Michal Říha
Charles University in Prague
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Food Chemistry | 2012
Kateřina Macáková; Přemysl Mladěnka; Tomáš Filipský; Michal Říha; Luděk Jahodář; František Trejtnar; Paolo Bovicelli; Ilaria Proietti Silvestri; Radomír Hrdina; Luciano Saso
Flavonoids, substantial components of the human diet, are generally considered to be beneficial. However, they may possess possible pro-oxidative effects, which could be based on their reducing potential. The aims of this study were to evaluate the ability of 26 flavonoids to reduce ferric ions at relevant pH conditions and to find a possible relationship with potentiation of hydroxyl radical production. A substantial ferric ions reduction was achieved under acidic conditions, particularly by flavonols and flavanols with the catecholic ring B. Apparently corresponding bell-shaped curves displaying the pro-oxidant effect of flavonols quercetin and kaempferol on iron-based Fenton reaction were documented. Several flavonoids were efficient antioxidants at very low concentrations but rather inefficient or pro-oxidative at higher concentrations. Flavonols, morin and rutin were progressively pro-oxidant, while 7-hydroxyflavone and hesperetin were the only flavonoids with dose-dependent inhibition of hydroxyl radical production. Conclusively, administration of flavonoids may lead to unpredictable consequences with few exceptions.
RSC Advances | 2014
Michal Říha; Jana Karlíčková; Tomáš Filipský; Kateřina Macáková; Liliana Rocha; Paolo Bovicelli; Ilaria Proietti Silvestri; Luciano Saso; Luděk Jahodář; Radomír Hrdina; Přemysl Mladěnka
Copper is an essential trace element involved in plenty of redox reactions in living systems, however, unbound copper ions cause damage to various biomolecules via excessive generation of reactive oxygen species. Flavonoids, ubiquitous plant secondary metabolites, possess complex effects on human health and chelation of transient metal ions is one of their proposed mechanisms of action. In this in vitro study, 26 flavonoids from various subclasses were screened for their interactions with both copper oxidation states at four (patho)physiologically relevant pH conditions (4.5, 5.5, 6.8 and 7.5) by two spectrophotometric approaches and compared with the clinically used copper chelator trientine. In a slightly competitive environment, the majority of flavonoids were able to chelate cupric ions, however, under more competitive conditions, only flavones and flavonols were able to chelate both cupric and cuprous ions. Apparently, the 2,3-double bond was essential for stable copper chelation. The most efficient copper chelation sites were the 3-hydroxy-4-keto group in flavonols and the 5,6,7-trihydroxyl group in flavones. On the other hand, the 3′,4′-dihydroxyl group was associated only with a weak activity. 3-Hydroxyflavone, kaempferol and partly baicalein were even more potent than trientine in the acidic environment, however, none of the tested flavonoids was able to surpass it at physiological pH or slightly acidic conditions. In conclusion, flavonoids possessing appropriate structural characteristics were efficient copper chelators and some of them were even more potent than trientine under acidic conditions.
Journal of Inorganic Biochemistry | 2013
Michal Říha; Jana Karlíčková; Tomáš Filipský; Kateřina Macáková; Radomír Hrdina; Přemysl Mladěnka
Copper is an essential trace element involved in many physiological processes. Since disorder of copper homeostasis is observed in various pathologies, copper chelators may represent a promising therapeutic tool. This study was aimed at: 1) formation of an in vitro methodology for screening of copper chelators, and 2) detailed analysis of the interaction of copper with clinically used D-penicillamine (D-PEN), triethylenetetramine (trientine), experimentally tested 8-hydroxyquinolines, and the disodium salt of EDTA as a standard chelator. Methodology based on bathocuproinedisulfonic acid disodium salt (BCS), usable at (patho)physiologically relevant pHs (4.5-7.5), enabled assessment of both cuprous and cupric ions chelation and comparison of the relative affinities of the tested compounds for copper. In the case of potent chelators, the stoichiometry could be estimated too. Clioquinol, chloroxine and EDTA formed very stable complexes with Cu(+)/Cu(2+) at all tested pHs, while copper complexes with trientine were stable only under neutral or slightly acidic conditions. Non-substituted 8-hydroxyquinoline was a less efficient copper chelator, but still unequivocally more potent than D-PEN. Both 8-hydroxyquinoline and D-PEN chelation potencies, similarly to that of trientine, were pH-dependent and decreased with pH. Moreover, only D-PEN was able to reduce cupric ions. Conclusively, BCS assay represents a rapid, simple and precise method for copper chelation measurement. In addition, lower binding affinity of D-PEN compared with 8-hydroxyquinolines and trientine was demonstrated.
