Dissertations / Theses on the topic 'Biological activity of chamomile extracts'

У оквиру ове докторске дисертације изведено јеиспитивање различитих екстракционих поступака заизоловање апигенина из цвета камилице, као и евалуацијабиолошке активности добијених екстраката. Полазнибиљни материјал сачињавале су две групе латицакамилице: ферментисане и неферментисане (нативне).Екстракција ферментисаних цветова је извођена применомултразвучне екстракције користећи етанол као екстрагенс,а добијени екстракти су се одликовали изузетно високимсадржајем апигенина. Оптимизација екстракције је билаизведена применом методе одзивне површине. Применомелектрон-спин резонанце испитана је антирадикалскаактивност екстраката. Додатно, фармаколошка вредностдобијених екстраката је потврђена и одређивањем њиховогантимикробног и антипролиферативног потенцијала.Нативни цветови камилице су екстраховани применомразличитих екстаркционих техника: микроталасне,ултразвучне, Soxhlet екстракције као и екстракцијесубкритичном водом. Eкстрaкција водом у субкритичномстању се показала супериорнијом у односу на све осталетехнике у погледу садржаја укупних фенола и флавоноида.У циљу добијања екстраката са максималним садржајемапигенина изведена је оптимизација овог екстракционогпроцеса. Изоловање чистог апигенина је изведено изекстракта добијеног под оптималним екстракцијомусловима (однос дрога:растварач 1:30, брзина мешања 3 Hz,притисак 45 bar, температура 115&ordm;C, време 30 мин,концентрација модификатора 0,001 М) применом поступкаколонске хроматографије на стубу полиамида. Хемијскипрофил као и садржај појединачних полифенолнихкомпонената у екстрактима добијеним на различитимпритисцима, температурама и уз присуство модификатораразличитих концентрација одређен је применом UHPLCDAD-HESI-MS/MS. У свим анализираним екстрактимадетектован је велики број полифенолних компонената, докје апигенин у свима био доминантно једињење. Садржајапигенина у екстракту добијеном под оптималнимекстракционим условима је износио 1.700,34 mg/kg.Применом седам различитих тестова извршена јеевалуација антиоксидативног и антирадикалскогпотенцијала екстраката. Антимикробни потенцијалекстраката је одређен за осам различитих микробнихлинија. in vitro тестовима испитана је способностинхибиције &alpha;-амилазе, &alpha;-глукозидазе и тирозиназе.Деловањем на раст три хистолошки различите ћелијскелиније, испитана је антипролиферативна активностекстраката добијених субкритичном водом.Антимотилитетна активност обе групе екстраката(ферментисаних и неферментисаних цветова) одређена је уin vitro условима.<br>U okviru ove doktorske disertacije izvedeno jeispitivanje različitih ekstrakcionih postupaka zaizolovanje apigenina iz cveta kamilice, kao i evaluacijabiološke aktivnosti dobijenih ekstrakata. Polaznibiljni materijal sačinjavale su dve grupe laticakamilice: fermentisane i nefermentisane (nativne).Ekstrakcija fermentisanih cvetova je izvođena primenomultrazvučne ekstrakcije koristeći etanol kao ekstragens,a dobijeni ekstrakti su se odlikovali izuzetno visokimsadržajem apigenina. Optimizacija ekstrakcije je bilaizvedena primenom metode odzivne površine. Primenomelektron-spin rezonance ispitana je antiradikalskaaktivnost ekstrakata. Dodatno, farmakološka vrednostdobijenih ekstrakata je potvrđena i određivanjem njihovogantimikrobnog i antiproliferativnog potencijala.Nativni cvetovi kamilice su ekstrahovani primenomrazličitih ekstarkcionih tehnika: mikrotalasne,ultrazvučne, Soxhlet ekstrakcije kao i ekstrakcijesubkritičnom vodom. Ekstrakcija vodom u subkritičnomstanju se pokazala superiornijom u odnosu na sve ostaletehnike u pogledu sadržaja ukupnih fenola i flavonoida.U cilju dobijanja ekstrakata sa maksimalnim sadržajemapigenina izvedena je optimizacija ovog ekstrakcionogprocesa. Izolovanje čistog apigenina je izvedeno izekstrakta dobijenog pod optimalnim ekstrakcijomuslovima (odnos droga:rastvarač 1:30, brzina mešanja 3 Hz,pritisak 45 bar, temperatura 115&ordm;C, vreme 30 min,koncentracija modifikatora 0,001 M) primenom postupkakolonske hromatografije na stubu poliamida. Hemijskiprofil kao i sadržaj pojedinačnih polifenolnihkomponenata u ekstraktima dobijenim na različitimpritiscima, temperaturama i uz prisustvo modifikatorarazličitih koncentracija određen je primenom UHPLCDAD-HESI-MS/MS. U svim analiziranim ekstraktimadetektovan je veliki broj polifenolnih komponenata, dokje apigenin u svima bio dominantno jedinjenje. Sadržajapigenina u ekstraktu dobijenom pod optimalnimekstrakcionim uslovima je iznosio 1.700,34 mg/kg.Primenom sedam različitih testova izvršena jeevaluacija antioksidativnog i antiradikalskogpotencijala ekstrakata. Antimikrobni potencijalekstrakata je određen za osam različitih mikrobnihlinija. in vitro testovima ispitana je sposobnostinhibicije &alpha;-amilaze, &alpha;-glukozidaze i tirozinaze.Delovanjem na rast tri histološki različite ćelijskelinije, ispitana je antiproliferativna aktivnostekstrakata dobijenih subkritičnom vodom.Antimotilitetna aktivnost obe grupe ekstrakata(fermentisanih i nefermentisanih cvetova) određena je uin vitro uslovima.<br>In the frame of this thesis different extraction approaches forapigenin isolation from chamomile ligulate flowers wereexamined and biological activity of obtained extracts wasevaluated. Starting plant samples included fermented andnonfermented (native) flowers.Extraction of fermented flowers was performed by usingultrasound-assisted extraction with ethanol. The concentrationof apigenin was high in obtained extracts. Optimization of theextraction procedures was performed by response surfacemethodology. Antiradical activity of observed extracts wasexamined by electron-spin resonance spectroscopy.Furthermore, pharmacological potential of obtained extractswas confirmed by determining their antimicrobial andantiproliferative activity.Native chamomile flowers were extracted by differentextraction techniques: microwave, ultrasound, Soxhlet andsubcritical water extraction. Subcritical water extractionshowed to be superior in comparison to other applied techniquesin respect to total phenols and flavonoids content. Optimizationof the subcritical water extraction was directed to maximizationof apigenin content. Isolation of pure apigenin from extractsobtained under optimal extraction conditions (sample-tosolventratio 1:30, agitation rate 3 Hz, temperature 115&ordm;C,pressure 45 bar, extraction time 30 min) was performed bypreparative chromatography. Chemical profiles and content ofindividual polyphenolic components in extracts obtained atdifferent pressures, temperatures, and with differentconcentrations of a modifier was determined by UHPLC-DADHESI-MS/MS. In all analyzed extracts the great number ofpolyphenolic components was detected while apigenin was thedominant compound in all extracts. Content of apigenin in theextract obtained under optimal extraction condition was1,700.34 mg/kg. Antioxidant and antiradical potential ofextracts was evaluated according to different mechanisms.Antimicrobial potential of extracts was determined against eightdifferent microbial strains. Ability of extracts to inhibit &alpha;-amylase, &alpha;-glucosidase and tyrosinase was determined by invitro assays. Antiproliferative activity of subcritical waterextracts was defined by testing their influence on the growth ofthree histologically different cell lines.Anti-intestinal motility activity of both group of extracts (nativeand fermented) was determined by in vivo experiments.

