Perennial plants from the Amaryllidaceae family are generally known for their beauty but also like herbs which contain wide range of alkaloids. To these days more than 500 alkaloids have been isolated. Amaryllidaceae alkaloids (AmA) are derivatived from aminoacid tyrosine and divided into nine basic groups. Biological activity of these substances icludes antitumor, antibacterial, antifungal, antiviral, antimalarial activity and some of them are used for treatment Alzheimer's disease (AD). Narcissus cv. PROFESSOR EINSTEIN have been choosen thanks to previous research of summary extract. Twelve alkaloids have been detected by GC-MS and ten of them have been identified (e.g.: lykoramine, pluviine, haemanthamine, pancracine, homolycorine). Due to this diversity of alkaloids and the fact that summary extract has relatively high inhibitory activity (IC50 = 49,99 ± 5,38 μg/mL) against HuBuChE Narcissus have been appropriate for isolation of the alkaloids and for further study of their biological activity. Summary ethanolic extract for gain pure compounds was prepared from 34.3 kg fresh bulbs. Separation was initiated by column chromatography and extract was divided into almost 500 fractions. Some of them were put together owing to TLC analysis and finally 27 subfractions were formed. Subfraction Nr. 26...

Kohelová E: Alkaloids of Amaryllidaceae family: isolation, structural identification, biological activity. II Diploma thesis, Charles University in Prague, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany and Ecology, Hradec Králové, 2017, 77p. The aim of the diploma thesis was a preparation of alkaloidal extract from fresh bulbs of Narcissus cv. PROFESSOR EINSTEIN to izolate two Amaryllidaceae alkaloids by column and thin layer chromatography. Subsequently these alkaloids were subjected to structural MS and NMR analysis and tested for biological activity against human cholinesterases (HuAChE and HuBuChE) and for cytotoxic activity. In cooperation with the Faculty of Agrobiology, Food and Natural Resources, Czech University of Life Sciences Prague, antifungal and antimicrobial activities of alkaloids were tested, and in cooperation with Faculdade de Medicina da Universidade de Lisboa Portugal antimalarial activity of alkaloids was studied. Isolated substances were identified as galanthamine, caranine and pluviine. Pluviine was isolated in an amount allowing only structural analysis. In the test for determining cholinesterase inhibitory activity of caranine the following values were obtained: IC50, HuAChE = 320 ± 42 μM a IC50, HuBuChE = 486 ± 56 μM. Galanthamine (IC50,...

Schickerová Julie: Alkaloids of Narcissus pseudonarcissus cv. Dutch Master: isolation, structural identification, preparation of analogues, biological activity. Diploma thesis, Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany, Hradec Králové, 2018, 73 p. The purpose of this diploma thesis was to isolate the substances from the fraction ND 15- 9, which was obtained by column chromatography of the alkaloid extract of Narcissus pseudonarcissus cv. Dutch Master. The method of preparative TLC was used to separate this fraction, and the three purified compounds were isolated in the pure state NDS1-NDS3. NMR, GC/MS and optical rotation were used to determine their structure. The obtained data were compared with data in the literature and further studies on their biological activity were performed. Isolated substances were identified as epimaritidine, crinine and tetrahydromasonine. Their inhibitory activities (IC50, AChE > 1000 μM, IC50, BuChE > 1000 μM) versus human erythrocyte AChE and plasma BuChE were inactive compared to galanthamine standards (IC50, AChE = 1.71 ± 0.07 μM, IC50, BuChE = 42.30 ± 0.10 μM), huperzin A standards (IC50, AChE = 0.033 ± 0.001 μM, IC50, BuChE > 1000 μM) and berberin standards (IC50, AChE = 0.71 ± 0.01 μM, IC50, BuChE = 30.7 ± 3.5...

