Václav Diopan

Electrochemical and spectrometric study of antioxidant activity of pomiferin, isopomiferin, osajin and catalposide

The antioxidant properties of pomiferin, isopomiferin, osajin and catalposide are evaluated. The electrochemical behaviour of these compounds at a carbon paste electrode was studied using square wave voltammetry. Oxidative signals, optimized frequencies and appropriate pH acetate buffer conditions were determined. The detection limits (3S/N) for pomiferin, isopomiferin, osajin and catalposide were estimated to be 50 pg/ml, 800 pg/ml, 40 pg/ml and 10 ng/ml, respectively. Furthermore, spectrometric test was employed with 2,2-diphenyl-1-picrylhydrazyle (DPPH) to evaluate the antioxidant activities of these compounds. Based on the obtained results, the highest antioxidant activity measured by DPPH tests was found at pomiferin followed by isopomiferin. The activities of osajin and catalposide were undetectable. The protective effects of pomiferin, isopomiferin, osajin and catalposide on DNA exposed to oxygen radicals in vitro were also studied. Changes in height of oxidative signals for the four bases (guanine, thymine, adenine and cytosine) were measured for DNA exposed to oxygen radicals, generated by Fentons reaction, non-oxidized ssDNA (50 microg/ml) displayed well developed signals; however, after oxidative damage the observed oxidative signals decreased. Significant protective effects were observed for pomiferin and osajin. Decreased effect was observed for isopomiferin while a further reduced protective effect was seen for DNA exposed to catalposide. Based on the obtained results, pomiferin had the highest antioxidant activity followed by isopomiferin, osajin and catalposide.

Phytohormones as Important Biologically Active Molecules - Their Simple Simultaneous Detection

Phytohormones, their functions, synthesis and effects, are of great interest. To study them in plant tissues accurate and sensitive methods are required. In the present study we aimed at optimizing experimental conditions to separate and determine not only plant hormones but also their metabolites, by liquid chromatography coupled with a UV-VIS detector. The mixture we analyzed was composed of benzyladenine, kinetin, trans-zeatin, cis-zeatin, dihydrozeatin, meta-topolin, ortho-topolin, α-naphthalene acetic acid, indole-3-acetic acid, trans-zeatin-7-glucoside, trans-zeatin-O-glucoside, trans-zeatin-9-riboside, meta-topolin-9-riboside and ortho-topolin-9-riboside. We measured the calibration dependences and estimated limits of detection and quantification under the optimal chromatographic conditions (column: Polaris C18; mobile phase: gradient starting at 2:98 (methanol:0.001% TFA) and was increasing to 55:45 during twenty minutes, and then decreasing for 10 min to 35:65, flow rate: 200 µL·min-1, temperature: 50 °C, wavelength: 210 nm). The detection limits for the target molecules were estimated as tens of ng per mL. We also studied the effect of flax extracts on the phytohormones’ signals. Recovery of aliphatic and aromatic cytokinins, metabolites of cytokinins and auxins were within the range from 87 to 105 %. The experimental conditions were tested on a mass selective detector. In addition we analysed a commercial product used for stimulation of roots formation in cuttings of poorly rooting plants. The determined content of α-naphthalene acetic acid was in good agreement with that declared by the manufacturer.