Doutoramento em Engenharia Alimentar - Instituto Superior de Agronomia<br>The aim of this work was to develop methodologies for micropropagation and extraction of bioactive compounds from both wild gorse (Pterospartum tridentatum L.) and micropropagated gorse explants, as a means of valorisation of this species. Micropropagation was very effective on large scale propagation, having multiplication rates ranging from 1.6 to 43.9; number of shoots ranging from 1.5 to 10.4 and rooting above 96.7%. The extraction yield of aqueous extraction varied from 10.5 to 19.4 and 12.2 to 30.1% (w/w) for wild gorse and micropropagated gorse explants, respectively. Aqueous extracts showed high levels of total phenols, from 203.4 to 369.2 and 104.1 to 121.3 mg gallic acid eq g-1 ms, and FRAP antioxidant activity: 256-856 and 160-260 mmolFe2+ 100g-1 ms; DPPH: 1433-1551 and 1503-1509 mM Trolox 100g-1 ms for wild gorse and micropropagated gorse explants, respectively. Extracts showed no antifungal activity. However, these extracts showed inhibition of Bacillus subtilis, Pseudomonas aeruginosa and Staphylococcus aureus. Chitosan bioactive coatings with gorse extracts improved cherry postharvest preservation ability. The results allow topropose these methodologies to obtain extracts with bioactive properties from micropropagated material, without depleting natural resources

PhD (Department of Biotechnology, Faculty of Applied and Computer Sciences), Vaal Universtiy of Technology<br>The genus Tulbaghia has been used in traditional medicine to treat various ailments such as fever, earache, tuberculosis and esophageal cancer. However, there is limited scientific evidence to support its use. Therefore the objectives of this study were to perform phytochemical analysis, investigate the antioxidant, antimicrobial, anticancer, immunomodulatory activities and toxicity of crude acetone and water extracts from selected Tulbaghia species. Standard methods were used for preliminary phytochemical analysis. The total phenolic content of the plant extracts was determined using the folin ciocalteu method whereas the total flavonoids were determined by using the aluminium chloride colorimetric method. DPPH and ABTS assays were used to evaluate the antioxidant activity. The antimicrobial activity was assessed by agar well diffusion, microtiter dilution and time kill assays. For anticancer studies, the antiproliferative activity of the extracts was evaluated using the MTT assay on Hkesc-1 and KB cells. Morphological changes of the cancer cells treated with extracts were examined using light microscopy. Induction of apoptosis was assessed using fluorescence microscopy and acridine orange/ethidium bromide staining. Flow cytometry analysis was conducted to examine the multicaspase activity and cell cycle arrest. For immunomodulatory activity, the Greiss reagent and Luminex cytokine assays were used to determine the effect of the extracts on NO production and the concentration of the cytokines in the treated cells, respectively. Toxicity of selected Tulbaghia species was examined by investigating the effect of the extracts on the metabolic activity and cell membrane integrity on the treated RAW264.7 cells using the MTT and LDH assays, respectively. The zebrafish assay was used to evaluate the embryotoxicity and teratogenic effects of crude acetone and water extracts of T. violacea at 24 h intervals for 96 h post fertilisation (hpf). The percentage mortality, hatchability and heart rate were examined. Phytochemical screening of eight Tulbaghia species demonstrated the presence of flavonoids, glycosides, tannins, terpenoids, saponins and steroids. The amount of total phenol and flavonoid content varied in different plant extracts ranging from 4.50 to 11.10 milligrams gallic acid equivalent per gram (mg GAE/g) of fresh material and 3.04 to 9.65 milligrams quercetin equivalent per gram (mg QE/g) of fresh material respectively. The IC50 values based on DPPH and ABTS for T. alliacea (0.06 and 0.06 mg/mL) and T. violacea (0.08 and 0.03 mg/mL) were generally lower showing potential antioxidant activities. For antimicrobial activity, the acetone extracts of T. acutiloba, T. alliacea, T. leucantha, T. ludwigiana, T. natalensis and T. simmleri showed moderate antimicrobial activity against all test organisms while the water extracts showed moderate to no activity. One species, T. cernua, showed poor activity against all the tested microbes. The acetone and water extracts of T. violacea showed the greatest antibacterial and antifungal activity against all the tested microorganisms with minimum inhibitory concentration ranging from 0.1 mg/mL to 3.13 mg/mL. The acetone extracts of T. violacea also exhibited both bacteriostatic/fungistatic and bactericidal/fungicidal activity depending on the incubation time and concentration of the extract. The bactericidal/fungicidal activity was observed at x2 MIC. The results for anticancer activity showed that treatment of Hkesc-1 cells with acetone and water crude extracts had anti-proliferative activity with IC50 values of 0.4 mg/mL and 1.625 mg/mL, respectively while KB had 0.2 mg/mL and 1 mg/mL, respectively. Morphological changes such as blebbing, cell shrinkage and rounding were observed in the treated cells suggesting that apoptosis was taking place. AOEB staining showed that the level of apoptosis was dependent on the concentration of the extracts. The activation of multicaspase activity in both Hkesc-1 and KB treated cells was also concentration dependent leading to cell death by apoptosis and the induction of cell cycle arrest at the G2/M phase. Immunomodulatory activity results indicated that cell viability was above 80% when concentrations of 50 µg/mL or less of both acetone and water crude was used. Treatment with the acetone extract had no significant effect (p>0.05) on the LPS induced NO production in RAW264.7 cells except at 50 µg/mL where significant inhibition was observed. The water extract had no significant effect (p>0.05) on NO production at all the concentrations. Treatment of LPS–induced RAW264.7 cells with acetone extract stimulated the production of IL-1α, IL-6 and TNF-α, but had no significant effect (p > 0.05) on IL-1β. On the other hand, treatment with the water extracts stimulated the production of IL-1α, IL-6 but had no significant effect (p>0.05) on TNF-α and IL-1β. Treatment of LPS-induced RAW264.7 cells with the acetone extract had very little stimulatory effect on IL-4, IL-5 and IL-13 and no significant effect on IL-10 whereas for the water extract a significant stimulatory effect was only observed for IL-4 after 48 h of treatment. High concentrations (>10000 pg/mL) of MCP-1, MIP1-α, MIP1-β, MIP-2, GCSF, GM-CSF, RANTES and IP-10 were also observed in acetone and water extract treated RAW264.7 cells. For toxicity studies, acetone and aqueous crude leaf extracts from T. alliacea, T. simmleri, and T. violacea had a significant inhibitory (p<0.05) effect on the RAW264.7 cells after 48h treatment. Acetone extracts from T. alliacea, T. simmleri and T. violacea resulted in IC50 values of 0.48 mg/mL, 0.72 mg/mL and 0.1 mg/mL, respectively. Treatment with water extracts showed minimal toxic effect indicated by higher IC50 values of 0.95 mg/mL, 2.49 mg/mL and 0.3 mg/mL for T. alliacea, T. simmleri and T. violacea, respectively. The LDH release by macrophages after 24 h treatment with acetone extracts was observed to be concentration dependent while treatment with water extracts did not induce LDH release. The zebra fish assay showed a lethal dose (LD50) for the T. violacea acetone crude extract of 20 μg/mL whereas that for water extract was 85 μg/mL. The observed teratogenic effects included scoliosis, edema of the pericardial cavity, retarded yolk resorption, hook-like/bent tail and shorter body length. In conclusion, the results from this study indicate that the extracts from the eight Tulbaghia species examined contain phytochemicals that may have the antioxidant, antimicrobial, anticancer and immunomodulatory properties. Extracts from T. violacea were observed to be the most potent. This study thus supports the use of T. violacea in treating bacterial and fungal infections in traditional medicine. The results of this study also confirm the anticancer potential of T. violacea. The immunomodulatory activity of the acetone and water extracts from T. violacea indicated a dominantly pro-inflammatory activity. Traditional medicine prepared form T. violacea may be of benefit to individuals with weak immune systems. The toxicity of selected Tulbaghia species was observed to be concentration, extract and time dependent. Therefore, traditional medicine prepared from Tulbaghia extracts should be taken with caution preferably in small doses over a short period of time. Future studies will focus on the identification of the bioactive compound(s) responsible for the antimicrobial, anticancer and immunomodulatory activities.

