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As raízes de Pfaffia paniculata (Ginseng brasileiro) são comercialmente encontradas como cápsulas contendo as raízes pulverizadas, misturadas ou não ao extrato etanólico destas raízes. Estas raízes são popularmente recomendadas para vários propósitos, e têm sido utilizadas na terapia contra o câncer pela medicina popular. Os principais componentes encontrados nestas raízes já isolados incluem o ácido pfáffico e os pfaffosideos A, C, D, E e F; estes componentes inibiram o crescimento de células do melanoma B-16, demonstrado em um outro estudo. O objetivo deste estudo foi investigar os efeitos dos extratos etanólico, butanólico ou aquoso das raízes de Pfaffia paniculata sobre o crescimento das linhagens tumorais de células mamárias humanas MCF-7 e SKBR-3, utilizando método colorimétrico (cristal violeta) e quantificação das células que incorporaram bromodeoxiuridina (BrdU). A coloração por laranja de acridina/brometo de etídeo foi utilizada para avaliar morte celular, e as alterações subcelulares foram avaliadas por microscopia eletrônica. O extrato butanolico apresentou efeitos citotóxicos nas linhagens MCF-7 e SKBR-3. Morte celular foi observada pelo tratamento com o extrato butanólico por 1 h; alterações morfológicas foram observadas com 500µg/mL deste extrato, após 24 h de tratamento. Após o tratamento por 48 h com o extrato butanólico nesta mesma concentração, foi observado degeneração de componentes citoplasmáticos e alterações morfológicas sugestivas de morte celular. O tratamento com 1000µg/mL deste extrato levou a profundas alterações citoplasmáticas e nucleares, incompatíveis com a viabilidade celular. O tratamento com o extrato etanólico não causou efeitos significantes no crescimento das células MCF-7 e SKBR-3; o extrato aquoso, por outro lado, estimulou o crescimento das células MCF-7, após o tratamento por 1 h. Estes resultados indicam efeitos citotóxicos exercidos pelo extrato butanólico das raízes de Pfaffia paniculata em linhagens celulares de tumor de mama humano / The roots of Pfaffia paniculata (Brazilian ginseng) are commercially available as capsules containing the powdered roots, mixed or not with the ethanolic extract of the roots. These roots have been populary recommended for many purposes, and have also been used on cancer therapy by folk medicine. The main components found in these roots that have already been isolated include pfaffic acid and pfaffosides A, C, D, E and F; these components inhibited the growth of melanoma B-16 cells, as shown in another study. The aim of this study was to investigate the effects of ethanolic, butanolic or aqueous extract of the roots of Pfaffia paniculata on the growth of human breast tumor cell lines MCF-7 and SKBR-3, using a colorimetric method (crystal violet) and quantification of bromodeoxiuridine (BrdU) positive cells. Acridine orange/ethidium bromide staining was utilized to evaluate cell death, and subcellular alteration was evaluated by electron microscopy. The butanolic extract presented cytotoxic effects in MCF-7 and SKBR-3 cell lines. Cell death was observed following treatment with the butanolic extract for 1 h; morphologic alterations were observed with 500µg/mL of this extract, after 24 and 48 h of treatment. Following treatment for 48 h with the butanolic extract with this same concentration, degeneration of cytoplasmic components and morphological alterations suggestive of cell death were observed. Treatment with 1000µg/mL of this extract lead to profound cytoplasmic and nuclear alterations, incompatible with cell viability. The treatment with ethanolic extract didn´t lead to significant effects on the growth of MCF-7 and SKBR-3 cells; the aqueous extract, on the other hand, stimulated the growth of MCF-7 cells, following treatment for 1 hour. These results point to cytotoxic effects exerted by the butanolic extract from the roots of Pfaffia paniculata on human breast tumor cell lines