Journal of Controlled Release | 2013
Barbora Janůšová; Barbora Školová; Katarína Tükörová; Lea Wojnarová; Tomáš Šimůnek; Přemysl Mladěnka; Tomáš Filipský; Michal Říha; Jaroslav Roh; Karel Palát; Alexandr Hrabálek; Kateřina Vávrová
Transdermal permeation enhancers are compounds that temporarily decrease skin barrier properties to promote drug flux. In this study, we investigated enhancers with amino acids (proline, sarcosine, alanine, β-alanine, and glycine) attached to hydrophobic chain(s) via a biodegradable ester link. The double-chain lipid-like substances displayed no enhancing effect, whereas single-chain substances significantly increased skin permeability. The proline derivative l-Pro2 reached enhancement ratios of up to 40 at 1% concentration, which is higher than that of the well-established and standard enhancers Azone, DDAIP, DDAK, and Transkarbam 12. No stereoselectivity was observed. l-Pro2 acted synergistically with propylene glycol. Infrared studies revealed that l-Pro2 forms a separate liquid ordered phase in the stratum corneum lipids and has no significant effect on proteins. l-Pro2 action was at least partially reversible as measured by skin electrical impedance. Toxicity in keratinocyte (HaCaT) and fibroblast (3T3) cell lines showed IC(50) values ranging from tens to hundreds of μM, which is comparable with standard enhancers. Furthermore, l-Pro2 was rapidly decomposed in plasma. In vivo transdermal absorption studies in rats confirmed the enhancing activity of l-Pro2 and suggested its negligible skin toxicity and minimal effect on transepidermal water loss. These properties make l-Pro2 a promising candidate for potential clinical use.
Bioorganic Chemistry | 2013
Tomáš Filipský; Michal Říha; Radomír Hrdina; Kateřina Vávrová; Přemysl Mladěnka
The effects of iron-chelating agents on miscellaneous pathologies are currently largely tested. Due to various indications, different properties for chelators are required. A stoichiometry of the complex in relation to pH is one of the crucial factors. Moreover, the published data on the stoichiometry, especially concerning flavonoids, are equivocal. In this study, a new complementary approach was employed for the determination of stoichiometry in 10 iron-chelating agents, including clinically used drugs, by UV-Vis spectrophotometry at relevant pH conditions and compared with the standard Jobs method. This study showed that the simple approach based on absorbance at the wavelength of complex absorption maximum was sufficient when the difference between absorption maximum of substance and complex was high. However, in majority of substances this difference was much lower (9-73 nm). The novel complementary approach was able to determine the stoichiometry in all tested cases. The major benefit of this method compared to the standard Jobs approach seems to be its capability to reveal a reaction stoichiometry in chelators with moderate affinity to iron. In conclusion, using this complementary method may explain several previous contradictory data and lead to a better understanding of the underlying mechanisms of chelators action.
Biochimie | 2012
Kateřina Macáková; Zuzana Řeháková; Přemysl Mladěnka; Jana Karlíčková; Tomáš Filipský; Michal Říha; Ashok K. Prasad; Virinder S. Parmar; Luděk Jahodář; Petr Pavek; Radomír Hrdina; Luciano Saso
Platelets play a crucial role in physiological haemostasis. However, in coronary arteries damaged by atherosclerosis, enhanced platelet aggregation, with subsequent thrombus formation, is a precipitating factor in acute myocardial infarction. Current therapeutic approaches are able to reduce approximately one quarter of cardiovascular events, but they are associated with an increased risk of bleeding and in some resistant patients are not efficient. Some coumarins possess antiplatelet activity and, due to their additional antioxidant effects, may be promising drugs for use in combination with the present therapeutic agents. The aim of this study was to analyse a series of simple 4-methylcoumarins for their antiplatelet activity. Human plasma platelet suspensions were treated with different aggregation inducers [arachidonic acid (AA), collagen and ADP] in the presence of the 4-methylcoumarins. Complementary experiments were performed to explain the mechanism of action. 5,7-Dihydroxy-4-methylcoumarins, in particular those containing a lipophilic side chain at C-3, reached the activity of acetylsalicylic acid on AA-induced aggregation. Other tested coumarins were less active. Some of the tested compounds mildly inhibited either collagen- or ADP-induced aggregation. 5,7-Dihydroxy-4-methylcoumarins did not interfere with the function of thromboxane synthase, but were competitive antagonists of thromboxane A(2) receptors and inhibited cyclooxygenase-1 as well. 5,7-Dihydroxy-4-methylcoumarins appear to be promising candidates for the extension of the current spectrum of antiplatelet drugs.
Oxidative Medicine and Cellular Longevity | 2015
Jana Karlíčková; Kateřina Macáková; Michal Říha; Liliane Maria Teixeira Pinheiro; Tomáš Filipský; Veronika Horňasová; Radomír Hrdina; Přemysl Mladěnka
Isoflavones are commonly consumed in many Asian countries and have potentially positive effects on human being. Only a few and rather controversial data on their interactions with copper and iron are available to date. 13 structurally related isoflavones were tested in the competitive manner for their Cu/Fe-chelating/reducing properties. Notwithstanding the 5-hydroxy-4-keto chelation site was associated with ferric, ferrous, and cupric chelation, the chelation potential of isoflavones was low and no cuprous chelation was observed. None of isoflavones was able to substantially reduce ferric ions, but the vast majority reduced cupric ions. The most important feature for cupric reduction was the presence of an unsubstituted 4′-hydroxyl; contrarily the presence of a free 5-hydroxyl decreased or abolished the reduction due to chelation of cupric ions. The results from this study may enable additional experiments which might clarify the effects of isoflavones on human being and/or mechanisms of copper absorption.
