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Content of rutin in selected elderberry cultivars
Píšťková, Magdalena ; Hrstka, Miroslav (referee) ; Vespalcová, Milena (advisor)
The aim of this diploma thesis was to determine the rutin content in springs and leaves of 19 elderberry species (Sambucus nigra L.) with use of liquid chromatography. Furthermore the rutin content was determined in its significant sources – rue and buckwheat. The observed values were compared in order to answer the question whether it is useful to use the elderberry spring and leaves as a new source of rutin. Theoretical part of the thesis describes elderberry in general, its chemical composition and its usage in medicine. Following part concentrates on important substances contained in elderberry – flavonoids. Also principles of extraction techniques and HPLC are discussed. In practice the rutin was gained from plant matrix with use of pressurized hot water extraction (PHWE) which was performed in three five minutes long cycles. Conditions during the PHWE were set up to 80 °C and 150 bars. Obtained extract was purified with use of SPE columns and analyzed by HPLC with DAD detector (wave length – 360 nm). HPLC was performed with SUPELCOSILTMLC-8DB column. The mobile phase was composed of MeOH, H2O, and HCOOH in ration of 36:61,5:2,5. The set up flow of mobile phase was 1 ml/min. In the leaves the highest rutin content was determined in Pregarten – 26,34 ± 0,50 mg/g. On the other hand the lowest rutin content in leaves was determined in Körsör – 3,74 ± 0,13 mg/g. In springs the obtained results were as follow. The highest content was determined in Samyl – 4,87 ± 0,01 mg/g and the lowest content was determined in Heidegg 13 springs – 0,56 ± 0,01 mg/g. Also the HPLC detection limit – 6,3 . 10-5 mg/ml and quantitation limit – 2,1 . 10-4 mg/ml were determined for rutin.
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Assessment of rutin in elderberries
Kaňová, Kateřina ; Hrstka, Miroslav (referee) ; Vítová, Eva (advisor)
This work is focused on determination of the concentration of rutin in the fruits of elderberry (Sambucus nigra L.). The theoretical part provides an overview of the properties, occurrence and use of elderberry in natural healing, medicine and food industry. Furthermore, important substances contained in elderberry, especially flavonoids, including rutin, are mentioned. Properties, biological effects and the possibility of determination of rutin are briefly described. In the experimental part the optimization and validation of the method of high performance liquid chromatography (HPLC), which is used for identification and quantification of rutin in fruits of 18 elderberry species, was performed. The pressurized hot water extraction was used for the preparation and solid phase extraction (SPE) for purification of the sample. The largest amount of rutin was found in a variety of elderberry Albida, 6,70 mg per 1 g of dry matter. High concentrations of rutin included varieties Haschberg, Sambu, Pregarten, Sambo and Sampo. On the other hand, the lowest concentration was found in variety Aurea, 1,24 mg.g-1. Finally, rutin content in elderberries was compared with the content of rutin in elder leaves and branches andwith significant sources of rutin – buckwheat and rue.
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Content of rutin in the biological waste material from plantations of elderberry
Boháčová, Hana ; Vránová, Dana (referee) ; Vespalcová, Milena (advisor)
This thesis is focused on determining the concentration of rutin in twigs elderberry (Sambucus nigra L.). The theoretical part provides an overview of the properties, occurrence and use of elderberry in folk medicine, medicine and food. Furthermore, there are the most important substances contained in elderberry, especially flavonoids, including rutin and quercetin as well. There are briefly described their properties, biological effects and possibilities of determination. The content of the experimental part of the thesis was developed cheap extraction methods to obtain the maximum yield of dried twigs rutin elderberry. Rutin content was observed in twigs cultivated elder varieties and wild elderberry. The identification and quantification of rutin in extracts was optimized and validated method of high performance liquid chromatography (HPLC). As a comparative method to develop extraction was used a pressurized hot water extraction (PHWE). There were tested three particle sizes of shredded twigs. The particles were extracted differently hot water varying duration. Most rutin was found in samples milled at 4 mm particles. The optimum time and temperature of extraction was 7 hours at 80°C. The highest amount of rutin in bred elderberry was determined in a sample Mladší 2014, digestion using 1,49 ± 0,004 mg.100g-1 and method PHWE 5,75 ± 0,015 mg.100g-1. There was determined the most rutin in a sample Září 2014 in the wild variant of elderberry. By digestion of ground twigs was obtained 0,90 ± 0,022 mg.100g-1 of rutin, using PHWE 2,98 ± 0,022 mg.100g-1.
