Overpressured layer chromatography: from the pressurized ultramicro chamber to BioArena system

The pressurized ultramicro (UM) chamber as a closed adsorbent layer chamber enables the use of a special chromatoplate and a pump to increase and optimize the mobile phase flow velocity through an optional development distance in an adsorbent layer. This chamber is the basic instrument of overpressured-layer chromatography (OPLC), which is a separation technique that combines the advantages of conventional TLC/HPTLC with those of HPLC. The versions of OPLC instrument, the character and achievement of off-line and on-line OPLC systems in analytical and preparative use are described. The development of BioArena as a complex bioautographic system means an exploitation of the unique advantages of planar-layer system for detection, isolation and identification of new antimicrobials, antineoplastics, biopesticides and other biologically active substances as well as for studying fundamental biochemical reactions and mechanisms

Study of trace elements in BioArena system and in in vivo conditions

The adsorbent layer system is especially suitable for the biological evaluation of different compounds and trace elements as well. Present experiments showed that formaldehyde (HCHO) molecules participate in the antibiotic activity of Cu (II) ion, an „old antibiotic”. The elimination of HCHO from the chromatographic spots (e.g. by reduction or capturing) resulted in a characteristic decrease of the antibiotic effect of trace elements. The trace elements are HCHO carriers and generate a double effect (first step: deprivation of HCHO as also biological effect; second step: release of HCHO with big killing activity). These features offer good opportunities for influencing fundamental biochemical pathways. It has been established that the trace elements (mainly transition metal ions as e.g. Ni(II) ion) always generate quadruple, bioequivalent, specific immune-stimulating activity in plants with a non-linear dose-response. HCHO and its reaction products (mainly O3) are responsible also for this latter activity

Separation and identification of antibacterial chamomile components using OPLC, bioautography and GC-MS

Components of 50% ethanolic chamomile (Matricaria recutica L.) flower extract, previously found antibacterial in a TLC-bioautographic study, were separated and isolated by the use of on-line OPLC, which consisted of an OPLC 50 BS system, an on-line coupled flow-through UV detector, and a manual fraction collector. The collected peaks were investigated by GC-MS analysis and by TLC re-chromatography with subsequent visualization, performed after use of the vanillin-sulphuric acid reagent, or under UV illumination, or applying bioautographic detection. The main compounds of the collected 11 fractions were identified by GC-MS. The results showed that the antibacterial effect of 50% ethanolic extract of chamomile is ascribable to cis-, trans-spiroethers, and the coumarins like herniarin and umbelliferone

PROTEIN AND AMINO ACID CHEMICAL STUDIES

A cross-linking procedure not deteriorating the physico-chemical characteristics of wool has been elaborated. The thermal destruction processes in wool have been clarified. Solutions have been found for accelerating and inhibiting reactions taking place between wool and formaldehyde. The reactions between L-Iysine and formaldehyde as well as L-arginine and formaldehyde have been identified. Procedures have been elaborated for the practical utilization of the reaction products

A transz-rezveratrol és rokonvegyületek vizsgálata BioAréna-rendszerben

A vörös bor kedvező biológiai hatásaiért elsősorban a kékszőlő termés héjából a vörösborba kerülő sztilbénvázas transz-rezverátrol (TR) a felelős. A TR sokrétű hatásának értelmezése a hatás mechanizmus teljesen uj megközelítését igényelte. Feltételeztük korábban, majd bizonyítottuk , hogy egyrészt az antibiotikus hatású TR-rel együtt a szőlőnövényben a formaldehid (HCHO)kötött formában együttesen fordul elő, másrészt kémiai reakciókban pedig bizonyitottuk hogy a TR reakcióba lép a HCHO-val, amikor nagyon reaktiv hidroximetil-származékok képződnek.Az elmult időszakban a TR és a HCHO közötti reakciókat mikrokémiai szinten tanulmányoztuk. Az un BioAréna rendszer (a direkt bioautográfia továbbfejlesztett változata és a müveleti lépések koordinálása) lehetővé tette a TR és a HCHO közötti reakciók szisztematikus tanulmányozását. Megállapítottuk, hogy a TR kettős hatást ad, melynek első lépése a HCHO molekulák felvétele az adott rendszerből, s ez önmagában biológiai (védő) hatást jelent. Az igy keletkezett uj hidroximetil TR származékok antiproliferativ hatást mutatnak (pl. mikrobák és tumor sejtek ellen).Ozon-befogó molekulák adásával nem tudtuk a TR antibakteriális hatását csökkenteni. A BioAréna-rendszerben különböző hazai és külföldi fehér és vörös borfajták antibakteriális spektrumát is összehasonlitottuk: a fehér borfajták általában mérsékelt antibiotikus hatásukkal tüntek ki minőségi és mennyiségi szinten egyaránt.. | For the beneficial activities of red wine mainly a stilbene derivative, trans-resveratrol (TR) which is originating from the skin of red grapes into the red wine, is responsible. The explanation of the multiple biological activity of TR required a totally new approach to the mechanism of action. We supposed earlier and then proved that the TR in the grape plant occurs together with bound formaldehyde (HCHO) and we proved in chemical reactions that TR can react with HCHO forming very reactive hydroxymethyl derivatives. More recently the interaction between TR and HCHO can be studied in microchemical level: the BioArena system (the developed version of direct bioautography and the coordination of operating steps) gave a possibility for the systematic study of the interactions between TR and HCHO . It has been established that TR generates a double effect. The mobilisation of HCHO from a given biological unit (e.g. tissue) with TR (first step) may cause a chemopreventive /biological effect and the reaction products (hydroxymethyl derivatives) between TR and HCHO (second step) may exert a killing/inhibiting effects on pathogens and/or cancer cells (double effect). Using ozone (O3) capture molecules (e.g. limonene) in the chromatographic spots the antibacterial activity of TR didn't decrease. Comparison of antibacterial activity of white and red wines showed the moderate antibacterial activity of white wines quantitatively and qualitatively, alike.

Application of direct bioautography and SPME-GC-MS for the study of antibacterial chamomile ingredients

The isolation and characterization of antibacterial chamomile components were performed by the use of direct bioautography and solid phase microextraction (SPME)-GC-MS. Four ingredients, active against Vibrio fischeri, were identified as the polyacetylene geometric isomers cis- and trans-spiroethers, the coumarin related herniarin, and the sesquiterpene alcohol (-)-alpha-bisabolol

Bioassay-guided isolation and identification of antimicrobial compounds from thyme essential oil by means of overpressured layer chromatography, bioautography and GC-MS

A simple method is described for efficient isolation of compounds having an antibacterial effect. Two thyme (Thymus vulgaris) essential oils, obtained from the market, were chosen as prospective materials likely to feature several bioactive components when examined by thin layer chromatography coupled with direct bioautography as a screening method. The newly developed infusion overpressured layer chromatographic separation method coupled with direct bioautography assured that only the active components were isolated by means of overrun overpressured layer chromatography with online detection and fractionation. Each of the 5 collected fractions represented one of the five antimicrobial essential oil components designated at the screening. The purity and the activity of the fractions were confirmed with chromatography coupled various detection methods (UV, vanillin-sulphuric acid reagent, direct bioautography). The antibacterial components were identified with GC-MS as thymol, carvacrol, linalool, diethylphthalate, and alpha-terpineol. The oil component diethyl-phthalate is an artificial compound, used as plasticizer or detergent bases in the industry. Our results support that exploiting its flexibility and the possible hyphenations, overpressured layer chromatography is especially attractive for isolation of antimicrobial components from various matrixes