Tapinarof (3,5-dihydroxy-4-isopropylstilbene) is a therapeutic agent used in the treatment of psoriasis (VTAMA®). In this study, we examined the redox behaviour, (photo)stability, (photo)toxicity and (bio)transformation of tapinarof in the context of a structure-activity relationship study. Selected derivatives of the structurally related tapinarof were investigated, namely resveratrol, pterostilbene, pinosylvin and its methyl ether. Tapinarof undergoes electrochemical oxidation in a neutral aqueous medium at a potential of around +0.5 V (vs. Ag|AgCl|3M KCl). The anodic reaction of this substance is a proton-dependent irreversible and adsorption-driven process. The pKa value of tapinarof corresponds to 9.19 or 9.93, based on empirical and QM calculation approach, respectively. The oxidation potentials of tapinarof and its analogues correlate well with their HOMO (highest occupied molecular orbital) energy level. The ability to scavenge the DPPH radical decreased in the order trolox ≥ resveratrol > pterostilbene > tapinarof > pinosylvin ≫ pinosylvin methyl ether. It was also confirmed that tapinarof, being a moderate electron donor, is able to scavenge the ABTS radical and inhibit lipid peroxidation. The 4'-OH group plays a pivotal role in antioxidant action of stilbenols. During the stability studies, it was shown that tapinarof is subject to spontaneous degradation under aqueous conditions, and its degradation is accelerated at elevated temperatures and after exposure to UVA (315-399 nm) radiation. In aqueous media at pH 7.4, we observed an ∼50 % degradation of tapinarof after 48 h at laboratory temperature. The main UVA photodegradation processes include dihydroxylation and hydration. In conclusion, the phototoxic effect of tapinarof on a human keratinocytes cell line (HaCaT) was evaluated. Tapinarof exhibited a clear phototoxic effect, similar to phototoxic standard chlorpromazine. The IC50 values of the cytotoxicity and phototoxic effects of tapinarof correspond to 27.6 and 3.7 μM, respectively. The main HaCaT biotransformation products of tapinarof are sulfates and glucuronides.
- MeSH
- antioxidancia farmakologie chemie MeSH
- buněčné linie keratinocytů HaCaT MeSH
- fototoxická dermatitida MeSH
- keratinocyty * účinky léků metabolismus účinky záření MeSH
- kůže účinky léků metabolismus účinky záření patologie MeSH
- lidé MeSH
- oxidace-redukce * MeSH
- resveratrol farmakologie analogy a deriváty chemie MeSH
- stilbeny * farmakologie chemie MeSH
- ultrafialové záření MeSH
- vztahy mezi strukturou a aktivitou MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Some biologically active substances are unstable and poorly soluble in aqueous media, at the same time exhibiting low bioavailability. The incorporation of these biologically active compounds into the structure of a lipid-based lyotropic liquid crystalline phase or nanoparticles can increase or improve their stability and transport properties, subsequent bioavailability, and applicability in general. The aim of this short overview is (1) to clarify the principle of self-assembly of lipidic amphiphilic molecules in an aqueous environment and (2) to present lipidic bicontinuous cubic and hexagonal phases and their current biosensing (with a focus on electrochemical protocols) and biomedical applications.
- MeSH
- kapalné krystaly * chemie MeSH
- lipidy chemie MeSH
- nanočástice * chemie MeSH
- technologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
Lipid nitroalkenes - nitro-fatty acids (NO2-FAs) are formed in vivo via the interaction of reactive nitrogen species with unsaturated fatty acids. The resulting electrophilic NO2-FAs play an important role in redox homeostasis and cellular stress response. This study investigated the physicochemical properties and reactivity of two NO2-FAs: 9/10-nitrooleic acid (1) and its newly prepared 1-monoacyl ester, (E)-2,3-hydroxypropyl 9/10-nitrooctadec-9-enoate (2), both synthesized by a direct radical nitration approach. Compounds 1 and 2 were investigated in an aqueous medium and after incorporation into lipid nanoparticles prepared from 1-monoolein, cubosomes 1@CUB and 2@CUB. Using an electrochemical analysis and LC-MS, free 1 and 2 were found to be unstable under acidic conditions, and their degradation occurred in an aqueous environment within a few minutes or hours. This degradation was associated with the production of the NO radical, as confirmed by fluorescence assay. In contrast, preparations 1@CUB and 2@CUB exhibited a significant increase in the stability of the loaded 1 and 2 up to several days to weeks. In addition to experimental data, density functional theory-based calculation results on the electronic structure and structural variability (open and closed configuration) of 1 and 2 were obtained. Finally, experiments with a human HaCaT keratinocyte cell line demonstrated the ability of 1@CUB and 2@CUB to penetrate through the cytoplasmic membrane and modulate cellular pathways, which was exemplified by the Keap1 protein level monitoring. Free 1 and 2 and the cubosomes prepared from them showed cytotoxic effect on HaCaT cells with IC50 values ranging from 1 to 8 μM after 24 h. The further development of cubosomal preparations with embedded electrophilic NO2-FAs may not only contribute to the field of fundamental research, but also to their application using an optimized lipid delivery vehicle.