O uso de plantas medicinais no tratamento de doenças tem aumentado de forma significativa nos últimos anos, bem como as pesquisas sobre a caracterização química destas plantas. Neste estudo, foram determinados os constituintes inorgânicos presentes nas folhas e nos extratos de três espécies da planta do gênero Casearia (C. sylvestris, C. decandra e C. obliqua) coletadas em regiões distintas da Mata Atlântica, SP. As composições elementares dos solos em que estas plantas foram cultivadas também foram determinadas. Tradicionalmente, estas plantas são utilizadas devido às suas propriedades antiinflamatórias, antiácidas, anti-sépticas e cicatrizantes. Além disso, as atividades antiulcerogênicas e antitumorais do gênero Casearia, e sua capacidade de neutralizar venenos de serpentes e abelhas, também tem sido comprovadas cientificamente. A metodologia analítica empregada foi a análise por ativação com nêutrons. Irradiações longas e curtas das amostras e dos padrões foram realizadas utilizando fluxos de nêutrons do reator de pesquisas IEA-R1 do IPEN/CNEN-SP. Nas folhas, o elemento K foi encontrado em níveis de porcentagens, os elementos Ca, Cl, Mg e Na na ordem de mg g-1, Br, Fe, Mn, Rb e Zn em μg g-1 e os elementos As, Co, Cr, Cs, La, Sb, Sc e Se em teores mais baixos, da ordem de ng g-1. Os resultados obtidos nas análises dos extratos indicaram que os mesmos elementos determinados nas folhas de Casearia foram também encontrados em seus respectivos extratos. A comparação das composições elementares das folhas de Casearia sylvestris coletadas em três regiões diferentes da Mata Atlântica mostrou que as concentrações dos elementos encontrados nas plantas dependem da composição dos solos em que elas foram cultivadas. Por outro lado, espécies diferentes do gênero Casearia cultivadas em uma mesma região apresentaram composições elementares similares. Estes resultados mostram a importância da realização de estudos que avaliem as atividades farmacológicas de Casearias cultivadas em diferentes tipos de solos. A qualidade analítica dos resultados foi avaliada pela análise dos materiais de referência certificados NIST 1573a Tomato Leaves, NIST 1515 Apple Leaves, INCT-TL-1 Tea Leaves, IAEA-Soil-7 e USGS W1. Os desvios padrão relativos e os erros relativos obtidos nestas análises indicaram boa precisão e exatidão dos resultados. / The use of medicinal plants in the treatment of diseases has increased significantly in the last years, as has research concerning chemical characterization of these plants. In this study, inorganic constituents were determined in leaves and in extracts from three medicinal plant species of the Casearia genus (C. sylvestris, C. decandra and C. obliqua) collected in distinct regions of the Atlantic Forest, SP. The elemental compositions of the soils in which these plants were grown were also determined. Traditionally, these plants are used due to their antiinflamatory, antiacid, antiseptic and cicatrizing properties. The antiulcer and the antitumor activities of the Casearia genus and its capacity to neutralize snake and bee venoms, have also been scientifically confirmed. The analytical methodology used was neutron activation analysis. Long and short irradiation periods of the samples and the standards were carried out at IPEN’s IEA-R1 nuclear research reactor. In the leaf K was found at the percentage levels, Ca, Cl, Mg and Na at mg g-1 levels and the elements Br, Fe, Mn, Rb and Zn at the μg g-1 levels. As, Co, Cr, Cs, La, Sb, Sc and Se at the ng g-1 levels. Results obtained in the extracts indicated that the same elements present in the leaves are also found in their extracts. The comparison between the inorganic composition of Casearia sylvestris leaves collected from three different regions of the Atlantic Forest showed that the elemental concentrations in the plants leaves varied depending on the place where they were grown. Different Casearia species cultivated in a same region presented similar elemental compositions. Based on these findings it can be concluded that the studies about the pharmacological effect of Casearia genus plants grown in different types of soil are of great importance. The quality of the obtained results was assured by the analyses of the certified reference materials NIST 1573a Tomato Leaves, NIST 1515 Apple Leaves, INCT-TL-1 Tea Leaves, IAEA-Soil-7 and USGS W1. The relative standard deviations and the relative errors obtained in these analyses indicated good precision and accuracy of the results.

