkyl, α-hydroxyalkyl and α,β-dihydroxyalkyl radicals has been performed. Ionol and bis-phenol 2246 have been shown to be more effective than α-tocopherol or diphenol derivatives in suppressing hydrocarbon oxidation processes. At the same time, α-tocopherol and its water-soluble analogues, as well as diphenol-based substances, are more effective than phenol derivatives in regulating various homolytic processes involving carbon-centered radicals. This fact can be accounted for by taking into consideration the contribution to formation of the final product set and the respective yields made by semiquinone radicals and compounds with quinoid structure arising in the course of homolytic transformations in systems containing diphenol derivatives.

Reactions of Arylamine and Aminophenol Derivatives, and Riboflavin with Organic Radicals

Based on product yield data on radiolysis of hexane, ethanol and 3 M aqueous ethylene glycol solutions, the ability of a number of arylamine, aminophenol and quinonimine derivatives to affect processes involving peroxyl, alkyl or α-hydroxyalkyl radicals was assessed. It has been shown that the introduction of a hydroxyl group into aromatic amine structure enhances its antioxidant performance and makes it significantly more reactive with respect to carbon-centered organic radicals. Replacement of the hydrogen atom of a hydroxyl group by a methyl group decreases the anti-radical activity of aminophenols drastically. Compounds containing (or capable of forming) a quinonimine moiety interact with alkyl or α-hydroxyalkyl radicals most effectively, suppressing recombination and fragmentation reactions of the latter. In the sequence: aromatic amines--aminophenols--quinonimines, a trend towards enhancement of the ability of the compounds studied to react with carbon-centered radicals was noted. Also, this study presents for the first time evidence of riboflavin reactivity with respect to organic radicals.

22-NBD-cholesterol as a novel fluorescent substrate for cholesterol-converting oxidoreductases.

Docking simulations and experimental data indicate that 22-(N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-23,24-bisnor-5-cholen-3β-ol (22-NBD-cholesterol), a common fluorescent sterol analog, binds into active sites of bovine cytochrome P450scc and microbial cholesterol dehydrogenases (CHDHs) and then undergoes regiospecific oxidations by these enzymes. The P450scc-dependent system was established to realize N-dealkylation activity toward 22-NBD-cholesterol, resulting in 7-nitrobenz[c][1,2,5]oxadiazole-4-amine (NBD-NH(2)) formation as a dominant fluorescent product. Basing on LC-MS data of the probes derivatized with hydroxylamine or cholesterol oxidase, both pregnenolone and 20-formyl-pregn-5-en-3β-ol were deduced to be steroidal co-products of NBD-NH(2), indicating intricate character of the reaction. Products of CHDH-mediated conversions of 22-NBD-cholesterol were defined as 3-oxo-4-en and 3-oxo-5-en derivatives of the steroid. Moreover, the 3-oxo-4-en derivative was also found to be formed after 22-NBD-cholesterol incubation with pathogenic bacterium Pseudomonas aeruginosa, indicating a possible application of the reaction for a selective and sensitive detection of some microbes. The 3-keto-4-en derivative of 22-NBD-cholesterol may be also suitable as a new fluorescent probe for steroid hormone-binding enzymes or receptors.

Effects of various vitamins and coenzymes Q on reactions involving α-hydroxyl-containing radicals

Effects of vitamins B, C, E, K and P, as well as coenzymes Q, on formation of final products of radiation-induced free-radical transformations of ethanol, ethylene glycol, α-methylglycoside and glucose in aqueous solutions were studied. Based on the obtained results, it can be concluded that there are substances among vitamins and coenzymes that effectively interact with α-hydroxyl-containing radicals. In the presence of these substances, recombination reactions of α-hydroxyalkyl radicals and fragmentation of α-hydroxy-β-substituted organic radicals are suppressed. It has been established that the observed effects are due to the ability of the vitamins and coenzymes under study to either oxidize α-hydroxyl-containing radicals yielding the respective carbonyl compounds or reduce them into the initial molecules.

Evaluation of the fluorescent probes Nile Red and 25‐NBD‐cholesterol as substrates for steroid‐converting oxidoreductases using pure enzymes and microorganisms