Aromatic plants contain both volatile and non-volatile fractions and the chemical composition of these two fractions may be influenced by seasonal changes. The essential oil and solvent extracts of S. africana-caerulea, S. africana-lutea and S. lanceolata, collected at the same locality throughout the 2005 growing season, were compared in terms of essential oil composition, yields and biological activities. Mostly quantitative, rather than qualitative variationwas observed in the oil composition of each species.Major fluctuations in the composition of S. africana-caerulea oil included limonene (2–33%) and viridiflorol (2–24%). Levels of α-pinene (1–12%), myrcene (2–12%) and α-eusdesmol (trace-13%) fluctuated seasonally in the S. africana-lutea oil. In S. lanceolata, considerable changeswere noted for β-caryophyllene (1–19%), β-caryophyllene oxide (1–21%) and ledol (3–12%). The extract prepared from S. lanceolata harvested inwinterwas more active against Gram-positive bacteria. The S. africana-caerulea extract exhibited the most favourable antiplasmodial activity when harvested in winter (IC50 value: 12 μgml−1), which contrasts with the lowest anti-plasmodial activity of S. lanceolata obtained at the same period (IC50 value: 43 μgml−1). The anti-oxidant activity of the solvent extracts also displayed variation over seasons with thewinter collection of S. africana-lutea yielding the most favourable anti-oxidant activity (IC50 value: 10 μg ml−1). All the solvent extracts prepared from the winter collection exhibited the lowest toxicity (20bIC50 valuesb60 μg ml−1), while the three essential oils obtained from autumn collection were more toxic (0.03bIC50 valuesb0.4 μg ml−1).

The aim of the first project was to evaluate the ability of selected molecules to affect the secondary structure of Abeta(1-42). Such activity is desirable as it may provide new tools to study the role of the Abeta(1-42) conformers in Alzheimer disease. Although this study did not identify molecules able to modulate the conformation of Abeta (1-42), the reported studies have set the fundamental techniques for this project.<br>The goal of the second project was to compare the kavalactone content and the inhibition of human P450 enzymes for commercial and traditional kava extracts. We report that amounts and ratios of kavalactones are similar for solvents that mimic commercial extracts but different from aqueous extract (traditional extract). All extracts tested show similar inhibition of the P450 enzymes suggesting that kava extracts, commercial or traditional, may lead to herb-drug interactions. These results also suggest that because of their higher concentration of kavalactones, commercial kava extracts are more likely to cause adverse drug reactions via inhibition of drug metabolism.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior<br>Socratea exorrhiza (Mart.) H. Wendland é conhecida popularmente como paxiuba. Esta Arecaceae ocorre naturalmente na região amazônica. É muito utilizada por caboclos e índios de Roraima, porém pouco estudada quanto ao perfil químico e biológico. Neste trabalho investigou-se o potencial bioativo dos extratos da parte externa e interna da raiz e do broto da raiz de S. exorrhiza, mediante testes de atividade citotóxica (Artemia salina Laech), antioxidante (DPPH) e antimicrobiana contra as bactérias (Staphylococcus aureus ATCC 6538, Escherichia coli ATCC 25922, Proteus mirabilis ATCC 7002) e a levedura Candida albicans ATCC18804. Dos extratos mais ativos determinaram-se os teores de fenólicos e flavonóides por espectrofotometria. O processo de extração foi a quente em sistema de Soxhlet, usando solventes orgânicos de polaridades crescentes (hexano, clorofórmio, acetato de etila e metanol). Os resultados mostraram que os extratos acetato de etila da parte externa da raiz e do broto da raiz destacaram se nos testes de atividade citotóxica e antioxidante, sendo o do broto da raiz (DL50 473 e IC50 24,03 &#956;g/mL) o mais ativo para os dois testes. A ação positiva desses extratos nas atividades biológicas pode estar relacionada à presença de flavonoides evidenciada pelo elevado teor encontrado (76,8% e 64,66%, respectivamente). Os extratos metanólicos da parte externa da raiz e do broto da raiz apresentaram alto poder inibitório frente à levedura C. albicans (halos de18 e 25 mm) respectivamente. Todos os extratos do broto da raiz apresentaram ação inibitória moderada contra o crescimento das bactérias Gram-negativas E. coli e P. mirabilis com halos que variaram entre 10 e 17 mm, sendo que o extrato metanólico do broto da raiz obteve alto poder inibitório no crescimento de P. mirabilis (halo 22 mm). A ação positiva dos extratos da parte externa da raiz e do broto da raiz contra bactérias Gram-negativas corrobora com o conhecimento tradicional, que preconiza a utilização das raízes de S. exorrhiza na medicina popular para o tratamento de doenças sexualmente transmissíveis, o que torna ainda maior o valor dos resultados obtidos. Outro dado importante obtido com trabalho é o novo registro de ocorrência de S. exorrhiza para o estado de Roraima. Os resultados aqui apresentados são inéditos e importantes na ampliação do conhecimento sobre esta palmeira, estimulando a continuidade de estudos mais aprofundados.<br>Socratea exorrhiza (Mart.) H. Wendland is popularly known as paxiuba. This Arecaceae occurs naturally in the rain forest. It is widely used by the locals and the native Indians of Roraima, however it is not as studied as in regards to its biological and chemical profile. In this work, it was investigated that the extracts of the bioactive potential of the outer and inner parts of the root and of the sprout from the root of S. exorrhiza, by cytotoxic activity tests (Artemia salina Laech), antioxidant (DPPH) and antimicrobial against bacteria (Staphylococcus aureus ATCC 6538, Escherichia coli ATCC 25922, Proteus mirabilis ATCC 7002) and with Candida albicans yeast ATCC18804. Among the most active extracts were determined the levels of phenolics and flavonoids by spectrophotometry method. The extraction process was heat by the Soxhlet method, using organic solvents of increasing polarity (hexane, chloroform, ethyl acetate and methanolic). extracts of the outer part of the root and the sprout from the root stood out on the cytotoxic activity and the antioxidant tests, being the sprout from the root (DL50 473 and IC50 24,03 &#956;g/mL) the most active of the two tests. The positive Action of these two extracts on the biological activities can be related to the presence of flavonoids evidenced by the high content found on the of the outer part of the root extracts 76,8% and sprout from the root 64,66%. The methanolic extracts of the outer part of the root and the sprout from the root demonstrated high inhibitory power before the C. albicans yeast (halos of 18 and 25 mm) respectively. All the sprouts from the root extracts presented moderate inhibitory action against the growth of the Gram-negative E. coli and P. mirabilis bacteria with halos that varied between 10 and 17mm, and the methanolic extracts sprout from the root obtained high inhibitory power on the growth of P. mirabilis (halo 22 mm). The extracts&#8223; positive action of the outer part of the root and the sprout from the root against Gram-negative bacteria collaborates with the traditional knowledge that advocates the use of the S. exorrhiza roots in popular medicine for the sexually transmitted diseases (STD), which makes even higher the values of the results with these extracts. Another important finding in this work is the new record of occurrence of S. exorrhiza for the state of Roraima.The data herein presented is new and important in advancing knowledge about this palm tree, stimulating the continuity of deeper studies.

