Aims of the study: In South Africa, the genus Helichrysum is widely used in traditional medicine. The uses are well documented although renaming of species and the resulting confusing taxonomic nomenclature may cause uncertainty as to which specific species was referred to in some reports. The aim of this paper is to present a collated and coherent overview of the documented traditional uses of Helichrysum species and to update the botanical identity of previously studied species. Materials and methods: Databases (Scifinder, ISIWeb of Knowledge) and several bookswere used to collect in information on South African Helichrysum species. Results: The traditional uses, chemistry and biological activity of Helichrysum species have been summarized. It was attempted to give clarity as to exactly which species is refer to in the ethnobotanical literature. Conclusions: Although a largenumber of ethnopharmacological uses have beendocumentedand the chemistry of the genus has been studied extensively, only a few South African species have been investigated for their biological activity.

No description available.

Student Number : 0009899J - PhD thesis - School of Therapeutic Sciences - Faculty of Health Sciences / The genus Eriocephalus commonly known as ‘wild rosemary’, ‘Cape snow bush’, or ‘kapokbos’ is a member of the family Asteraceae (tribe Anthemideae). The genus is endemic to southern Africa, with the highest concentration of species in the Western and Northern Cape. The genus comprises 32 species and a total of 42 taxa, which are distributed in South Africa, Namibia, Botswana, and Lesotho. The characters used in species delimitation are purely based on morphological variation in floral and foliar parts and are highly homoplastic due to phenotypic plasticity. In many cases these features are not sufficiently distinctive, as some taxa tend to exhibit dimorphism in some character states such as the presence of opposite and alternate leaves. In some species there is extensive intergrading of the major diagnostic characters leading to uncertainty in species delimitation. Both chemical and molecular characters were used in this study in an attempt to evaluate current species delimitations in the genus, along with species-level relationships and affinities. The genus is also economically important with some of its members used as medicinals, fodder, perfumes, and cosmetics. This warrants investigation into the phytochemistry and biological activity of these species in order to determine a scientific rationale for their traditional uses. For this reason, the antimicrobial, antiinflammatory, antioxidant activities, and inhibition of acetylcholinesterase by the volatile oils and leaf extracts of the genus, which are relatively unknown for most members of the genus, were also investigated. Representatives of 22 species of the genus, eight of which were from Namibia and 14 from South Africa were collected from wild populations. In most cases multiple collections per population per species were considered. Aerial plant parts were hydrodistilled to obtain the essential oils, and phenolics were extracted from leaves using acetone. Essential oils were analysed by thin layer chromatography (TLC), gas chromatography (GC), gas chromatography coupled to mass spectroscopy (GC/MS), and phenolics were analysed using thin layer chromatography (TLC) and high performance liquid chromatography (HPLC/UV). Biological assays were carried out using the 5-lipoxygenase enzyme to evaluate antiinflammatory activity; disc diffusion and microtitre plate dilution assays were used to assess antimicrobial activities of selected fungi and bacteria; the TLC-DPPH and DPPHmicrotitre methods were used to investigate antioxidant activities and a TLC-bioautographic assay was used for testing the inhibition of the acetylcholinesterase enzyme. Total genomic DNA was extracted from silica dried leaf material. The non-coding plastid DNA regions, the psbA-trnH intergenic spacers and the internal transcribed spacer (ITS) region of nuclear ribosomal DNA were amplified, and sequenced and analysed using the parsimony algorithm. The essential oils are largely comprised of acyclic, monocyclic, and bicyclic regular and irregular mono- and sesquiterpenes of various structural groups. Two hundred compounds were noted in the essential oils with some of