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The electrochemical study of the degradation of the natural dyes from historical objects
Ramešová, Šárka ; Schwarzová, Karolina (advisor) ; Navrátil, Tomáš (referee)
The oxidation of quercetin (3,3',4',5,7-pentahydroxyflavone) and luteolin (3',4',5,7-tetrahydroxyflavone) has been studied. These intensively coloured polyphenolic compounds belong to a large group of naturally occuring flavonoid compounds. They were used with advantage as colourants in old tapestries. The study concerns the analysis of degradation processes of the original dyes. Degradation products formed during the ageing process can be used as fingerprints for the determination of the original colours. The degradation processes of flavonoid compounds were studied by electrochemical methods, absorption spectrometry and separation methods in aqueous media. The oxidation mechanism involves electron transfer coupled with chemical reaction and is influenced by the acidobasic properties of quercetin. The dissociation constants pK1 and pK2 of quercetin and pK1 and pK2 of luteolin were determined by combining of independent methods: acid-base titration and UV-Vis spectroscopy. The stability of both compounds significantly depends on the presence of the oxygen. The distribution of the oxidation products of quercetin in aqueous media within the exposure to oxygen was monitored by LC-MS/MS and LC- DAD analysis.
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NADP-dehydrogenases of medicinal plants.
Bartošová, Martina ; Hýsková, Veronika (advisor) ; Kubíčková, Božena (referee)
Medicinal plants usually contain metabolites formed by secondary biosynthetic pathways for example phenylpropanoid and shikimate pathway, which depend on NAPDH availability. In this work total content of phenolic compounds and flavonoids was determined in 10 medicinal plants common for Czech Republic. Activity of enzymes providing NADPH was determined as well. These enzymes were glucose-6-phosphate dehydrogenase (G6PDH, EC 1.1.1.49), NADP-isocitrate dehydrogenase (NADP-ICDH, EC 1.1.1.42), NADP-malic enzyme (oxalacetate decarboxylating) (NADP-ME, EC 1.1.1.40). The activity of shikimate dehydrogenase (SDH, EC 1.1.1.25), the main enzyme of shikimate pathway, was also determined. Extraction methods were optimized for each assessment. Applicable source for NADP-dependent enzymes are black radish (Raphanus sativus L. var. niger), garlic (Allium sativum L.) and mint (Mentha piperita L). The highest content of phenolic compounds and flavonoids was determined in sage (Salvia officinalis), lemon balm (Melissa officinalis L.) and Mentha citrata. Extraction with methanol was more effective than extraction with hot water. More extraction methods and studies are necessary for characterization of individual compounds and principals on which they work. Key words: medicinal plants, NADP-dependent enzymes, phenolic...
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Flavonoids in some cultivars of Sambucus nigra L. flowers.
Krulišová, Markéta ; Spilková, Jiřina (advisor) ; Martin, Jan (referee)
The aim of this work was to find out the content of flavonoids in the flowers of eleven cultivars of black elder (Sambucus nigra L.) and determine if their contents differ according to the statistical significance. Preparing the list of effects and substances that are contained in the flowers was the goal as well. The drug Sambuci nigrae flos is mostly used for its content of flavonoids and hydroxycinnamic acids in the therapy of colds and diseases of the urinary and respiratory tract. Its benefits were confirmed also by studies testing antibacterial and antiviral activity, diabetes and obesity, effects on the immune system and also protection against UV radiance. The content of flavonoids was established by the spectrophotometric method that is listed in the Czech Pharmacopoeia 2009 in the article Sambuci nigrae flos as the method for the determination of content. The statistic evaluation of the differences in the flavonoids content between the cultivars was made by ANOVA and the Bonferroni test. The demand of Czech Pharmacopoeia 2009 on the minimal content of flavonoids is 0,80%. This requirement was fulfilled in the flowers of these cultivars: Albida, Heidegg 13, Riese auß Voßloch, Sambu, Samdal, Sampo and Samyl. On the opposite side, flowers of the cultivars Allesö, Aurea, Dana and Juicy did...
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Secondary metabolites of plant tissue culture of Trifolium pratense L.