Made available in DSpace on 2016-06-02T19:30:13Z (GMT). No. of bitstreams: 1 6774.pdf: 4696015 bytes, checksum: 084c02944eff3cb2f25b1db2ce63280b (MD5) Previous issue date: 2015-03-31<br>Financiadora de Estudos e Projetos<br>Plants species from Brazilian savanna often have defense mechanisms in response to the low nutrient content of the soil, the light intensity, the competition for resources and drought, pronounced by deep water deficit, to which they are subjected. This area is a potential source of bioactive compounds that still little explored and potential phytochemical studies of these species are important to this preservation. Biological invasions are one of the main factors threatening the Brazilian savanna biodiversity. Many plants are initially introduced as ornamental species and subsequently become invasive. Invasive plants are generally more competitive than the other species and tolerate unfavorable conditions. This advantage can be explained according to the "Novel weapons hypothesis". It proposes that invasive plants have secondary metabolites that act as allelochemicals and present new challenges to native species. Thus, secondary metabolites of invasive species in the Brazilian savanna are also an important area of phytochemistry being investigated. Therefore, we hypothesized that native species from Brazilian savanna, Myrsine umbellata and Myrsine coriacea, and the alien specie, Tithonia diversifolia, produces secondary metabolites that have different biological activities, including insecticidal, antifungal and phytotoxic acitivities, favoring their establishment in this environment and have economic importance for agriculture and pharmaceutic industry. In the Chapter 1, the biological activity of leaf extracts of M. umbellata and M. coriacea, was evaluated. The in vitro antifungal effect of leaf extracts of these two species was evaluated against Pythium aphanidermatum, Fusarium oxysporum and Colletotrichum acutatum, three important crop pathogens. We also evaluated the insecticide potential of these extracts against Spodoptera frugiperda, known as fall armyworn, an important pest that causes damages to many crops. Finally, the total polyphenols and antioxidant activity of the leaf extracts of these two species was quantified by the DPPH method. In the Charpter 2 we evaluated the phytotoxic potential of leaf, stem and root extracts from T. diversifolia. Bioassays of wheat elongation coleoptile seed germination and seedling growth are performed. The Ethyl Acetate leaf extract was the most active extract and was selected to continue the isolation studies and the identification of compounds. The chromatographic separation of this extract allowed us the isolation of 12 sesquiterpene lactones (STSs), 1 secosesquiterpeno, and 1 diterpene. Two STSs are reported for the first time in this study, 8&#946;-O-(2-methylbutyroyl) and tirotundin 8&#946;-O-(isovaleroyl)tirotundin. The high phytotoxic activity of the extracts and compounds, especially the majority, 1&#946;-methoxydiversifolin, tagitinin A and tagitinin C, indicates that allelopathy may be responsible for their establishment in this habitat. In the Chapter 3, the T. diversifolia Ethyl Acetate fractions were tested for insecticidal activity against S. frugiperda and it was observed a significant reduction in the pupal weight of the caterpillar. This may be an indicative of the presence of toxic substances to this insect in these fractions, however, this reduction in the pupal weight is not associated with the majority compounds tested. Thus, we conclude that the studied plant species have secondary metabolites with promising biological activities and economic importance for agriculture and pharmaceutical industry.<br>Espécies vegetais provenientes do Cerrado costumam apresentar mecanismos de defesa em resposta ao baixo teor de nutrientes do solo à que estão submetidas, a intensa luminosidade, a maior competição por recursos e ao longo período de estiagem, marcada por profundo déficit hídrico. Este domínio é uma fonte potencial de compostos bioativos ainda pouco explorada e estudos do potencial fitoquímico destas espécies são importantes para a preservação da sua biodiversidade. Entre os principais fatores que ameaçam a biodiversidade do Cerrado, está a invasão biológica. As plantas invasoras são geralmente mais competitivas que as demais espécies e suportam condições adversas. Essa vantagem pode ser explicada segundo a hipótese das novas armas , que afirma que as plantas invasoras possuem metabólitos secundários que agem como aleloquímicos e apresentam novos desafios para as espécies nativas. Dessa forma, metabólitos secundários de espécies invasoras do Cerrado também são uma importante área da fitoquímica a ser investigada. Sendo assim, foi proposta a hipótese de que espécies nativas, Myrsine umbellata e Myrsine coriacea e uma invasora do cerrado, Tithonia diversifolia, produzem metabólitos secundários em resposta ao estresse à que estão submetidas e que estes apresentam diferentes funções biológicas, incluindo, atividade antifúngica, antioxidante, inseticida e fitotóxica, que favoreçam seu estabelecimento nesse ambiente e apresentam importância econômica para uso na agricultura e na indústria farmacêutica. Para isso, no primeiro capítulo, foi avaliada a atividade biológica de extratos foliares de M. umbellata e M. coriacea. Foi avaliado o efeito antifúngico in vitro de extratos foliares destas espécies sobre Pythium aphanidermatum, Fusarium oxysporum e Colletotrichum acutatum, importantes patógenos de culturas agrícolas. Também foi avaliado o potencial inseticida destes extratos sobre Spodoptera frugiperda, conhecida como lagarta-do-cartucho do milho, importante praga causadora de danos a esta cultura. E por último, foi determinado a concentração de fenóis totais e atividade antioxidante dos extratos foliares destas duas espécies pelo método de redução do radical DPPH (2,2-difenil-1-picrilidrazil). No segundo capítulo foi avaliado o potencial fitotóxico de extratos de folha, caule e raiz de T. diversifolia. Foram realizados bioensaios com coleóptilos de trigo, germinação de sementes e crescimento de plântulas. O extrato foliar preparado com Acetato de Etila foi selecionado para continuar os estudos de isolamento e identificação de compostos por apresentar melhor atividade. A separação cromatográfica deste extrato nos permitiu isolar 12 lactonas sesquiterpênicas (STS), 1 secosesquiterpeno, e um diterpeno. Duas STSs estão sendo relatadas pela primeira vez neste trabalho, 8&#946;-O-(2-metilbutiroil)tirotundina e 8&#946;-O- (isovaleroil)tirotundina. A alta atividade fitotóxica desta espécie, bem como dos compostos avaliados e, principalmente, os compostos majoritários 1&#946;-metoxidiversifolina, tagitinina A e tagitinina C, indica que a alelopatia pode ser uma das formas de seu estabelecimento nesse ambiente. No capítulo 3 foram testados a atividade inseticida de frações do extrato Acetato de Etila de T. diversifolia e seus compostos majoritários contra S. frugiperda e foi observado uma redução significativa no peso pupal da lagarta. A redução da massa pupal pode ser o indicativo da presença de substâncias tóxicas ao inseto nessas frações, no entanto, essa redução não está associada à presença dos compostos majoritários testados. Dessa forma, concluímos que as espécies vegetais estudadas, possuem metabólitos secundários com atividades biológicas promissoras e importância econômica para serem utilizados na agricultura e na indústria farmacêutica.

The aim of this research was to investigate the phytochemistry of two species Sanicula europaea and Teucrium davaeanum which are traditionally used in treatment of wounds. Four compounds were isolated from the 80% methanolic extract of S. europaea; bis-(2-ethylhexyl) phthalate (1), palmitic acid (2), rosmarinic acid (3), saniculoside N (4). Compounds 1 and 2 were isolated for the first time from this species. The structure elucidation of the isolated compounds was on the basis of 1D, 2D NMR spectroscopy and mass spectrometry measurements. Two compounds were isolated from the crude glycosides extract of T.davaeanum; 6 is a phenylethanoid glycoside and 8 is an iridoid glycoside, from the data available these may be new compounds for which the names davaeanuside A and davaeanuside B are proposed respectively." The total polyphenol content of S. europaea L, T. davaeanum leaves-flowers and T. davaeanum stem were found to be 5.0, 1.20 and 0.65 mg per 100 mg dried plant material respectively. A study of the antioxidant activity of the 50 % ethanol extracts of S. europaea and T. davaeanum showed that on a mg/mg basis S. europaea and T. davaeanum have approximately 5%, 8 % antioxidant capacity of Trolox respectively. A study of the cytotoxic activity of davaeanuside A (6), iridoid glycoside (7), davaeanuside B (8) and saponin compound (10) isolated from the crude glycosides extract of T. davaeanum revealed that saponin compound (10) inhibited the growth of Hela cells by 50 % at 50 μg/ml, P< 0.001, but the other compounds did not show activities against the tested cell lines at 100 μg/ml. The results of this work provide some basis for the traditional use of these species in the treatment of wounds.

