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  • Author or Editor: Zsuzsa Király-Véghely x
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A method, including solid phase extraction sample preparation, overpressured layer chromatographic separation, and subsequent densitometric evaluation, was developed for measurement of total resveratrol (cis- and trans-isomers) content of wine. The amount of resveratrol was determined in wine samples from different winemaking regions of Hungary. The total resveratrol was high in Hungarian red wines (3.6–11 mg L−1), and much lower in white ones (0.04–1.5 mg L−1).

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A simple method is described for separation and determination of the isomers of trans -resveratrol and related compounds from different types of red wine by overpressured layer chromatography (OPLC). Comparison of OPLC with TLC clearly showed the advantages of the forced-flow technique (higher theoretical plate number, good resolution, etc.) over conventional planar layer liquid chromatography. It was established that the glycosides of resveratrol isomers were always present in higher concentrations than free stilbene isomers in red wine samples. This was especially true for the Pinot Noir wine. Exploitation of the advantages of OPLC provides further possibilities of analysis and isolation of stilbene isomers from grapes and wine.

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On the basis of our earlier observations, preliminary experiments were conducted with the fire blight pathogen Erwinia amylovora with the BioArena system using the potential antibacterial substances 1′-methylascorbigen (MeAG) as formaldehyde (HCHO) precursor and trans-resveratrol (RES) as HCHO mobilizer, capturer, and scavenger. Results showed characteristic, strong, dose-dependent antibacterial effects of both MeAG and RES present as chromatographic spots on the adsorbent layer. It is supposed that the weak antibacterial effect of AG may have originated from MeAG formed in situ on the layer by partial enzymatic methylation of AG. Addition of HCHO capture compounds (l-arginine, glutathione, glucosamine) to the culture medium before inoculation partially or totally reduced the antibacterial effect of both molecules, that is, this simplest aliphatic aldehyde, the endogenous HCHO molecule, participates in the antibacterial activity of these compounds. It follows from these results that there is a possibility of interaction between endogenous H2O2 and HCHO in the chromatographic spots and it is supposed that these reactive molecules and potential derivatives, e.g. singlet oxygen (1O2) and ozone (O3), may be the causes of the destructive effect of the fire blight pathogen on fruit trees.

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JPC - Journal of Planar Chromatography - Modern TLC
Authors: Ernő Tyihák, Ágnes Móricz, Péter Ott, György Kátay, and Zsuzsa Király-Véghely

New results with BioArena as a complex separation and detection (evaluation) system support earlier observations that formaldehyde (HCHO) and its reaction products play a special and crucial role in the effects of antibiotic in general. It has been established that antibiotic-like compounds (e.g. trans -resveratrol, Cu(II) ions) have a duplicate inhibiting effect on pathogen cells as a result of the action of HCHO. HCHO as a key molecule of the formaldehydome participates in series of interactions which can be screened by means of different spectroscopic and chromatographic techniques; accumulation of HCHO and its reaction products in TLC spots is limited, however, so indirect detection is advantageous. In BioArena the planar stationary phase bed after TLC/HPTLC and, mainly, OPLC separation can be used for manifestation of deprivation of HCHO from antibiotic spots, for observation of the duplicate effect of substances with a direct effect, and for demonstration of cell proliferation promotion and retardation.

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The BioArena system, which integrates the modern technique and biological results of bioautography with layer liquid chromatography is especially suitable for investigating biochemical interactions. Formaldehyde (HCHO) and its reaction products play a crucial role in the antibiotic activity of trans -resveratrol and other molecules — when HCHO-capturing molecules are used in culture media the antimicrobial activity of antibiotic-like compounds decreases substantially. HCHO and hydrogen peroxide are present as normal endogenous compounds in cells, so there is a possibility of interaction in which singlet oxygen ( 1 O 2 ) and excited HCHO can be formed. The 1 O 2 can oxidize water molecules and so H 2 O 3 can be formed, from which, by disproportion, among other reactions, ozone (O 3 ) also can be formed in the chromatographic spots. Elimination of HCHO and/or O 3 from the spots results in a decrease in the antiproliferative effect.

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The BioArena system, which integrates the up-to-date methodology and biological results of bioautography with OPLC as an efficient planar separation technique (compact spots, etc.), is especially suitable for investigating biochemical interactions in an adsorbent bed after chromatographic separation. The first results from BioArena show that formaldehyde (HCHO), which can originate from pathogen cells in some situations, can play a special role in the antibiotic activity of trans -resveratrol. When l -arginine and glutathione were used as endogenous HCHO-capturing molecules in the culture medium the antimicrobial activity of trans -resveratrol on the adsorbent layer decreased substantially. It has been observed that trans -resveratrol generates a time-dependent, and therefore concentration-dependent, duplicate inhibiting effect on the pathogen, and that the BioArena system was suitable for illustration of this new phenomenon. It is probable that this effect occurs as a result of HCHO, with special emphasis on the possibility of interaction between the HCHO and H 2 O 2 of endogenous origin on the adsorbent layer incubated with pathogen cells. It seems that the BioArena system will be an important, indispensable complement to the basic separation technique OPLC.

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