Pancreas is known to be an organ producing a variety of exocrine and endocrine substances, where also insulin belongs. This hormone is produced in the body almost solely by specialized β-cells of the Langerhans islets and is stored here in secretory granules. As the β-cells contain large number of these vesicles, an organism can quickly respond to the glucose stimulation. Completely processed insulin is formed in the secretory granules probably as a hexamer, where six insulin molecules are coordinated along two zinc bivalent cations. Appropriate β-cell response to higher glucose level and following insulin secretion is one of the key processes that regulate metabolism in the body. In order to study insulin production, its effects or secretion, permanent pancreatic cell lines are often used as biological models, out of primary cells from islets of Langerhans. This diploma thesis is focused on two permanent cell lines INS-1E and BRIN-BD11. We searched for the ability of the cells to produce insulin, if the hormone is fully processed, as well as zinc content, which could have a great influence on insulin's processing. Using different methods we compared these two cell lines with cells from the Langerhans islets. We succeeded in isolation of secretory granules from all three cell types and we plan to...

Hanusová P.: Alkaloids of Amaryllidaceae family: isolation, structural identification, biological activity. III. Diploma thesis, Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany and Ecology, Hradec Králové, 2017. For phytochemical study was selected plant cultivar Narcissus cv. PROFESSOR EINSTEIN. The aim of the work was to isolate Amaryllidaceae alkaloids in pure form and screening of their biological activities. Summary alkaloidal extract was prepared from 34 kg of fresh bulbs and separated by column chromatography. Preparative TLC and crystallization were used for the isolation of substances from subfraction 6/1. Two pure alkaloids of galanthamine structural type were obtained. The alkaloids were identified based on their MS, NMR analysis and optical rotation as narwedine and lycoraminone. Lycoraminone has been isolated for the first time from natural source. Both substances were tested for their acetylcholinesterase, butyrylcholinestease and prolyl oligopeptidase inhibition activity. Anticancer activity against two gastrointestinal cancer cell lines Caco-2 and HT-29 (colorectal adenocarcinoma) and antimalarial activity has been also measured. Unfortunately, none of isolated alkaloids showed some significant activity in biological assays. Keywords:...

Dohnalova A: Alkaloids of Amaryllidaceae family: isolation, structural identification, biological activity. I; Diploma thesis, Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany and Ecology, Hradec Králové, 2017, 64 p. The Amaryllidaceae family includes bulbous, beautifully flowering plants that have been used for thousands of years in traditional medicine. The major chemical compounds found in this particular plant family are alkaloids, namely Amaryllidaceae alkaloids. Until now, more than 500 isoquinoline alkaloids have been discovered, which exhibit a diverse biological activity including antitumor and antibacterial. They are also able to inhibit acetylcholinesterase. The aim of the diploma thesis was to process 34 kg of fresh bulbs of Narcissus cv. PROFESSOR EINSTEIN and to prepare an alkaloidal extract. This extract was further divided by column chromatography to almost 500 fractions which were merged based on TLC into 27 subfractions. The subfractione No. 17 was selected for isolation of at least one pure alkaloid. Preparative TLC was used for the the isolation. One pure compound was obtained in crystalic form which was then subjected to structural analysis by EI-MS and NMR methods. Further studies of biological activities were performed in...

Charles University, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany Author: Hana Šimková Supervisor: prof. Ing. Lucie Cahlíková, Ph.D. Title of diploma thesis: Alkaloids of genus Narcissus: isolation, structural identification, biological activity Key words: Narcissus, alkaloids, biological activity, Alzheimer's disease, cytotoxic activity The aim of the diploma thesis was an isolation of alkaloids with a focus on minor fractions. These fractions were obtained from the summary alkaloid extract of Narcissus pseudonarcissus cv. Carlton. The method of preparative TLC was used for the isolation of alkaloids. Three substances of alkaloid origin marked as Fj 3-4/kr, F 7/2-1, F 7/2-3 were isolated from the assigned fractions. These substances were identified as alkaloids of homolycorine type lycorenine, homolycorine and hippeastrine by using GC-MS, NMR and optical rotation. The results were also compared with data in the literature. These three alkaloids were tested for their inhibitory activity against AChE, BuChE, POP and GSK-3β. The inhibitory activity against AChE and BuChE was compared with the reference substances galanthamine (IC50 AChE = 1,71 ± 0,07 μM, IC50 BuChE = 42,3 ± 1,3 μM) and huperzine A (IC50 AChE = 0,033 ± 0,001 μM, IC50 BuChE> 1000 μM). The inhibitory...