Orientador: Helena Teixeira Godoy<br>Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos<br>Made available in DSpace on 2018-08-11T10:31:24Z (GMT). No. of bitstreams: 1 Scherer_Rodrigo_D.pdf: 928592 bytes, checksum: 5a302dc29cabc12b55ee37f7e292ef66 (MD5) Previous issue date: 2008<br>Resumo: No presente trabalho foram avaliados alguns extratos vegetais com relação à atividade biológica (antioxidante e antimicrobiana) e sua composição. Do carrapicho (Xanthium strumarium) foram estudados 18 extratos diferentes obtidos com extração supercrítica, hidrodestilação, maceração, maceração dinâmica e soxhlet com solventes de diferentes polaridades, incluindo metanol, etanol, acetato de etila e clorofórmio/diclorometano (1/1), enquanto que das plantas cravo-da-índia (Caryophillus aromaticus L.), citronela (Cymbopogon winterianus) e palmarosa (Cymbopogon martinii) apenas o óleo essencial. Também foram avaliadas as frações isoladas eugenol e eucaliptol. Para determinação da atividade antioxidante foi desenvolvido um novo índice utilizando o radical DPPH (2,2-difenil-1-picril-hidrazila), chamado de Índice de Atividade Antioxidante (IAA), através do qual foi possível comparar a capacidade antioxidante dos diferentes extratos vegetais com compostos de referência. A determinação da atividade antimicrobiana foi realizada através da determinação da concentração inibitória mínima (CIM), através da qual a ação antimicrobiana dos extratos foi comparada com a ação de alguns antibióticos frente aos microrganismos Staphylococcus aureus, Escherichia coli, Salmonella thyphimurium, Pseudomonas aeruginosa e Clostridium perfringens. Na avaliação da composição das frações voláteis, foi utilizada a cromatografia a gás com detector de massas para a identificação e com detector de ionização em chama para a quantificação dos compostos. Nos extratos fixos, foi utilizada a cromatografia líquida com detector de arranjo de diodos e detector de massas para a confirmação da identidade dos compostos fenólicos avaliados. Os resultados mostraram que os extratos hidrodestilados e extratos supercríticos de X. strumarium apresentaram uma composição qualitativa muito semelhante, entretanto, foram diferentes quantitativamente, pois no extrato hidrodestilado o _-guaieno foi encontrado com 79,6% da composição, enquanto que nos extratos supercríticos a concentração foi de 0,2 a 0,5%. Os extratos hidrodestilados e os extratos supercríticos de X. strumarium apresentaram ação antioxidante muito fraca, porém forte ação antimicrobiana, entretanto, os extratos fixos de X. strumarium apresentaram forte ação antioxidante e forte ação antimicrobiana. Nos extratos fixos de X. strumarium foram identificados dois compostos fenólicos majoritários, o ácido ferúlico e o ácido clorogênico, sendo que esses compostos foram encontrados em maiores quantidades nos extratos metanólicos e etanólicos do que nos extratos obtidos com acetato de etila e diclorometano/clorofórmio. Além disso, foi encontrado um alto grau de correlação entre o teor desses compostos com a atividade antioxidante dos extratos. Foi encontrado nos cotilédones e nas sementes de X. strumarium o composto hepatotóxico carboxiatractilosideo, mas não foi encontrado nas folhas em estádio adulto, sendo assim, o uso medicinal dessa planta deve ser restrito apenas as folhas na fase adulta. Tanto o óleo de cravo quanto o eugenol apresentaram atividade antioxidante muito forte e ação antimicrobiana forte a moderada sobre os microrganismos avaliados. Os óleos de citronela e palmarosa e o eucaliptol apresentaram fraca ação antioxidante, porém forte a moderada ação antimicrobiana. O eugenol é o componente majoritário do óleo de cravo seguido pelo _-cariofileno, enquanto que nos óleos de citronela e palmarosa, o _-citronelal e o geraniol foram os componentes majoritários, respectivamente. Considerando que o eucaliptol e o eugenol apresentaram boa atividade antimicrobiana, e o fato de serem frações isoladas, o que facilita o controle de qualidade e a aquisição de lotes homogêneos de amostras, os dois compostos foram escolhidos para serem testados como alternativa aos antibióticos utilizados como promotores de crescimento em frangos de corte no período de 1 a 21 dias. No experimento, foram utilizados 392 pintos machos distribuídos em 7 tratamentos onde foram utilizados o eucaliptol e o eugenol microencapsulados e o antibiótico avilamicina. Os resultados mostraram que a associação dos dois compostos afetou negativamente o desempenho das aves, entretanto, a utilização de eucaliptol (500 mg/kg) ou eugenol (500 mg/kg) melhorou o desempenho das aves em relação ao controle. Não foram verificadas diferenças significativas com o tratamento de 10 mg/kg de avilamicina, portanto, é possível substituir a avilamicina pelos extratos vegetais testados sem comprometer o desempenho das aves na fase inicial<br>Abstract: In this work the biological activity (antioxidant and antimicrobial), as well as the composition of several plant extracts were studied. For cocklebur (Xanthium strumarium), 18 different extracts were studied, including supercritical extraction, hydrodistillation, maceration, dynamic maceration and soxhlet with different extracting solvents, including methanol, ethanol, ethyl acetate and chloroform/dichloromethane (1/1), while clove (Caryophillus aromaticus L.), citronella (Cymbopogon winterianus) and palmarosa (Cymbopogon martinii) only the essential oil. The isolated fractions eucalyptol and eugenol were analyzed too. To the antioxidant activity evaluation, a new Antioxidant Activity Index (AAI) using the DPPH radical (2,2-diphenyl-1-picrylhydrazyl) was proposed, that use was possible to compare the antioxidant strength between plant extracts, as well as between pure compounds. The antimicrobial activity of samples was evaluated by the determination of the minimal inhibitory concentration (MIC), by the way the extracts were compared with antibiotics against the microorganisms Staphylococcus aureus, Escherichia coli, Salmonella thyphimurium, Pseudomonas aeruginosa e Clostridium perfringens. The composition of the volatile fractions was carried out using a gas chromatography coupled with mass spectrometer detector for the identification and coupled with flame ionization detector (FID) for the quantification of the compounds. For non-volatile fractions, a liquid chromatography coupled with diode array detector and coupled with mass spectrometer detector was used in the phenolic compounds analysis. The results showed that the composition of the X. strumarium extracts obtained by supercritical extraction and by hydrodistillation presented little qualitative difference, but they did differ quantitatively, 79.6% of _-guaiene was found in the hydrodistilled, while as 0.2 to 0.5% in the supercritical extracts. Hydrodistilled and supercritical extracts of X. strumarium presented poor antioxidant activity, but strong antimicrobial activity, however, the non-volatile fractions exhibit strong antioxidant and antimicrobial activities. In the non-volatile fractions, two phenolic compounds was indentified, the ferulic and chlorogenic acids, since that the concentrations were higher in methanolic and ethanolic extracts than the ethyl acetate and chloroform/dichloromethane extracts, moreover, high correlation between these compounds and antioxidant activity was found. The results show that carboxyatractyloside, a high liver toxic compound, is indeed present in the seeds and in the cotiledonary stage (two leaves stage) but not in adult leaves, therefore for medicinal use, only the adult leaves should be used. The clove oil and the eugenol presented very strong antioxidant activity and strong to moderate antimicrobial activity against tested microorganisms. The citronella and palmarosa oils and the eucalyptol showed poor antioxidant activity and strong to moderate antimicrobial activity. The eugenol was found as the majority compound in the clove oil followed by B-caryophyllene, while as in the citronella and palmarosa oils the B-citronellal and geraniol were the majority compounds, respectively. Considering that the eucalyptol and eugenol showed good antimicrobial activity and are isolated compounds, that make easy the control quality and the acquisition of the homogeneous samples batches, they are chosen compounds to be tested as alternative growth promoters in broilers from 1 to 21 days of age. In this experiment were used 392 male broilers (Ross), distributed in 7 treatments where used eucalyptol and eugenol microencapsulated and the antibiotic avilamicyn. The results showed that the association of the two compounds had negative effect in the broilers performance, however, when the eucalyptol (500 mg/kg) or eugenol (500 mg/kg) was used, the broilers performance were better than control group, and no significant difference to 10 mg/kg of avilamicyn was observed. So, the avilamycin can be substituted by the tested plant extracts without to lose the performance of birds in the growing phase<br>Doutorado<br>Doutor em Ciência de Alimentos