Novotná, Hana ; Kašparová, Marie (advisor) ; Siatka, Tomáš (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Department of Pharmacognosy Candidate: Hana Novotná Supervisor: PharmDr. Marie Kašparová, Ph.D. Title of diploma thesis: Secondary metabolites of plant tissue culture of Trifolium pratense L. Explant cultures are perspective sources of secondary metabolites. Nevertheless production of flavonoids and isoflavonoids by the suspension culture of Trifolium pratense L. is not high. Elicitation is one of the methods used to enhance the biosynthesis of secondary metabolites. Elicitation induces physiological changes, stimulates defensive or stress-induced reactions in plants and subsequently triggers the synthesis of secondary metabolites. The objective of this study was to observe the influence of two elicitors - abscisic acid and ascorbic acid - on the production of flavonoids and isoflavonoids by the Trifolium pratense L. suspension culture (Sprint variety). The culture was cultivated in Gamborg medium to which 2 mg.l-1 of 2,4- dichlorophenoxyacetic acid and 2 mg.l-1 of 6-benzylaminopurine were added, at the temperature of 25 řC and 16 hours light / 8 hours dark period. The best elicitation effect of abscisic acid on the production of flavonoids and isoflavonoids was observed after a 6-hour application of the highest 500 µmol.l-1...
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Preparative diastereomeric separation of silybin and its sulfates by RP-HPLC
Kolářová, Petra ; Tesařová, Eva (advisor) ; Kalíková, Květa (referee)
Silymarin is standardized extract isolated from fruits of Milk Thistle plant (Silybum marianum). The principal component of silymarin is silybin. This flavonolignan is mainly responsible for the medicinal effects of Milk Thistle fruits: antioxidant, hepatoprotective, anticancer and chemoprotective activities. Natural silybin exists as an equimolar mixture of diastereomers A and B whose preparative separation is very hard. It was shown that the biological activity of silybin A and B are different. Silybin in the blood conjugates mainly to sulfates. The structure or biological activity of the sulfates is not yet known. The aim of this work is to develop practically applicable method for preparative separation of diastereomers of silybin A and B, and sulfates, which are considered as one of the major metabolites of silybin. The preparative method for separation of silybin A and B in the mobile phase consisting of 50% MeOH on the chromatographic column Labio C18 25x250 mm, was developed. In addition, preparative method for separation of mixture of products accured from the sulphation of silybin-23-acetate in the mobile phase consisting of MeOH/H2O 60/40 (v/v) with addition of 10 ml/l HCOOH, was optimized.
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The effect of quercetin metabolite on the rat isolated arteries.
Rudišarová, Simona ; Vopršalová, Marie (advisor) ; Pourová, Jana (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacology & Toxicology Student: Simona Rudišarová Supervisor: PharmDr. Marie Vopršalová, CSc. Title of diploma thesis: The effect of quercetin metabolite on the rat isolated arteries This study is focused on the research of the effects of flavonol quercetin and its selected metabolites (3-(3-hydroxyphenyl)propionic acid and 3-hydroxybenzoic acid) on blood vessels in vitro. A standardized method of an isolated rat aorta was used. The aim of this study was to find out how substances act on the isolated aorta and if the effect of the effective metabolite is dependent on the presence of the endothelium in the vessel. After precontracting with noradrenaline, individual concentrations of the test substances were added to the aortic ring bath (final concentrations in the bath ranging from 10-7 to 10-3 M). From the measured values EC50 values (quercetin = 3,63 . 10-5 M, 3-(3-hydroxyphenyl)propionic acid = 3,21 . 10-6 M, 3-hydroxybenzoic acid = 4,38 . 10-5 M) were obtained and the corresponding DRC curves were constructed using S.P.E.L. Advanced Kymograph Software. The experiment was performed once again with quercetin and 3-(3-hydroxyphenyl)propionic acid on the endothelium-denuded vessel. According to the results is...