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Extraction of selected elderberry flavonoids for food industry purposes
Grulichová, Hana ; Hrstka, Miroslav (referee) ; Vespalcová, Milena (advisor)
Amount of rutin and quercetin in bark and leaves from different type of Sambucus nigra was determined in this diploma thesis. Records of amounts were compared with amounts in significant sources of these flavonoids. Matters from raw matrix were obtained with pressurized solvent extraction (PSE) and pressurized hot water extraction (PHWE). There was use temperature 120°C, pressure 150 bar, three times 5-min cycles for PSE. There were used 80°C, 150 bar, three 5-min cycles for rutin extracted with PHWE and the same condition were used for extraction of quercetin only the temperature was changed to 100 °C. Analysis of rutin and quercetin were carry out high performance liquid chromatography (HPLC) with diode array detection (DAD), where was used SUPELCOSILTM LC 8DB column (5µm; 250 x 4.6 mm). Methanol: water: formic acid (36: 61.5: 2.5, pH 2.17 2.28) was used like a mobile phase. The flow of the mobile phase was set up 0.7 ml/min. Size of sample was 10 µl. The most amount of rutin was determined in the leaves of wild kind of Sambucus nigra with methanol extraction and hot water too. Extraction with PHWE was more effectively. There were determined 5.58 mg of rutin in one gram of sample from wild Sambucus nigra. There were determined the smallest amount of rutin in Sambucus nigra Körsör. There was found only 0.13 mg/g with extraction PHWE. The quercetin wasn’t determined in any Sambucus nigra leaves. There were found both of the flavonoids (rutin and quercetin) in the branch of Sambucus nigra. The rutin was determined in the small amount in wild kind but it was found in the important amount in Sambucus nigra cultivar. Whereas the quercetin was found in higher concentration than rutin when was finding the ideal conditions for extraction of branch. The quercetin was chosen like more important in light of amount. In the end the quercetin wasn’t detected in any Sambucus nigra cultivars. The most amount of quercetin was determined in wild sambucus nigra, 0.24 mg/g. The most amount of rutin was found in Sambucus nigra Albida, 2.33 mg/g.
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Possibility of determination of selected flavonoides
Smělá, Margita ; Vitoulová, Eva (referee) ; Vespalcová, Milena (advisor)
The theoretic part of the bachelor‘s thesis deals with description of selected flavonoids as apigenin, quercetin, luteolin, keampferol, rutin, myricetin and with options of their determinations. There were described typical properities and effects of several flavonoids at first. In the next part there are principles of methods, especially high-performance liquid chromatography and capillary electrophoresis. The biggest attention was made to literary survey of determination flavonoids coumpounds mentioned above, by high-performance liquid chromatography and capillary electroproresis. There were described the collection of plant material for extraction of flavonids in the practical part. The white and blue grapes were frozen by dry ice as soon as they were separated from a busch. Grapes were made in a nitrogenous atmosphere and lyophilizated after the frozen.
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Srovnání antioxidačních vlastností a absorbce UV záření u extraktů vybraných bylin, Spiruliny a jejich směsí
Farinasso, Giulia
This study aimed to compare the antioxidant properties and UV radiation absorption of se-lected herbal extracts, blue-green algae spirulina extract and their mixtures. Extracts were ob-tained by ultrasound assisted extraction. The total polyphenol (TPC) and flavonoid contents (TFC) were determined, and the Trolox equivalent antioxidant capacity (TEAC) was meas-ured. The best antioxidant properties has been showed by C. officinalis individual extract and its mixture with spirulina. Additionally, the UV-absorbing substances at 355 nm were quanti-fied. The extracts from C. officinalis and S. nigra were further analysed for their rutin content by HPLC-DAD. The rutin content in S. nigra and C. officinalis extracts was found to be 1935,97 ± 98,39 μg.g-1 and 233,035 ± 8,839 μg.g-1 of dry weight of flos, respectively. The highest content of phenolic compounds was found in the S. nigra extract, and the highest total antioxidant capacity was determined for the C. officinalis extract. Pearson's correlation re-vealed that the overall antioxidant capacity and UV absorption results were significantly cor-related with the total phenolic compound content only for the elderflower extract. The syner-gistic effect of the herbs used in combination with spirulina was evaluated.