The fluorescent probes Nile Red (nonsteroidal dye) and 25‐{N‐[(7‐nitrobenz‐2‐oxa‐1,3‐diazol‐4‐yl)‐methyl]amino}‐27‐norcholesterol (25‐NBD‐cholesterol) (a cholesterol analog) were evaluated as novel substrates for steroid‐converting oxidoreductases. Docking simulations with autodock showed that Nile Red fits well into the substrate‐binding site of cytochrome P450 17α‐hydroxylase/17,20‐lyase (CYP17A1) (binding energy value of −8.3 kcal·mol−1). Recombinant Saccharomyces cerevisiae and Yarrowia lipolytica, both expressing CYP17A1, were found to catalyze the conversion of Nile Red into two N‐dealkylated derivatives. The conversion by the yeasts was shown to increase in the cases of coexpression of electron‐donating partners of CYP17A1. The highest specific activity value (1.30 ± 0.02 min−1) was achieved for the strain Y. lipolytica DC5, expressing CYP17A1 and the yeasts NADPH‐cytochrome P450 reductase. The dye was also metabolized by pure CYP17A1 into the N‐dealkylated derivatives, and gave a type I difference spectrum when titrated into low‐spin CYP17A1. Analogously, docking simulations demonstrated that 25‐NBD‐cholesterol binds into the active site of the microbial cholesterol oxidase (CHOX) from Brevibacterium sterolicum (binding energy value of −5.6 kcal·mol−1). The steroid was found to be converted into its 4‐en‐3‐one derivative by CHOX (Km and kcat values were estimated to be 58.1 ± 5.9 μm and 0.66 ± 0.14 s−1, respectively). The 4‐en‐3‐one derivative was also detected as the product of 25‐NBD‐cholesterol oxidation with both pure microbial cholesterol dehydrogenase (CHDH) and a pathogenic bacterium, Pseudomonas aeruginosa, possessing CHOXs and CHDHs. These results provide novel opportunities for investigation of the structure–function relationships of the aforementioned oxidoreductases, which catalyze essential steps of steroid bioconversion in mammals (CYP17A1) and bacteria (CHOX and CHDH), with fluorescence‐based techniques.

Effect of phenolic compounds and quinones on radiation-induced oxidation processes of hexane and ethanol

The radiation-chemical yields were determined for the products of γ-radiation induced oxidation of hexane and ethanol in the presence of natural and synthetic phenolic compounds and quinones. It was found that the commercial antioxidants ionol and bisphenol 2246 more effectively inhibit the hexane oxidation process while α-tocopherol and quinones are superior to other test compounds in the suppression of ethanol oxidation. The results indicate that the antioxidant properties of phenolic compounds and quinones depend not only on their structure but also on the nature of the oxidation substrate. The cause of this phenomenon is discussed.

A New Mechanism for Photo- and Radiation-Induced Decomposition of Sphingolipids

Data have been obtained showing regularities in product formation following radiolysis of serinol, lysosphingomyelin and photolysis of N-(2-hydroxypropyl)hexanamide, sphingomyelin, which point to the possibility of photo- and radiation-induced destruction of the named substrates via a C–C bond rupture. The key stage of this process is the formation and decomposition of N-centered radicals generated from the starting compounds.

Uptake and metabolism of fluorescent steroids by mycobacterial cells.

HighlightsMycobacteria can convert BODIPY‐cholesterol and similar NBD‐labeled steroids.4‐En‐3‐one derivatives are major fluorescent products of the bioconversion.NBD fluorophore of the compounds undergoes degradation in the mycobacteria.The steroids cause staining of envelope lipids and intracellular lipid droplets.3‐NBD‐cholestane don’t converted by M. tuberculosis H37Rv and M. smegmatis mc2 155. Abstract Fluorescent steroids BODIPY‐cholesterol (BPCh) and 7‐nitrobenzoxadiazole‐4‐amino‐(NBD)‐labeled 22‐NBD‐chelesterol (22NC) as well as synthesized 20‐(NBD)‐pregn‐5‐en‐3&bgr;‐ol (20NP) were found to undergo bioconversions by Mycobacterium tuberculosis H37Rv and M. smegmatis mc2 155. The major fluorescent products were determined to be 4‐en‐3‐one derivatives of the compounds. Degradation of NBD fluorophore was also detected in the cases of 22NC and 20NP, but neither NBD degradation nor steroidal part modification were observed for the synthesized 3‐(NBD)‐cholestane. Mycobacterial 3&bgr;‐hydroxysteroid dehydrogenases were concluded to be responsible for the formation of the 4‐en‐3‐one derivatives. All the compounds tested were found to cause staining both membrane lipids and cytosolic lipid droplets when incubated with mycobacteria in different manner, demonstrating ability of the steroids to reside in the compartments. The findings reveal a potential of the compounds for monitoring of steroid interactions with mycobacteria and provide information for design of new probes for this purpose.

Free-radical Destruction of Sphingolipids Resulting in 2-hexadecenal Formation

The action of hypochlorous acid (HOCl) and γ-radiation on aqueous lysosphingolipid dispersions was found to produce 2-hexadecenal (Hex). This process includes the stages of formation of nitrogen-centered radicals from the starting molecules and the subsequent fragmentation of these radicals via the rupture of C–C and O–H bonds. These findings prove the existence of a nonenzymatic pathway of sphingolipid destruction leading to the formation of Hex, which possesses a wide spectrum of biological activity. Analysis of the effect of HOCl on transplantable rat glioma C6 cells and human embryonic kidney 293 cells points to the formation of Hex. This suggests that the described mechanism of free-radical destruction of sphingolipids may be replicated on cell culture under the stress of active chlorine forms.