Submitted by CARITA LIBERATO DO AMARAL null (caritaliberato@gmail.com) on 2017-04-11T17:12:57Z No. of bitstreams: 2 Tese_Carita_Liberato_do_Amaral.pdf: 5356975 bytes, checksum: ea2885d6ecdf894dd43cb282fa498e09 (MD5) Tese_Carita_Liberato_do_Amaral.pdf: 5356975 bytes, checksum: ea2885d6ecdf894dd43cb282fa498e09 (MD5)<br>Rejected by Luiz Galeffi (luizgaleffi@gmail.com), reason: Solicitamos que realize uma nova submissão seguindo as orientações abaixo: Foram submetidos 2 arquivos PDF’s, apenas 1 arquivo deve ser submetido. O arquivo PDF não deve estar protegido e a dissertação/tese deve estar em um único arquivo, inclusive os apêndices e anexos, se houver. Corrija estas informações e realize uma nova submissão contendo o arquivo correto. Agradecemos a compreensão. on 2017-04-17T19:13:40Z (GMT)<br>Submitted by CARITA LIBERATO DO AMARAL null (caritaliberato@gmail.com) on 2017-04-17T19:31:41Z No. of bitstreams: 1 Tese_Carita_Liberato_do_Amaral.pdf: 5356975 bytes, checksum: ea2885d6ecdf894dd43cb282fa498e09 (MD5)<br>Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-04-17T19:42:22Z (GMT) No. of bitstreams: 1 amaral_cl_dr_jabo.pdf: 5356975 bytes, checksum: ea2885d6ecdf894dd43cb282fa498e09 (MD5)<br>Made available in DSpace on 2017-04-17T19:42:22Z (GMT). No. of bitstreams: 1 amaral_cl_dr_jabo.pdf: 5356975 bytes, checksum: ea2885d6ecdf894dd43cb282fa498e09 (MD5) Previous issue date: 2017-02-20<br>Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)<br>Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)<br>As condições ambientais, assim como o contraste sazonal nas fitofisionomias do Cerrado entre a época seca e úmida colaboram para a diversificada sintetize e seleção de metabólitos secundários em plantas. Os metabólitos secundários atuam como uma interface química entre plantas e o meio, desta forma a sua síntese pode ser comumente alterada pelas condições ambientais. Fatores como temperatura, disponibilidade hídrica, luminosidade, características do solo, nutrientes e poluição atuam diretamente sobre a produção e liberação desses compostos. Os microrganismos do solo podem transformar os compostos tóxicos liberados, desativando-os ou dando origem a compostos ainda mais tóxicos, assim como esses compostos podem alterar as características e atributos do solo quando presentes em quantidades adequadas. Dentre as plantas nativas do Cerrado destaca-se a espécie Vernonia ferruginea, uma agressiva infestante de pastagem, que possui indícios de produção de compostos com potencial tóxico. Desta forma, esta pesquisa foi desenvolvida para verificar a atividade tóxica de V. ferruginea e se a mudança de algumas características como disponibilidade hídrica, tipo de solo, condições de luminosidade e concentração de alumínio no solo influenciam a produção desses compostos. Para tal, plantas de V. ferruginea foram desenvolvidas sobre diferentes condições para posterior extração e/ou fracionamento do material vegetal para verificação de atividade biológica por meio de bioensaios e identificação de grupos químicos. Considerando os resultados dos parâmetros de germinabilidade e de desenvolvimento das espécies testadas é possível indagar sobre a possibilidade de V. ferruginea ter a capacidade de liberar substâncias tóxicas, uma vez que, consequências diretas da presença desses compostos nos extratos foram verificadas no bioensaio, como: a inibição/redução da germinação de sementes, injúria ou necrose no sistema radicular e redução do desenvolvimento e do acúmulo de massa seca das plântulas. Os resultados também dão base para constatar que algumas interações da espécie com as alterações nas suas condições de desenvolvimento causam desvios de rotas biossintética de metabólitos modificando e/ou potencializando sua capacidade fitotóxico.<br>The environmental conditions, as well as the seasonal contrast in the Cerrado phytophysiognomies between the dry and wet seasons, contribute to the diversified synthesis and selection of secondary metabolites in plants. Secondary metabolites act as a chemical interface between plants and the environment, so their synthesis can be commonly altered by environmental conditions. Factors such as temperature, water availability, luminosity, soil characteristics, nutrients and pollution act directly on the production and release of these compounds. Soil microorganisms can transform the released toxic compounds, deactivating them or giving rise to even more toxic compounds, as these compounds can alter the characteristics and attributes of the soil when present in adequate amounts. Among the native plants of the Cerrado, the species Vernonia ferruginea, an aggressive pasture weed, stands out, with indications of production of compounds with toxic potential. Thus, this research was developed to verify the toxic activity of V. ferruginea and if the change of some characteristics such as water availability, soil type, light conditions and aluminum concentration in the soil influence the production of these compounds. For this, plants of V. ferruginea were developed on different conditions for later extraction and / or fractionation of the vegetal material for verification of biological activity by means of bioassays and identification of chemical groups. Considering the results of the parameters of germinability and development of the tested species it is possible to inquire about the possibility of V. ferruginea being able to release toxic substances, since direct consequences of the presence of these compounds in the extracts were verified in the bioassay, such as: The inhibition / reduction of seed germination, injury or necrosis in the root system and reduction of the development and accumulation of dry mass of the seedlings. The results also provide evidence that some interactions of the species with changes in their developmental conditions cause biosynthetic pathway deviations of metabolites modifying and / or potentiating their phytotoxic capacity.<br>FAPESP: 12/25298-0

碩士<br>國立臺灣大學<br>園藝學研究所<br>102<br>Mushrooms have been not only used as food materials with their unique flavor and texture, but also recognized as an important source of biologically active compound of medicinal value. Mushrooms have a variety of accumulated secondary metabolites such as phenolic compounds, polypeptides, terpenes, and steroids. Mushrooms also have lectins, polysaccharides, polysaccharide-peptides, and polysaccharide-protein complexes which are known to have immunomodulatory and anticancer activities. Mushroom is rich in polysaccharides, and the polysaccharides has been recently researched for many biological and medical tests, such as treating diabetes, lowering the cholesterol, anti-aging, cancer-inhibiting and so on. Pink oyster mushroom (Pleurotus djamor) is a kind of mushrooms emerging on the market, and it is one of the oyster mushroom in Pleurotus. Its polysaccharide content and the biological activity has yet to be fully discovered, so that this test through the MTT cell viability test by the cervix cancer cells (Hela) to observe the biological activity of pink oyster mushroom and also we observe the morphology change of different cancer cell lines through adding the polysaccharides.

碩士<br>亞洲大學<br>保健營養生技學系碩士班<br>99<br>Tridax procumbens Linn. is used in folk medicine. It is reported that the plant shows antibacterial activity and contains antitumor chemical components. The purpose of this research was to evaluate antibacterial, antioxidation, anti-inflammation and anticancer activity of T. procumbens Linn. The flowers , leaves and stems of T. procumbens were extracted with ten times of ethyl alcohol. The extract was then partitioned by n-hexane, ethyl acetate, n-butyl alcohol and water to evaluate its antimicrobial and antioxidative activities. We also evaluate its anti-inflammation and anticancer activity of the flower. The result showed that most of extracts had antimicrobial activities extracts from flower. The ethyl acetate extract was the most effective among all the extracts. Most extracts from flowers, leaves and stems showed the good antioxidative activities especially the flower. The the ethyl acetate extracts presented the best on DPPH scavenging activity and reducing power and had the highest total phenolic compounds content . We used the ethyl alcohol and water extracts of T. procumbens to treat RAW264.7 macrophage, which induced inflammation by Lipopolysaccharide (LPS).In the NO assay, the extracts of T. procumbens flower had better inhibit In MTT assay of SK-Hep-1, we found that extract of T. procumbens flower had the best inhibitory ability. We also found the ethyl alcohol extract of T. procumbens had the best to diminish the expression of matrix metalloproteinase (MMP)-9 and MMP-2. The ethyl alcohol extracts of flower had good anti-migration and anti-invasion ability.