the common constituents being; α- and β-pinene, yomogi alcohol, ρ-cymene, 1,8-cineole, camphor, 4-terpineol, spathulenol, caryophyllene oxide, α-copaene and β-caryophyllene. Most of the species have a relatively high content of 1,8-cineole and camphor. Twenty-two chemotypes were noted and the potential for commercial development in the flavour, fragrance and pharmaceutical industries has been recorded. Among the favourable chemotypes noted includes the camphor, 1,8-cineole, bisabolol oxide B and nerolidol rich oils. However, due to the extensive variability in the essential oil profiles, standardization of oils in commercial development is crucial. The leaf extracts comprised of flavonoids with the flavones and flavanones as the major structural types present in most species. The terpene and flavonoid chemistry of the genus is highly divergent even among multiple individuals of the same species and hence not a good taxonomic marker for specific delimitation as no coherent groups was evident although some phytochemical congruence has been noted between some of the taxa. The DNA sequence data revealed lack of variability in the non-coding regions psbA-trnH and trnL-F among species of the genus. The nuclear DNA region (ITS) was variable but the number of characters separating taxa was too few for resolution of relationships between taxa. Presence of highly divergent paralogous repeats of ITS were also noted in some taxa. The combination of molecular and chemical data did not resolve the species delimitation problems due to the highly variable distribution of characters within a single species. The patterns of variation observed in the genus may be attributed to chemical convergence, divergence, hybridisation, differential gene expression, polymorphism and allelochemical diversification among other factors. The lack of coherence in the phylogenetic and phenetic groupings of the various taxa implies that the current species boundaries may not be a true reflection of natural taxonomic entities. The use of multiple taxa in taxonomic studies is strongly recommended due to the extensive variability noted in the chemical profiles of the taxa that is also depicted in the phylogenetic histories. It also implies that caution should be taken in bioprospecting for new natural products for commercial development, as plant chemical profiles especially from the same species can be very variable. This implies carrying out exhaustive population and genetic studies for evaluation of diversity in the study group. In the antimicrobial assay, the oils were more active against the Gram-positive bacteria (2-16 mg/ml) and yeasts (1-16 mg/ml). Bacillus cereus and Cryptococcus neofomans were the most susceptible pathogens to the oils. The extracts exhibited low activity against the test pathogens except E. aromaticus and E. pinnatus with activity of 0.2 mg/ml against Staphylococcus aureus and Bacillus cereus respectively. The susceptibility of the fungal pathogens Cryptococcus neoformans and Candida albicans and the Gram-positive bacteria Bacillus cereus to the oils and extracts is an indication of the potential for use of the members of the genus as natural antibiotics. The essential oils exhibited antiinflammatory activities with IC50 values ranging between 19.0-98.6 μg/ml. The oils did not show antioxidant activity at the starting concentration of 100 μg/ml but the acetone leaf extracts exhibited antioxidant activities with IC50 values ranging between 21.5-79.6 μg/ml. The essential oils showed inhibitory activity against acetylcholinesterase enzyme. The biological activity of the oils indicates that most of the traditional uses are influenced by the presence of the oils. The in vitro biological activity of the essential oils and extracts against the test pathogens provides a scientific basis for the use of some of the members in traditional herbal remedies and validates the use of some of the members of the genus for treatment of respiratory tract infections, gastro-intestinal disorders, mental conditions, dermal infections, and inflammation. The study records the biological activities for some of the species for the first time and their potential for use in flavourings, perfumery, cosmetics, as sources of antimicrobial drugs, permeability enhancers in pharmaceutical formulations and for use as industrial oils.