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Rutinosidase from Aspergillus niger - analysis of active site and its mutagenesis
Šidáková, Anna ; Bojarová, Pavla (advisor) ; Palyzová, Andrea (referee)
Rutinosidases (α-L-rhamnosyl-β-D-glucosidases) from Aspergillus niger (AnRut) are glycosidases (EC 3.2.1) that catalyze the hydrolysis of the glycosidic bond between the aglycone and the disaccharide residue rutinose. The dual substrate specificity of this enzyme group describes the parallel activity towards the substrates rutin (carrying a rutinosyl disaccharide residue) and isoquercitrin (carrying a glucosyl residue). The active site of AnRut is more complex than that of other glycosidases and is composed of the catalytic amino acids Glu210 and Glu319 in the active-site cleft and a side tunnel. This untraditional structure with distinct interactions in the tunnel and active-site cleft is the probable reason for the enzyme exceptional substrate specificity. Through point or multiple mutations of the enzyme, we can modify its primary and secondary structure, thus causing a significant shift in substrate specificity. The main goal of this thesis is the analysis of three distinct mutant variants of AnRut rutinosidase; their production, purification, and the study of the influence of the mutations on the substrate specificity of the enzymes. All variants were designed based on molecular modeling. The substrate specificity was determined by reactions of the mutant variants with previously unstudied...
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Antioxidative parameters of fruit model product.
KRATOCHVÍLOVÁ, Lenka
The content of the main components (anthocyanins, quercetin, chlorogenic acid, L-ascorbic acid, rutin, total polyphenols) was determined in the syrup of elderberries (Sambucus nigra L.) by spectrophotometry or high-performanceliquid chromatography. Furthermore, the antioxidant activity of this syrup was determined using three methods: ABTS, FRAP, and DPPH. The measurement results were statistically processed and compared using correlation analysis, specifically using the Pearson correlation coefficient. It was found that the prepared product contains 2160 mg/kg anthocyanins, 201 mg/kg rutin, 26 mg/kg chlorogenic acid and 679 mg/l polyphenols. No free quercetin was found. The content of L-ascorbic acid in the original elderberries was relatively low and therefore for study purposes, the syrup was enriched with this substance in the amount of 0.5% (w/w) to study the connection of its content with other substances. It is clear from the statistical processing that the concentration of chlorogenic acid correlates with the antioxidant activity measured by the FRAP method and the concentration of polyphenols to the antioxidant activity measured by the DPPH method.
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Enzymatic and Metabolic Transformation of Silybin and its Congeners
Purchartová, Kateřina
Natural flavonoids and flavonolignans feature beneficial properties for living organisms such as antioxidant and hepatoprotective effects, anticancer, chemoprotective, dermatoprotective and hypocholesterolemic activities. Their metabolism in mammals is complex, the exact structure of their metabolites still remains partly unclear and the standards are usually not commercially available. Hence, this project focused on the preparation of potential and defined biotransformation Phase II sulfated metabolites of silymarin flavonolignans: silybin, 2,3-dehydrosilybin, isosilybin, silychristin, silydianin and flavonoids quercetin, taxifolin, rutin and isoquercitrin. Pure sulfated derivatives were prepared using aryl sulfotransferase from Desulfitobacterium hafniense and aryl sulfotransferase from rat liver. Using heterologously expressed PAPS (3'-phosphoadenosine-5'-phosophosulfate) - independent arylsulfotransferase from Desulfitobacterium hafniense and cheap p-nitrophenyl sulfate as sulfate donor, sulfated flavonolignans and flavonoids were obtained in high yields. Silymarin flavonolignans afforded exclusively monosulfates at the position C-20 (C-19 in the case of silychristin), except 2,3-dehydrosilybin that yielded also the 7,20-O-disulfated derivative. Isoquercitrin and rutin were selectively sulfated...
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