碩士<br>國立高雄海洋科技大學<br>水產食品科學研究所<br>105<br>Squid is important catches in Taiwan's offshore fishing ranked third in the world catch of squid. However, the squid processing will produce a lot of waste, such as viscera and ink sac that were discarded or as byproducts. Among them, squid ink (SI) accounted for about 1.3% of body weight has fish stench and is difficult to utilize. Some reports had been revealed that Cephalopods ink contained many bioactive substances, such as antioxidant, antibacterial and antitumor activity. Therefore, the antioxidant, antibacterial, angiotensin-I-converting enzyme (ACE) inhibitory activity and antitumor effects of neon flying squid (Ommastrephes bartramii) SI extracts extracted by methanol, ethanol, hexane and acetone as well as SI polysaccharide were evaluated. In ACE inhibitory activity, the SI extracts had no or very low ACE inhibitory activity (<0.2%), except for 1000 ppm of SI polysaccharide (SIP) with better ACE inhibitory activity (15.98%). No antibacterial activity against food pathogens was observed in all SI extracts. Cytotoxicity assay was performed using MTT assay on HeLa cell line and HL-60 cell line (Human promyelocytic leukemia cells). The cell viability of HeLa cells treated with SIP and SI methanol or ethanol extracts was decreased and the highest inhibitory rate reached 30% at SIP treatment. However, no or very low inhibitory activity on HL-60 cells treated with SIP and SI methanol or ethanol extracts was observed. In antioxidant activity, except for hydroxyl radical scavenging ability, the SIP showed higher antioxidant ability in scavenging DPPH, ABTS radical scavenging, and ferrous ion-chelating ability than SI methanol and ethanol extracts. When the SIP was separated using ultrafiltration into two fractions (<5 KDa and >5 KDa), the fractions with MW >5 KDa showed to elevate DPPH radical scavenging activity and hydroxyl radical scavenging ability. The SIP contained 65.03% total sugar, 0.14% protein, 0.1% lipid, 11.36% ash, 14.24% uronic acid, and 1.84% sulfate.

碩士<br>中山醫學大學<br>營養科學研究所<br>91<br>Propolis is a very popular nature health food currently. However, the composition and biological actives of propolis extracts and their digested products are still deficient. Therefore,the composition and biological actives of propolis extracts and their digested products will be discussed in this study. 95EtOH, 60EtOH, 30EtOH, aqueous and glycerol extracts of propolis were used in this study. Firstly,the phenolic composition and biological actives in various propolis extracts and their digested products will be determined . Secondly ,We discuss the change of phenolic composition and biological actives of the extracts after artifical gastric digestion or gastrointestinal digestion . Because phenolics are found to be the major available physiological contributor of propolis, we try to understand the difference of propolis composition of propolis by the digesting treatment. Biological evaluations includes total antioxidant capacity, reducing power, free redical scavenging efficiency and suppression efficiency for COX-2 expression. Result revealed that 95％EtOH and 60％EtOH extracts of propolis showed similar phenolic composition and biological actives.Both 30％EtOH and aqueous extract of propolis showed lessof phenolics and lower biological actives than that of 95％EtOH and 60％EtOH extract. The phenolic composition and biological effect of glycerol extract are between the above two groups. By the analysis of HPLC, phenolics were greatly decreased and changed, especially in 30％EtOH and aqueous extracts. The original extracts and digested extract of 95％EtOH and 60％EtOH treatments total antioxidant capacity,H2O2 scavenging, reducing power, DPPH, free redical scavenging and suppression efficiency for COX-2 expression.When Chelating , Fe+2 and superoxide anion scavenging is considered, before digestion processing ,performance of the aqueous extract is the best.Original aqueous extract showed perfect effect on cheating for ferrous ions , scavenging the superoxide anions.However, the functions the digests of the aqueous extract was found rednced evidently. By comparing the results of HPLC, the tannin-like materials with high polarity in the range of 0-50 minutes were probably the main contributor for cheating ferrous ions and scavenging superoxide anion.

Tese de doutoramento, Bioquímica (Bioquímica Farmacêutica e Toxicológica), Universidade de Lisboa, Faculdade de Ciências, 2012<br>The Plectranthus barbatus herbal tea is traditionally used to treat a wide range of health conditions, including psychiatric problems, gastrointestinal disturbances and inflammation‐related conditions. The aim of this work was to determine if P. barbatus herbal tea may be useful in the treatment of acetylcholinesterase or inflammation related diseases, such as Alzheimer’s disease. In vitro activities of the P. barbatus aqueous extract were determined, namely antiacetylcholinesterase activity, antioxidant activity as radical scavenger and preventing lipid peroxidation, and anti‐inflammatory activity by decreasing the amount of hypochlorous acid produced by activated neutrophils. The values obtained for the in vitro activities were very promising and related with its content in rosmarinic acid, flavonoid glucuronides and abietane diterpenoids. After in vitro digestion with gastric and pancreatic juices, bacterial β‐glucuronidase and metabolization by Caco‐2 cells, the anti‐acetylcholinesterase activity of the plant extract suffered a slight decrease due to the loss of one active diterpenoid. The bioavailability of the plant extract was determined by administering it to rats and analyzing the rat plasma and brain. The extract components suffered glucuronidation, sulfation and methylation by the intestine and by the liver, but the plant compounds were found in rat brains and brain acetylcholinesterase activity showed an inhibition reaching 30%. As the bioavailability of rosmarinic acid was not the same when in the extract and when alone, the interference of plant phenolics on the permeability of each other was tested in Caco‐2 monolayers using rosmarinic acid and two flavonoids, apigenin and luteolin. This study showed that the compounds have higher intestinal permeability when in a mixture due to the inhibition of the efflux transporters that limit their bioavailability. The compounds from the P. barbatus extract can bind to the protein structure of acetylcholinesterase, human serum albumin and lysozyme by weak interactions such as hydrophobic interactions and hydrogen bonds. These interactions are the cause of the reversible inhibition of the enzymatic activity of acetylcholinesterase and lysozyme; therefore the compounds are less susceptible to cause side effects when used therapeutically. The P. barbatus herbal tea may be used to treat cholinesterase‐related problems such as gastrointestinal conditions and Alzheimer’s disease as its active components may reach the target organs, and brain acetylcholinesterase inhibition was detected. The compounds may circulate in the plasma bound to albumin and lysozyme, and may decrease inflammation by their radical scavenger activity, by decreasing neutrophil‐produced hypochlorous acid and by inhibiting lysozyme activity.<br>Infusões e decocções de Plectranthus barbatus são usadas tradicionalmente para uma grande diversidade de fins terapeuticos, incluido o tratamento de problemas psiquiátricos, distúrbios gastro‐intestinais e doenças relacionadas com processos inflamatórios. O objectivo deste estudo é determinar se o extracto aquoso de P. barbatus, preparado como decocção, poderá ser útil no tratamento de problemas relacionados com a actividade do enzima acetilcolinesterase ou com processos inflamatórios. Um extracto aquoso de P. barbatus foi preparado como decocção e foram determinadas, in vitro, as actividades anti‐acetilcolinesterase, antioxidante e anti‐inflamatória. Nestas actividades in vitro foram obtidos resultados muito promissores, com valores de IC50 baixos para a inibição da actividade do enzima acetilcolinesterase, actividade antioxidante no sequestro de radicais livres e protegendo lipidos de peroxidação, e na actividade antiinflamatória pela diminuição da quantidade de ácido hipocloroso produzido por neutrófilos activados. Estas actividades estavam relacionadas com a composição do extracto de P. barbatus, sendo ácido rosmarinico o composto maioritário, mas encontrando‐se também presentes flavonóides glucuronados (apigenina 7‐O‐glucurónido, luteolina 7‐O‐glucurónido e acacetina 7‐O‐glucurónido) e diterpenoides. Após a digestão in vitro do extracto vegetal com sucos gástrico e pancreático artificiais, da acção de β‐glucuronidase de bactérias da microflora intestinal e da metabolização por células Caco‐2, como modelo de células intestinais humanas, a actividade do extracto sofreu uma diminuição devido à perda de um diterpenoide activo, no entanto os outros compostos activos permaneceram intactos. A biodisponibilidade da decocção de P. barbatus foi determinada por administração intragástica e intraperitoneal a ratos, recolhendo‐se e analisando‐se o plasma e o cérebro dos ratos por HPLC. Os componentes da decocção foram metabolisados no intestino e no fígado, sofrendo glucuronidação, sulfatação e metilação. Encontrou‐se ácido rosmarínico no cérebro dos ratos após a administração do extracto, e o cérebro apresentava uma inibição da actividade da acetilcolinesterase atingindo aproximadamente 30%, sugerindo que outros compostos ou metabolitos dos componentes da decocção pudessem estar presentes em quantidades inferiores ao limite de detecção, mas que mesmo assim iriam influenciar a actividade enzimática. Como se encontraram algumas alterações entre a biodisponibilidade do ácido rosmarínico administrado no extracto ou administrado isolado, procedeu‐se ao estudo da interferência que compostos fenólicos possam ter na permeabilidade uns dos outros quando administrados em misturas, em membranas de células Caco‐2. Nesse estudo recorreu‐se ao método de “central composite design” CCD para determinar se existiriam xii diferenças entre a permeabilidade e metabolização de ácido rosmarínico, apigenina e luteolina em membranas de células Caco‐2, induzidas pela presença uns dos outros. Foram também coadministrados substractos de dois sistemas transportadores conhecidos, o transportador de ácidos monocarbóxilicos (MCT) e a glicoproteina‐P (Pgp), e concluiu‐se que os compostos da decocção de P. barbatus tinham maior permeabilidade por estarem em conjunto, uma vez que uns compostos inibiam os transportadores de efluxo dos outros. A biodisponibilidade dos substratos dos transportadores de efluxo presentes na membrana apical aumenta com a inibição dos transportadores, pois estes transportam activamente os seus substratos para o lúmen do intestino, limitando assim a passagem para a circulação sanguínea. A interacção entre os compostos do extracto de P. barbatus e proteinas foi avaliado através de técnicas de espectrometria de fluorescência e FTIR. Os componentes da decocção de P. barbatus têm a capacidade de se ligar à estrutura proteica da acetilcolinesterase, da albumina de soro humano e da lisozima através de interacções fracas, nomeadamente interacções hidrofóbicas e pontes de hidrogénio. Por se tratarem de ligações fracas e não se terem encontrado alterações da estrutura secundária das proteinas, estas interacções serão responsáveis pela inibição reversivel da actividade enzimática da acetilcolinesterase e da lisozima. Este tipo de interacções são as mais recomendadas para compostos a ser utilizados para fins terapeuticos pois evitam efeitos secundários resultantes da inibição destes enzimas por inibidores que formam complexos através de ligações mais fortes, como ligações covalentes, e induzem alterações profundas na estrutura proteica levando à desnaturação. Decocções de P. barbatus poderão ser utilizadas para tratar problemas relacionados com a actividade da acetilcolinesterase, como a doença de Alzheimer, pois após a administração os seus componentes poderão ser encontardos a a nivel do intestino, plasma e cérebro, inibindo a actividade da acetilcolinesterase no cérebro. Os componentes da decocção poderão circular na corrente sanguínea associados à albumina e à lisozima, e poderão reduzir processos inflamatórios devido à sua actividade sequestradora de radicais livres, à sua capacidade de diminuir a quantidade de àcido hipocloroso produzido por neutrófilos activados, e por inibir a atividade enzimática da lisozima.<br>Fundação para a Ciência e a Tecnologia (FCT, SFRH/BD/37547/2007)