Charles University Faculty of Pharmacy in Hradec Králové Department of Pharmacognosy Candidate: Michaela Teplanská Supervisor: PharmDr. Daniela Hulcová, Ph.D. Title of diploma thesis: Derivatives of Amaryllidaceae alkaloid vittatine as potential drugs Haemanthamine type Amaryllidaceae alkaloids are characterized by interesting biological activity. This group also includes alkaloid vittatine with antitumor, antibacterial, antifungal and antimalarial effects. Although vittatine does not inhibit cholinesterases, its derivatives have shown promising activity against butyrylcholinesterase, which is one of the targets of potential drugs in the treatment of Alzheimer's disease. Another series of semisynthetic vittatine derivatives was prepared in order to examine their biological activity. Reactions with acyl chlorides gave 11 aromatic esters. Identification of the prepared substances was performed by ESI HRMS, NMR and optical rotation measurements. The derivatives were tested for inhibitory activity against human cholinesterases. The results show that the substances were not active against acetylcholinesterase, but almost all of them inhibited butyrylcholinesterase. The most active was 3-O-(6-chloro-2- fluoro-3-methylbenzoyl)vittatine with an IC50 value 0.29 ± 0.03 μM. According to the calculated value...

From the reaction of ZnCl2 with the non-steroidal anti-inflammatory drug diflunisal (Hdifl), complex [Zn(difl-O)(2)(MeOH)(4)], 1 was formed, while in the presence of a N,N'-donor heterocyclic ligand 2,2'-bipyridylamine (bipyam), 2,2'-bipyridine (bipy), 1,10-phenanthroline (phen) and 2,2'-dipyridylketone oxime (Hpko), the complexes [Zn(difl-O,O')(2)(bipyam)], 2, [Zn(difl-O,O')(2)(bipy)], 3, [Zn(difl-O,O')(2)(phen)], 4 and [Zn(difl-O)2(Hpko)(2)], 5 were isolated, respectively. The complexes were characterized by physicochemical and spectroscopic techniques and the crystal structures of complexes 2, 3 and 5 were determined by X-ray crystallography. The ability of the complexes to scavenge 1,1-diphenyl-picrylhydrazyl, 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) and hydroxyl radicals and to inhibit soybean lipoxygenase was studied and the complexes were more active than free Hdifl. The interaction of the complexes with serum albumins was monitored by fluorescence emission spectroscopy and the corresponding binding constants were calculated. UV-vis spectroscopy, viscosity measurements and fluorescence emission spectroscopy for the competitive studies of the complexes with ethidium bromide were employed to investigate the interaction of the complexes with calf-thymus DNA and revealed intercalation as the most possible DNA-binding mode. Computational techniques were used to identify possible binding sites of albumins and DNA, and determine the druggability of human and bovine serum albumins with the five novel complexes. The majority of the complexes are stronger binders than the free Hdifl. This is the first study incorporating experimental and computational results to explore the binding activity of metal-NSAID complexes with DNA and serum albumins, suggesting their application as potential metallodrugs.

Author: Petra Nekolná Title: Alkaloids of family Amaryllidaceae: genus Lycoris Diploma thesis Charles University in Prague, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany and Ecology 2015, 79 p. The aim of this diploma thesis was to summarize the findings about alkaloids which were isolated from Lycoris plants of Amaryllidaceae family. It contains a botanical characteristics of species of genus Lycoris which were studied phytochemically, a file of alkaloids which were isolated from these plants and findings about the biological activity of these compounds. Within the genus Lycoris 11 species were studied phytochemically and 118 alkaloids were isolated from these plants. Alkaloids which were isolated from Lycoris plants are divided in several structural groups. The lycorine-, homolycorine-, crinine-, galanthamine- and pancratistatine-type alkaloids occur the most numerously. Anticancer, acetylcholinesterase-inhibitory and antimalarial activity of the alkaloids were described. The biological activity of alkaloids is connected with their structure. The most significant anticancer activity was observed in alkaloids from lycorine-, crinine- and pancratistatine-type. Acetylcholinesterase-inhibitory activity was pronounced the most in galanthamine-type alkaloids. The most...

Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Pharmaceutical Botany and Ecology Candidate: Zuzana Kavková Supervisor: doc. Ing. Lucie Cahlíková, Ph.D. Title of diploma thesis: Alkaloids of family Amaryllidaceae and their analogues as potential drugs The object of this diploma thesis was to prepare derivatives of alkaloids of Amaryllidaceae family and to deal with their biological activity. These alkaloids are famous for their antibacterial, antiinfectives, antifungal, antimalarial and inhibitory activity against AChE, BuChE and POP and also for cytotoxic effect against cell lines. In the current studies about anticancer activity it was found that the most active alkaloids are Amaryllidaceae alkaloids of these types: lycorine, crinane and pancratistatine. Their biological activity relates closely with their structure. The changes of different parts of the structure can explain the relationship between structure and activity, and also the importance of their organization which is necessary for starting the activity. Based on this finding were for the experiments chosen alkaloids like haemanthamine, haemanthidine and lycorine. An eleven derivatives were prepared and identified mostly by GC-MS and NMR. These derivatives were tested on a wide spectrum of tumor lines....