- MeSH
- dusíkaté sloučeniny MeSH
- faktor 2 související s NF-E2 MeSH
- KEAP-1 MeSH
- lidé MeSH
- mastné kyseliny * MeSH
- oxid dusnatý * metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Nitro-fatty acids modulate inflammatory and metabolic stress responses, thus displaying potential as new drug candidates. Herein, we evaluate the redox behavior of nitro-oleic acid (NO2-OA) and its ability to bind to the fatty acid transporter human serum albumin (HSA). The nitro group of NO2-OA underwent electrochemical reduction at -0.75 V at pH 7.4 in an aqueous milieu. Based on observations of the R-NO2 reduction process, the stability and reactivity of NO2-OA was measured in comparison to oleic acid (OA) as the negative control. These electrochemically-based results were reinforced by computational quantum mechanical modeling. DFT calculations indicated that both the C9-NO2 and C10-NO2 positional isomers of NO2-OA occurred in two conformers with different internal angles (69° and 110°) between the methyl- and carboxylate termini. Both NO2-OA positional isomers have LUMO energies of around -0.7 eV, affirming the electrophilic properties of fatty acid nitroalkenes. In addition, the binding of NO2-OA and OA with HSA revealed a molar ratio of ~7:1 [NO2-OA]:[HSA]. These binding experiments were performed using both an electrocatalytic approach and electron paramagnetic resonance (EPR) spectroscopy using 16-doxyl stearic acid. Using a Fe(DTCS)2 spin-trap, EPR studies also showed that the release of the nitro moiety of NO2-OA resulted in the formation of nitric oxide radical. Finally, the interaction of NO2-OA with HSA was monitored via Tyr and Trp residue electro-oxidation. The results indicate that not only non-covalent binding but also NO2-OA-HSA adduction mechanisms should be taken into consideration. This study of the redox properties of NO2-OA is applicable to the characterization of other electrophilic mediators of biological and pharmacological relevance.
- MeSH
- dusíkaté sloučeniny chemie metabolismus MeSH
- kyselina dusičná chemie metabolismus MeSH
- lidé MeSH
- lidský sérový albumin chemie metabolismus MeSH
- mastné kyseliny chemie metabolismus MeSH
- oxid dusnatý chemie metabolismus MeSH
- oxidace-redukce * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
The lipidic liquid-crystalline cubic phase (LCP) is a membrane-mimetic material useful for the stabilization and structural analysis of membrane proteins. Here, we focused on the incorporation of the membrane ATP-hydrolysing sodium/potassium transporter Na+/K+-ATPase (NKA) into a monoolein-derived LCP. Small-angle X-ray scattering was employed for the determination of the LCP structure, which was of Pn3m symmetry for all the formulations studied. The fully characterized NKA-LCP material was immobilized onto a glassy carbon electrode, forming a highly stable enzyme electrode and a novel sensing platform. A typical NKA voltammetric signature was monitored via the anodic reaction of tyrosine and tryptophan residues. The in situ enzyme activity evaluation was based on the ability of NKA to transform ATP to ADP and free phosphate, the latter reacting with ammonium molybdate to form the ammonium phosphomolybdate complex under acidic conditions. The square-wave voltammetric detection of phosphomolybdate was performed and complemented with spectrophotometric measurement at 710nm. The anodic voltammetric response, corresponding to the catalytic ATP-hydrolysing function of NKA incorporated into the LCP, was monitored at around + 0.2V vs. Ag/AgCl in the presence or absence of ouabain, a specific NKA inhibitor. NKA incorporated into the LCP retained its ATP-hydrolysing activity for 7 days, while the solubilized protein became practically inactive. The novelty of this work is the first incorporation of NKA into a lipidic cubic phase with consequent enzyme functionality and stability evaluation using voltammetric detection. The application of LCPs could also be important in the further development of new membrane protein electrochemical sensors and enzyme electrodes.
- MeSH
- adenosintrifosfát metabolismus MeSH
- biosenzitivní techniky metody MeSH
- enzymatické testy metody MeSH
- enzymy imobilizované chemie metabolismus MeSH
- glyceridy chemie MeSH
- hydrolýza MeSH
- kapalné krystaly chemie MeSH
- molekulární modely MeSH
- prasata MeSH
- sodíko-draslíková ATPasa chemie metabolismus MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Flavonolignans, silybin and its derivatives (2,3-dehydrosilybin, 7-O-methylsilybin, 20-O-methylsilybin) and isosilybin were studied using ex situ (adsorptive transfer, AdT) cyclic and square wave voltammetry (SWV). The two oxidation steps were described for flavonolignans at potentials E(p1) +0.5 V and E(p2) +0.85 V depending on experimental conditions. An additional oxidation peak at E(p3) +0.35 V was observed only for 2,3-dehydrosilybin. The anodic currents of flavonolignans are related to their electron transfer processes (oxidation of hydroxyl groups), which was supported by density functional theory (DFT) and B3P86 theory level. Our electrochemical results confirmed that 2,3-dehydrosilybin is a relatively strong antioxidant, which is strictly associated with oxidation at E(p3). The oxidation processes and antioxidant parameters of flavonolignans can be affected by transition metal complexation via hydroxyl groups. We found that silybin and 2,3-dehydrosilybin are able to chelate transition metals, especially Cu(2+). The formation of silybin/Cu complexes was studied by AdT SWV and the observation was also confirmed using fluorescence spectroscopy. The electrochemical investigation of DNA interactions and damage caused in the presence of silybin/Cu complex and hydrogen peroxide is described. We present evidence that flavonolignans are involved not only in antioxidant abilities but also in the prooxidation effects under in vitro conditions.