In most countries of subtropical Africa, fungal infections represent an increasing problem. Candida albicans is the most common fungal pathogen and has developed an extensive array of putative virulent mechanisms that allows successful colonization and infection of the host under suitable predisposing conditions. Crude acetone, water and hexane extracts of eight plant species, used as traditional medicine to treat fungal related diseases were evaluated for antifungal activity against C. albicans standard strain (ATCC 10231) and six clinical isolates using a serial microplate dilution method. Antioxidant activity was also determined by using the TEAC and DPPH assays and cytotoxicity was determined against mouse fibroblast cells with a MTT assay. MIC’s below 1 mg/ml were observed for the acetone and water extracts of Sclerocarya birrea (stem-bark), as well as the acetone extracts of Harpephyllum caffrum (leaves), Ptaeroxylon obliquum (leaves) and Rapanea melanophloeos (leaves). Bioautography confirmed the presence of compounds with antifungal activity in extracts of S. birrea and P. obliquum. S. birrea extracts had the highest antioxidant activity and the lowest cytotoxicity. However, P. obliquum contained the highest number of active compounds against C. albicans standard strain (ATCC 10231) and clinical isolates. MIC data also indicated that P. obliquum was the most active of all the plant species tested against C. albicans standard strain (ATCC 10231) and clinical isolates. Therefore, P. obliquum was chosen for bioassay-guided fractionation and isolation of antifungal compounds from P. obliquum acetone leaf extract. Two compounds were isolated from the most active fraction obtained from P. obliquum acetone leaf extract, using column chromatography with silica gel as the stationary phase. Compound 1 and Compound 2 inhibited the growth of C. albicans standard strain (ATCC 10231) at MIC 0.004 mg/ml compared to amphotericin B, MIC 0.11 mg/ml. The cytotoxicity of the compounds was determined against mouse fibroblast cells using the MTT assay. Berberine was used as a positive control and was found to be toxic with a LC50 of 9 04 μg/ml. The cytotoxicity of Compound 1 (against mouse fibroblast cells) was 0.002 μg/ml and that of Compound 2, 7.23 μg/ml. Selectivity Index (SI) of Compound 1, against both C. albicans standard strain (ATCC 10231) and clinical isolates, was 0.0004 and for Compound 2 it was 1.8 against the standard strain (ATCC 10231) and >0.03 against the clinical isolates tested. The structures of the compounds were determined by extensive nuclear magnetic resonance (NMR) techniques and chemical methods mainly by 1D NMR (1H, 13C and DEPT) and 2D NMR (HSQC, HMBC and COSY) and by comparison with the literature data. Compound 1 is an isomeric mixture of β-amyrin and lupeol. Compound 2 is a novel compound and was identified as: 8,11-Dihydro-5-hydroxy-12-hydroxymethyl-2-methyl-4H-pyrano[2,3-g] [1]benzoxepin-4-one 12-O-acetate. Candidal adherence to human buccal epithelial cells (HBEC) is considered the critical initial step in the pathogenesis of oral candidiasis, which may eventually lead to a systemic infection, especially in immuno-compromised individuals. Therefore, it was important to determine the ability of extracts and isolated compounds on the adherence of C. albicans standard strain (ATCC 10231) and clinical isolates to HBEC cells. P. obliquum acetone leaf extract at concentrations of 15.6 mg/ml and 7.8 mg/ml inhibited the proliferation and hyphae formation of C.albicans and thus adhesion. Natural products inhibiting adhesion of C. albicans to the oral mucosa may be beneficial in managing oral candidiasis. The high antifungal activity of the acetone leaf extract of P. obliquum indicates that it could be considered as a potential therapeutic agent against oral Candida infections. The therapeutic effects of this extract may be limited only to low concentrations to avoid cytotoxic reactions as the cytotoxicity was less favourable (LC50 = 35.58 μg/ml against mouse fibroblast cells). However, as oral rinses are used topically and not swallowed the systemic effect may not be of critical importantance.<br>Thesis (PhD)--University of Pretoria, 2012.<br>Paraclinical Sciences<br>Unrestricted

The control of Pratylenchus goodeyi a common nematode parasite of banana crop in Madeira Island can benefit from searching for natural nematicides through plants extracts. With this aim we submitted Solanum nigrum and S. sisymbriifolium dried plants to a sequential extraction in the solvent sequence of dichloromethane, acetone, ethanol and water, and to na aqueous extraction of the fresh and dried plants. Analyses with the extracts at several concentrations were used to assess mobility and mortality on P. goodeyi. Results showed that the water extract and aqueous extracts from both plants at a concentration of 10 mg/mL affected nematode mobility and caused mortality but the acetone extract from S. nigrum was the most efficient, causing 100% mortality whereas dichloromethane had no effect on P. goodeyi. Determination of the lipophilic and phenolic compounds present in the two most effective Solanum extracts (acetone and water) and in dichloromethane extract revealed that some of these compounds had nematicidal activity. S. nigrum acetone extract (10 mg/mL) was used to find out the nematicidal potential following the effect at gene expression level and nematode behaviour. Genes coding for calreticulin and beta-1,4- endoglucanase related to parasitism and translocon-associated protein putatively connected to stress were obtained and its relative expression assessed in nematodes exposed to the extract. Results revealed that expression of Pg-CRT decreased showing to influence the infection, Pg-ENG remained steady and Pg-TRAPδ was induced over time exposure. Biological assays showed that P. goodeyi mobility and ability to infect the banana roots were affected as a decrease in the number of nematodes that reached the roots was obtained with the increased exposure time to the extract being implicated in the infection success. The information obtained from this thesis showed that S. nigrum has potential to be used for the development of a new control strategy against plant-parasitic nematodes.<br>Agência Regional para o Desenvolvimento da Investigação, Tecnologia e Inovação