Charles University in Prague, Faculty of Pharmacy in Hradci Králové Department of Pharmaceutical Botany and Ecology Lucie Gábrlová Diploma thesis: Alkaloids of family Amaryllidaceae and their biological activity The aim of this diploma thesis was to compile assigned fraction of alkaloidal extract obtained from Chlidanthus fragrans and isolation of at least two alkaloids for testing their biological aktivity. For the processing of the extract and isolation of the alkaloids contained in it was used preparative TLC. The obtained substances were then undegone structure analysis, specifically, there were used EI-MS and NMR methods. Based on the results obtained were isolated substances identified and prepared for screening of their biological activity, which was not part of this diploma thesis anymore. The isolated alkaloids were identified as 6α-hydroxybuphanidrine, crinamidine and crinine. 6α-Hydroxybuphanidrine failed to get enough for biological testing. Inhibitory activity of crinamidine and crinine against erythrocyte AChE and serum BuChE was quite weak. Equally, neither of alkaloids showed stronger ability to inhibit the POP, where for crinamidine was determined IC50 = 0,790 ± 0,062 mM and for crinine IC50 = 1,473 ± 0,122 mM. The measurment of cytotoxic activity has been carried out only with...

Mesmo antes do advento da escrita, o homem já buscava na natureza a cura para suas doenças. Dentre as diferentes classes de produtos naturais usados para esse fim, destaca-se a dos alcalóides, por exibir relevante diversidade estrutural e farmacológica, e que tem como fonte importante o gênero Aspidosperma, do qual foram isolados alcalóides que têm sido usados nas terapias atuais. Nesse contexto, o presente trabalho teve como objetivo investigar os alcalóides obtidos a partir de diferentes partes de duas espécies ainda pouco exploradas: Aspidosperma ramiflorum e Aspidosperma tomentosum quanto à sua composição e atividades antifúngica, antimicrobiana e antitumoral. Foram identificados através de Cromatografia à Gás acoplada à Espectrometria de Massas (CG-EM) e Ressonância Magnetica Nuclear os alcalóides beta-ioimbina, nos pericarpos e arilos de A. ramiflorum, uleína, nas folhas, ramos e sementes de A. tomentosum, além de quebrachamina e razinilama, nos arilos e sementes dessa espécie. Adicionalmente, com base na literatura e nos resultados das análises por CG-EM verificou-se em A. tomentosum a presença de desmetileno-oxa-uleína, nas folhas, de eburnamonin-19-ona nos ramos e de di-hidro-razinilama, de aspidospermidina e também de di-hidro-razinilama e de 1-acetil-17-metóxi-aspidospermidina nas sementes e arilos dessa mesma espécie. Nenhum dos extratos de alcalóides totais ou alcalóides isolados apresentou atividade antibacteriana relevante. Verificou-se atividade antitumoral em extratos de alcalóides dos pericarpos de A. ramiflorum e dos ramos, folhas e arilos de A. tomentosum. Observou-se ainda atividade antifúngica em todos os extratos de alcalóides ensaiados, além de atividade específica contra isolados de Criptococcus neoformans resistentes ao fluconazol para os alcalóides quebrachamina e beta-ioimbina. O extrato de alcalóides totais das folhas de A. tomentosum, além da atividade frente a essas leveduras, também se mostrou ativo frente a duas espécies de Candida (C. krusei e C. parapsilosis). Concluiu-se a partir dos resultados obtidos que as duas espécies estudadas podem ser consideradas fontes promissoras de alcalóides que poderão ser utilizados como modelos para novos fármacos antifúngicos e antitumorais. / Even before the advent of writing, man already searched the nature for the cure of his illnesses. Among the different classes of natural products used for this purpose, monoterpenoid indole alkaloids exhibit large structural and pharmacological diversities and these compounds are commonly found in the Aspidosperma genus. In this context, the present work had as main goals to investigate the alkaloids from different parts of two poorly studied species, Aspidosperma ramiflorum and Aspidosperma tomentosum, regarding their composition and antibacterial, antifungal, and antitumoral activities. Gas Chromatography coupled to Mass Spectrometry (GC-MS) and Nuclear Magnetic Ressonance analysis allowed the identification of the following alkaloids: beta-yohimbine, in the pericarp and arils of A. ramiflorum; uleine, in leaves, branches and seeds of A. tomentosum; quebrachamine and rhazinilam, in the arils and seeds of the same species. Moreover, based on literature data and the results of the GC-MS analysis was characterized in A. tomentosum the presence of oxo-uleine, in the leaves and stems, eburnamonine, in the stems, and rhazinilam, aspidospermidine, dihydrorhazinilam and 1-acetyl-17-methoxy-aspidospermidine like alkaloids in the seeds and arils. Neither crude alkaloid extracts nor isolated alkaloids presented a relevant antibacterial activity. An antitumoral activity could be detected in crude alkaloids extracts from pericarps of A. ramiflorum and from stems, leaves and arils of A. tomentosum. On the other hand, antifungal activity was observed in all crude alkaloid extracts assayed, besides a specific activity of the alkaloids quebrachamine and beta-yohimbine against fluconazol resistant Criptococcus neoformans. Additionally, it was observed that crude alkaloid extract from leaves of A. tomentosum was also active against two Candida species (C. krusei e C. parapsilosis), as well as to the C. neoformans isolate. From these results we can conclude that the two studied species has to be considerated as promising sources for antifungal and antitumor alkaloids to be used as models to design new drugs.