Mestrado em Controlo de Qualidade / MSc in Quality Control / Piper betle L., conhecida como bétele, é uma espécie que se desenvolve largamente nos países do Sudeste Asiático, onde as suas folhas são económica e medicinalmente importantes. Para determinar o maior número possível de compostos voláteis e semivoláteis, as folhas foram sujeitas as diferentes processos de extracção, nomeadamente headspace - microextracção em fase sólida (HS-SPME), hidrodestilação e extracção por Soxhlet, e posteriormente analisadas por GC/MS, o que permitiu identificar 50 compostos, distribuídos por várias classes químicas, 23 dos quais foram descritos pela primeira vez. As diferentes técnicas permitiram a extracção de compostos distintos, sendo a HS-SPME aquela com que se obteve o perfil mais completo e com que se determinou maior teor de compostos. Dentro deste processo os melhores resultados foram obtidos utilizando a fibra revestida com divinilbenzeno/polidimetilsiloxano (DVB/PDMS). Considerando que a espécie é vulgarmente mascada, o seu extracto aquoso também foi analisado, tendo sido caracterizados apenas sete compostos, sendo o eugenol o composto principal. A composição em ácidos orgânicos deste extracto foi determinada por HPLC/UV e os ácidos oxálico, aconítico, cítrico, pirúvico, málico, chiquímico, acético e fumárico foram descritos pela primeira vez. O extracto aquoso também revelou capacidade para inibir a acetilcolinesterase de modo dependente da concentração. / Piper betle L., popularly known as “Paan”, is a species widely growing in South East Asia, where its leaves are economically and medicinally important. In order to screen the highest possible number of volatile and semi-volatile components, the leaves were subjected to headspace solid-phase microextraction (HS-SPME), hydrodistillation and Soxhlet extraction, prior to their analysis by GC/MS, which allowed the identification of 50 compounds, distributed by several chemical classes, 23 of them described for the first time. The different techniques lead to distinct compounds' extraction, with HS-SPME extracting highest amounts and providing the most complete profile. Within this procedure, best results were obtaine using Divinylbenzene/Polydimethylsiloxane (DVB/PDMS) fibre. Considering the use of the species as masticator, an aqueous extract was also analysed, in which only seven compounds were characterize, being eugenol the main one. The organic acids composition of this extract was determine by HPLC/UV and oxalic, aconitic, citric, pyruvic, malic, shikimic, acetic and fumaric acids are reported for the first time in this species. The aqueous extract also displayed AChE inhibitory capacity, in a concentration-dependent way.

Thesis ( |b(MSc. (Biochemistry)) -- University of Limpopo, 2017 / Medicinal plants have been used as a key source for medication and they remain to provide new therapeutic remedies to date. Extracts of Olea europaea subspecies africana leaves are used extensively in South Africa to treat various diseases traditionally. The diseases have been noted to be associated with free radicals, bacterial infections, and inflammation. However, there is little information about the antioxidant, antibacterial, and anti-inflammatory activities of the leaves of this plant in literature and the cytotoxicity of the leaf extracts is still a concern. The information about the Isolated compounds is also minimal hence this study was aimed at filling in those gaps in relation to the traditional use of the leaves in southern Africa and subsequently isolating and identifying the active compounds using bioassay-guided fractionation. Preliminary screening of the crude extracts for antioxidant, antibacterial and antiinflammatory activities indicated that the extracts possessed all biological activities. The presence of major phytochemicals in the crude extracts was determined through the use of standard chemical methods and TLC analysis. The colorimetric methods (Folin-Ciocalteau and Aluminum chloride) were used for quantification purposes. TLC-DPPH assay was used to screen antioxidant activities of the crude extracts. The observed activity was quantified using the spectrophotometric method of DPPH and reducing power. The antibacterial properties of the leaf extracts were determined by direct bioautography and the serial broth microdilution assay using E. coli, P. aeruginosa, E. faecalis and S. aureus as test bacteria. Screening of the acetone crude extract for anti-inflammatory activities was done using the LPSstimulated RAW 264.7, cells where the inhibition of ROS generation was studied. MTT assay was used to determine the cytotoxicity effects of the leaves. Isolation of bioactive compounds started with serial exhaustive extraction, followed by column chromatography packed with silica gel. NMR analysis was conducted to identify the isolated compound. The results revealed the presence of tannins, terpenoids, steroids and flavonoids with the total phenolic (99.67 ± 2.52 mg of GAE/g) and tannin content (114.33 ± 9.02 mg of GAE/g) found in high amounts. All crude extracts exhibited antioxidant activities and the antioxidant activity quantified via the DPPH assay demonstrated to xxi have EC50 value of 1.05 ± 0.0071 mg/mL. The reducing capacity was found to be dose-dependent and great significance was seen at concentration 0.5 mg/mL to 1 mg/mL that was about 2/3 of that of L-ascorbic acid (standard) at a similar concentration. Screening of the crude extracts for antibacterial activity revealed that all crude extracts except n-hexane and water extracts, inhibited the growth of the tested bacteria on the previously developed TLC plates. The activity was seen as clear zones on the bioautograms. Serial broth microdilution assay indicated that dichloromethane, acetone and ethanol had average MIC values of 0.30, 