Phytomedicine | 2017
Lenka Applová; Jana Karlíčková; Michal Říha; Tomáš Filipský; Kateřina Macáková; Jiřina Spilková; Přemysl Mladěnka
BACKGROUND One reason for the lower incidence of cardiovascular diseases in Asian countries may be the high intake of isoflavonoids and their antiplatelet effects may be an important factor. To date, there is limited comparison of a range of isoflavonoids and knowledge of their effects at different levels of platelet aggregation. PURPOSE To screen the antiplatelet effects of a number of isoflavonoids on the arachidonic acid based aggregation pathway and investigate how the antiplatelet activity might occur. METHODS The antiplatelet effects were first screened in whole human blood where platelet aggregation was induced by arachidonic acid. Further analysis was targeted at search of the mechanism of action. RESULTS Thirteen of the eighteen tested isoflavonoids had significant inhibitory effect on platelet aggregation in whole human blood. Genistein had the same potency as clinically used acetylsalicylic acid (ASA) while tectorigenin was clearly stronger than ASA. Further analyses showed that the effect of tectorigenin was not based on inhibition of cyclooxygenase-1 in contrast to ASA or thromboxane synthase but by competitive antagonism at thromboxane receptors. CONCLUSION Tectorigenin is a more potent antiplatelet compound than ASA and thus an interesting substance for further testing.
Oxidative Medicine and Cellular Longevity | 2016
Michal Říha; Pavlína Hašková; Jan Martin; Tomáš Filipský; Kateřina Váňová; Jaroslava Vávrová; Magdaléna Holečková; Pavel Homola; Libor Vitek; Vladimir Palicka; Tomáš Šimůnek; Přemysl Mladěnka
Iron and copper release participates in the myocardial injury under ischemic conditions and hence protection might be achieved by iron chelators. Data on copper chelation are, however, sparse. The effect of the clinically used copper chelator D-penicillamine in the catecholamine model of acute myocardial injury was tested in cardiomyoblast cell line H9c2 and in Wistar Han rats. D-Penicillamine had a protective effect against catecholamine-induced injury both in vitro and in vivo. It protected H9c2 cells against the catecholamine-induced viability loss in a dose-dependent manner. In animals, both intravenous D-penicillamine doses of 11 (low) and 44 mg/kg (high) decreased the mortality caused by s.c. isoprenaline (100 mg/kg) from 36% to 14% and 22%, respectively. However, whereas the low D-penicillamine dose decreased the release of cardiac troponin T (specific marker of myocardial injury), the high dose resulted in an increase. Interestingly, the high dose led to a marked elevation in plasma vitamin C. This might be related to potentiation of oxidative stress, as suggested by additional in vitro experiments with D-penicillamine (iron reduction and the Fenton reaction). In conclusion, D-penicillamine has protective potential against catecholamine-induced cardiotoxicity; however the optimal dose selection seems to be crucial for further application.
Redox Report | 2017
Tomáš Filipský; Michal Říha; Pavlína Hašková; Veronika Pilařová; Lucie Nováková; Vladimír Semecký; Jaroslava Vávrová; Magdaléna Holečková; Vladimir Palicka; Tomáš Šimůnek; Radomír Hrdina; Přemysl Mladěnka
Objectives: Rutin, quercetin-3-O-rutinoside, a natural flavonol glycoside, has shown various in vitro benefits with potential use treating human diseases, especially cardiovascular system disorders. Antioxidant properties are assumed to underlie the majority of these benefits. Yet rutin pro-oxidant properties have been reported as well. Our research group has recently shown aggravating effects on isoprenaline (ISO)-induced cardiotoxicity in Wistar:Han rats after 24 hours. Methods: This study was designed to examine in more detail the reasons for the negative effects of rutin (11.5 and 46 mg/kg, i.v.) after administration of ISO (100 mg/kg, s.c.) in rats within 2 hours of continuous experiment and in the H9c2 cardiomyoblast-derived cell line. Results: Like our previous findings, rutin did not (11.5 or 46 mg/kg, i.v.) reduce the ISO-induced mortality within 2 hours although the lower dose significantly reduced cardiac troponin T (cTnT) and partly improved the histological findings. In contrast, the higher dose increased the mortality in comparison with solvent (1.26% w/v sodium bicarbonate). This was not caused by any specific haemodynamic disturbances. It appears to be associated with oxidative stress as rutin enhanced intracellular reactive oxygen species formation in vitro and had the tendency to increase it in vivo. Conclusions: Rutin, likely due to its pro-oxidative effects, can exacerbate catecholamine cardiotoxicity depending on the dose used.