O presente trabalho enfocou o isolamento, a análise da atividade biológica e a caracterização dos componentes majoritários do Photogem® através de Cromatografia Líquida de Alta Eficiência (CLAE), espectroscopia eletrônica na região do ultravioleta-visível (UV-Vis) e espectrometria de massa. O Photogem® é um derivado de hematoporfirina usado como fotossensibilizador em Terapia Fotodinâmica (TFD). O derivado de hematoporfirina é composto por uma mistura de monômeros, dímeros e oligômeros mas as estruturas químicas destes componentes ainda não estão bem caracterizadas. Empregou-se extração em fase sólida para limpeza e fracionamento dos componentes do Photogem®. As três frações principais foram separadas de acordo com suas respectivas polaridades. Em seguida, testes preliminares de atividade biológica foram desenvolvidos com as respectivas frações do Photogem® em células tumorais (HEp-2) e normais (VERO). A fração 2 apresentou citotoxicidade cinco vezes maior do que o Photogem® tanto nas células tumorais quanto nas células normais. Os componentes majoritários do Photogem® foram isolados por meio de cromatografia líquida de alta eficiência (CLAE) e analisados por espectroscopia eletrônica na região do ultravioleta-visível (UV-Vis) e por espectrometria de massa. O componente que corresponde à fração 1 pode ser atribuído à hematoporfirina IX em função do seu espectro de absorção eletrônica (banda de Soret em 396nm e bandas Q em 497, 531, 568 e 619nm), seu coeficiente de absortividade molar ((1,30±0,07)x105 cm-1 mol-1 L) e seu espectro de massa (íon pseudomolecular em m/Z 599Da). É importante mencionar que este trabalho proporcionou a elaboração de um procedimento acessível à maioria dos laboratórios brasileiros no que se refere a purificação, separação, isolamento e caracterização das frações constituintes do Photogem®, sendo uma contribuição relevante para o avanço da TFD no Brasil. As etapas da metodologia completa são comentadas e discutidas em detalhes. / The present work focused the isolation, the biological activity analysis and the characterization of the major components of Photogem® through High Performance Liquid Chromatography (HPLC), UV-Vis electronic absorption spectroscopy (UV-Vis) and mass spectrometry. Photogem® is a hematoporphyrin derivative used as photosensitizer in Photodynamic Therapy (PDT). The hematoporphyrin derivative is composed by a mixture of monomers, dimers and oligomers but the chemical structures of these components are not well-characterized yet. It was employed solid phase extraction to clean-up and fractionation of the Photogem® components. Three principal fractions was separated as function of their respective polarities. Subsequently, initial biological tests were developed with the respective Photogem® fractions in tumoral (Hep-2) and normal (VERO) cells. Fraction 2 presented citotoxicity approximately 5-fold higher than Photogem® as in tumoral as in normal cells. The major components of Photogem® were isolated by High Performance Liquid Chromatography (HPLC) and analyzed by UV-Vis electronic absorption spectroscopy (UV-Vis) and mass spectrometry. The component that corresponds to fraction 1 can be assigned to hematoporphyrin IX as function of its electronic absorption spectrum (Soret band in 396nm and Q bands in 497, 531, 568 and 619nm), its molar absortivity coefficient ((1,30?0,07)x105 cm-1 mol- 1 L) and its mass spectrum (pseudomolecular ion in m/z 599Da). It is important to mention that this work provided the elaboration of an accessible procedure to the most of the brazilian laboratories regarding purification, separation, isolation and characterization of the constituent fractions of Photogem®, being a relevant contribution to the advancement of PDT in Brazil. The steps of the complete methodology are commented and discussed in details.