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Sea buckthorn (Hippophae rhamnoides L.) as a source of biochemically active substances
Blecha, Jan ; Hudeček, Jiří (advisor) ; Hodek, Petr (referee)
Sea Buckthorn (Hippophae Rhamnoides L.) as a source of biochemically active substances ABSTRACT This work is primarily focused on summarizing of existing data in the literature about the chemical composition of sea buckthorn (Hippophae rhamnoides L.) fruits, in particular of its oil. It was found that the content of individual chemical components substantially varies in different cultivars and is also very dependent on the conditions in which the plant is grown. Various sea buckthorn products can be rich sources of antioxidants, such as the vitamin C (in amounts up to 2500 mg/100g of fresh fruit) and tocopherols (up to 481 mg/100g of fresh fruit), flavonoids (up to 1100 mg/100g of fresh fruit) and carotenoids (up to 2139 mg/100g of oil). The fatty acids of this plant are mostly unsaturated (up to 89% of all fatty acids), in dominant amounts linolenic and linoleic acid can be found in seed oil and palmitooleic and saturated palmitic acid in pulp oil. Content and effects of significant flavonoids in the plant are also summarized. Among the most important biological effects of sea buckthorn, its antioxidant activity can be included as well as effects on the function of cardiovascular system, hepatoprotective and anticancer effects proven for some of the substances. Thanks to that the sea buckthorn has a high...
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Properties of plant extracts
Bělonožníková, Kateřina ; Ryšlavá, Helena (advisor) ; Liberda, Jiří (referee)
Plants synthesize a wide variety of compounds called secondary metabolites. These compounds comprise products of the phenylpropanoid pathway (flavonoids, stilbens, anthocyanines and tannins), isoprenoids, and alkaloids. Since plant secondary metabolites seem to affect human health in a positive way, decrease the risk of some diseases such as cancer, diabetes type 2 and disorders of the cardiovascular system, their studies attract a lot of attention. The mechanism of the effect of individual compounds on human health has not been clarified yet, the positive effect is probably caused by their antioxidant capacity. In this work 22 extracts from different kinds of fruit and vegetables were prepared. The total content of phenolic compounds and flavonoids, the antioxidant capacity by methods FRAP (ferric ion reducing antioxidant power) and ABTS [2,2'-azino-bis(3-ethylbenzthiazoline-6- sulphonic acid)] were determined. High content of phenolic compounds was established in the extract from strawberry, cherry and kiwi, the highest antioxidant capacity was found in the extract from strawberry. Another studied attribute of extracts from fruit and vegetables was their influence on the activity of digestive enzymes pancreatic lipase and α-amylase. The extracts from lemon, red currant, jostaberry, lime and...
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Effect of post-harvest processing on quality of Sambuci fructus
Brňáková, Lenka ; Spilková, Jiřina (advisor) ; Kašparová, Marie (referee)
Elderberry is very popular plant, that is due to its beneficial effects to human organism very reputable. The main content substances are flavonoids and anthocyanins. Fruits of this flower are available only seasonally (which is usual for most of flowers), so we have to search for alternative solution of getting them. One of these solutions is conservation. Nowadays there are lots of types of postharvest adjustment, so we have more opportunities to find an optimal concept for every one specific plant. The diploma thesis is focused on assessment of content of phenolic substances harvested in different parts of Hradec Králové. They were conservated at laboratory temperature, elevated (40řC, 60řC) and reduced (-18řC) temperature. Next part of thesis is focused on assessing of the content of anthocyanins in fruits, that had been conservated for 3 years in refrigerator. The theme was finding the best concept of postharvestal adjustment. Despite to the individual places of harvest, there were not mentionable differences found out in content of phenolic substances in fruits. Main differences in content of substances were caused by different temperature of conservating area. With the upper temperature the content of phenolic substances was lower. Storage in the fridge for 3 years did not have any influence...
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Analysis of sulfated flavonoids by HPLC-MS/MS
Petránková, Barbora ; Bojarová, Pavla (advisor) ; Kozlík, Petr (referee)
The aim of this work is to find the optimal separation conditions in high performance liquid chromatography for the analysis of sulfated polyphenolic substances, especially flavonoids, phenolic acids and catechols. Three chromatographic columns were used in the development of the method. ZIC-HILIC column with polar organic phase and C18 and Kinetex PFP columns with reverse phase. We focused in particular on a short time of separation, sharp tailless peaks and separation of all components of the mixture. Using a Kinetex PFP column, the optimal method for separating the test mixtures was found. The use of a mobile phase of 10 mM ammonium acetate/0,1% fromic acid and methanol was shown to be the most appropriate, with a mobile phase flow rate of 0,6 ml/min. By modifying the gradient and temperature, all components of the tested mixtures were separated, the peaks were sharp, and the separation time in most cases ranged do not exceed 10 min. The method is also directly applicable in conjunction with a mass detector. Keywords: High Performance Liquid Chromatography (HPLC), flavonoids, polyphenolic substances, sulfation, separation
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