0.32 and 0.35 mg/mL against all tested bacteria, respectively. Good anti-inflammatory activity of the crude extract was demonstrated at the highest concentration of 0.90 mg/mL. MTT assay indicated that the crude extract had no adverse cytotoxic effects. This was demonstrated by the LC50 values greater than 20 µg/mL and considered non-cytotoxic according to the National Cancer Institute (NCI). Isolation following the bioassay-guided-fractionation resulted in the selection of acetone extract to isolate the bioactive compounds from as it demonstrated good antioxidant and antibacterial activities. Fractionation of the compound by column chromatography yielded three combinations (pools) of fractions and of the three from which only pool 1 was considered for further fractionation. NMR spectra information identified the isolated compound as a mixture of ursolic acid (minor) and oleanolic acid (major). This compound had antibacterial and anti-inflammatory activities and no cytotoxic effects. The leaves of Olea europaea subspecies africana have been proven to possess antioxidant, antibacterial and anti-inflammatory properties. Evaluation of the biological activities of the crude extracts was to validate the use of the leaves traditionally to treat free radical and bacterial-related diseases and potential drug that are safe and has less side effects may be produced from the leaves. / National Research Foundation (NRF)

Thesis (M.Sc.) --University of Limpopo, 2008 / Senna species, a member of the Fabaceae family (subfamily Caesalpinaceae), is widely used traditionally to treat a number of disease conditions such as sexually transmitted diseases and some forms of intestinal complications. In this study the roots of Senna species, collected from Zebediela region of the Limpopo province (R.S.A), were ground to a fine powder and extracted with acetone by cold/shaking extraction method. The phytochemical composition of the extract was then determined by thin layer chromatography (TLC). The chromatograms were visualised with vanillin-sulphuric acid and p-anisaldehyde reagents. The total phenolic content of the extract was determined by Folin-Ciocalteu method and expressed as TAE/g of dry plant material. The extract was assayed for the in vitro anticancer activity using Jurkat T cells. The antioxidant activity was evaluated using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) assay and the antibacterial activity determined by both bioautographic and the microtiter plate methods. The acetone extract of the roots of Senna species inhibited the growth of Jurkat T cells in a dose- and time-dependent manner. The extract was shown to possess free radical scavenging activity and antibacterial activity against Pseudomonas aeruginosa, Enterococcus faecalis, Escherichia coli and Staphylococcus aureus with MIC values of 0.16, 0.078, 0.078 and 0.16 mg/ml, respectively. A compound with free radical scavenging activity was isolated from the acetone extract of the roots of Senna species through bioassay-guided fractionation. The isolated compound was identified as 1, 3-diphenol-2-propen-1-one. Thus, the study has systematically shown the biological activity of the roots of Senna species and the isolation and identification of the bioactive compound.

Masters of Science / Plant genome analysis is currently experiencing a boost due to reduced costs associated with the development of next generation sequencing technologies. Knowledge on genetic background can be applied to guide targeted plant selection and breeding, and to facilitate natural product discovery and biological engineering. In medicinal plants, secondary metabolites are of particular interest because they often represent the main active ingredients associated with health-promoting qualities. Plant polyphenols are a highly diverse family of aromatic secondary metabolites that act as antimicrobial agents, UV protectants, and insect or herbivore repellents. Most of the genome mining tools developed to understand genetic materials have very seldom addressed secondary metabolite genes and biosynthesis pathways. Little significant research has been conducted to study key enzyme factors that can predict a class of secondary metabolite genes from polyketide synthases. The objectives of this study were twofold: Primarily, it aimed to identify the biological properties of secondary metabolite genes and the selection of a specific gene, naringenin-chalcone synthase or chalcone synthase (CHS). The study hypothesized that data science approaches in mining biological data, particularly secondary metabolite genes, would enable the compulsory disclosure of some aspects of secondary metabolite (SM). Secondarily, the aim was to propose a proof of concept for classifying or predicting plant genes involved in polyphenol biosynthesis from data science techniques and convey these techniques in computational analysis through machine learning algorithms and mathematical and statistical approaches. Three specific challenges experienced while analysing secondary metabolite datasets were: 1) class imbalance, which refers to lack of proportionality among protein sequence classes; 2) high dimensionality, which alludes to a phenomenon feature space that arises when analysing bioinformatics datasets; and 3) the difference in protein sequences lengths, which alludes to a phenomenon that protein sequences have different lengths. Considering these inherent issues, developing precise classification models and statistical models proves a challenge. Therefore, the prerequisite for effective SM plant gene mining is dedicated data science techniques that can collect, prepare and analyse SM genes.

Thesis (M. Sc.(Agric))(Horticulture)-University of Pretoria, 2005. / Includes bibliographical references. Available on the Internet via the World Wide Web.