Polymerase chain reaction (PCR) is molecular diagnostic method which allows the identification of lactic acid bacteria used in food industry. In this work species-specific PCR primers (targeted on highly conserved 16S rDNA region) were used for identification of bacteria of species Streptoccocus thermophilus in 10 randomly commercially accessible fermented milk products and for identification of species Streptococcus thermophilus in 25 lyophilisates collected in Culture Collection of Dairy Microorganisms Laktoflora (CCDM, Tábor, Czech Republic). The PCR products (968 bp) were detected using electrophoresis in 1,2 % agarose gel. Bacterial DNA was isolated from crude cell lysates by magnetic carriers P(HEMA co GMA) containing carboxyl groups. DNA was reversibly bind on their surface in the presence of high concentrations of poly(ethylene glycol) (PEG 6000) and sodium chloride. Phenol extraction of DNA was used as control. Streptococcus thermophilus strains were identificated using PCR in all analysed samples.

Práce se zabývá návrhem magnetoreologické (MR) vzpěry vibroizolačního systému (VIS) pro kosmický nosič. V rešeršní části jsou popsány vybrané VIS a vzpěry těchto systémů, které byly v kosmických nosičích využity v minulosti. Každá z těchto vzpěr obsahující kapalinu byla těsněna pomocí statických těsnění a pružných vlnovců vyrobených z oceli. Důkladněji byla analyzována vzpěra pasivního systému VIS s označením ELVIS, jehož konstrukce se stala inspirací pro tuto práci. Jedná se o tříparametrický systém, v němž je tlumič uložen na pružině, jejíž tuhost přibližně odpovídá objemové tuhosti vlnovců respektive jejímu průmětu do axiálního směru (pressure thrust stiffness). V práci je představena metodika pro stanovení “pressure thrust stiffness” na základě geometrie vlnovce a také uvedeny parametry vlnovce díky kterým je možné měnit poměr mezi axiální a “pressure thrust stiffness” vlnovce. Tento poměr ovlivňuje v dané koncepci vzpěry její dynamické chování a tím i chování celého VIS. Pro predikci dynamického chování vzpěry byl vytvořen multi-body model VIS založeného na Stewartově plošině a detailnější model jediné vzpěry. Simulace provedené v tomto modelu odhalily parametry, které mají vliv na výkonost tlumiče ve VIS: časová odezva a dynamický rozsah. Díky modelu byl určen rozsah těchto parametrů, ve kterých bude zaručena efektivní funkce vzpěry ve VIS, konkrétně: časová odezva: 0-5ms, dynamický rozsah: 5-10. Před finálním návrhem vzpěry byla sestrojena vzpěra experimentální vzpěra, jejíž parametry byly přesně naměřeny a využity pro verifikaci jednotlivých modelů. Poznatky získané během experimentů byly využity při návrhu finální vzpěry. Jeden z nejdůležitějších poznatků byla nutnost náhrady feritového magnetického obvodu s ohledem na jeho křehkost. Proto byl odvozen tvarový přístup k navrhování rychlých magnetických obvodů z oceli s využitím 3D tisku, který byl následně patentován. Navržená vzpěra obsahuje magnetoreologický ventil jehož odezva je predikována na 1.2 ms a dynamický rozsah 10. V závěru práce je představena metodika, díky které byla vzpěra navržena.