Dissertations / Theses on the topic 'Plant-based phytochemicals and extract'

The current state of agriculture, where demand for safe food is increasing rapidly as a consequence of growing population, raises a number of questions related to the one health approach and sustainable animal production with minimal impact on the environment. Swine production is an important branch of food production where weaning is the most vulnerable phase for piglets, often associated with decrease of growth performance and diarrhoea. The maintenance of gut health is therefore a complex endeavour where nutrition is crucial in order to reduce the intestinal disorders. Antimicrobial resistance is also a significant global concern. Reducing antibiotic use in animal production systems decreased prevalence of antibiotic-resistant bacteria in animals about 15%. In the last decade, the European Union banned the antibiotic use as growth promoters in livestock (EU Reg. 1831/2003). The first antibiotic alternative was the wide application of essential nutrients such as zinc (Zn) and copper (Cu) salts in the form of premix in the diets of animals to control digestive disorders. Due to their low bioavailability, Zn and Cu are commonly found in animal’ manure as a reflection of their content in the feed. The use of Zn and Cu in feed may also have contributed to the emergence of methicillin-resistant Staphylococcus aureus (MRSA). Despite antibacterial and anti-inflammatory activities, the first adopted alternative against in-feed antibiotics became unsafe due to heavy metal’ pollution in livestock wastewater. In order to reduce the high concentration of Zn and Cu and the antibiotic use in animal diets, plant extracts and different phytochemicals are of potential interest due to their antimicrobial, antioxidant and anti-inflammatory properties. However, if nutritional ecology’ strategy is not sufficient to reduce the wastewater pollution of heavy metals from livestock production, the development of efficient methods such as multidisciplinary phytoremediation approach is required. First, the preliminary aim was to overview of the role and the main challenges related to the content of essential heavy metals in animal feed and to evaluate the concentration of heavy metals from feed and faeces in animal rearing systems in northern Italy. Based on an overview, the main second aim was to develop a plant-based integrated approach to reduce the input and output of both Zn and Cu as well as the use of antibiotic compounds in pig production. Hence, in order to reduce input, the first aim was to test several natural plant-based phytochemicals compounds (tannins and leonardite) in vivo and to test of the anti-inflammatory effects of peppermint oil and spearmint oil with porcine alveolar macrophages in vitro. The last aim was to assess the ability of two aquatic species, Typha latifolia and Thelypteris palustris to control the Zn and Cu output from contaminated livestock wastewaters as a cost-efficient phytoremediation strategy. The in vivo data revealed that natural plant extracts (leonarditre and tannins) improved animal health. High doses of tannins (1.25%) supplementation showed slight reduction of diet digestibility and protein utilization, however this did not influence on feed intake and growth performance of animals. The inclusion of 0.25% leonardite improved the zootechnical performance, serum lipid profile and gut epithelium integrity, indicating a good general health status. In vitro study results showed that both mint oils significantly reduced TNF-α secretion from macrophages. To conclude, leonardite supports an improved stress response in weaned piglets, high dose of tannins did not impair growth performance and both peppermint and spearmint oils had anti-inflammatory activities in vitro. Moreover, results obtained from the phytormediation trial showed that Typha latifolia and Thelypteris palustris can accumulate and translocate Zn and Cu from contaminated wastewater. Thus, phytoremediation was effective to counteract the output of zinc and copper, and possibly other heavy metals from the livestock industry. Hence, an integrated nutritional ecology strategy and phytoremediation approach, in accordance with the modern principles of agroecology is needed to reduce the antibiotics use and heavy metals pollution in food-producing animals. Moreover, plant-based strategy guarantees the improvement of the health status of human and animal and leads to increase of the sustainability in animal rearing systems.

>Magister Scientiae - MSc
Nanotechnology has emerged as a promising field in the quest to address health conditions. Green nanotechnology is a fairly new branch of nanotechnology, which aims to produce and utilize nanomaterials in a way that is safe for living organisms and their environment. Plant extracts are increasingly used in the green synthesis of gold nanoparticles (AuNPs), which involves the reduction of sodium tetrachloroaurate (III) dehydrate by phytochemicals present in the plant extract. It is probable that the green synthesised AuNPs are more biocompatible than chemically synthesised AuNPs as biomolecules of plant origin are involved in the synthesis process. Therefore, this study aimed to explore various water extracts from indigenous South African plants, which included Perlagonium capitatum, Otholobium bracteolatum, Gerbera linnae, Morrella quercifolia, Searsia lucida, Phylica bubescens, Euclea racemosa, Tetragonia fruticosa, and Searsia glauca for their potential to synthesize AuNPs and to investigate their toxicity towards several microorganisms known to cause skin infections. These organisms play a significant role in delaying the healing of wounds. The antimicrobial properties of nanoparticles are increasing exploited in the production of wound treatments.

This thesis represents studies on aspects of the defenses against insect herbivores in species of the plant family Meliaceae, particularly with regard to phytochemicals. Methanolic extracts of foliage from thirty species in twenty-two genera were bioassayed for toxicity and growth inhibitory activity against the variegated cutworm, Peridroma sauciaf and for feeding inhibition against the migratory grasshopper, Melanoplus sanguinipes. All but three species were inhibitory to P. saucia, members of the tribe Melieae being most inhibitory. Members of the subfamily Melioideae were more active than members of the Swieteniodeae. Newly identified species with activity comparable to neem (Azadirachta indica) foliage extracts included Aglaia odorata and Turreae holstii. Deciduous species produced extracts which were significantly more active than evergreen species, indicating a greater reliance on phytochemical-based defenses. Evidence is also presented to suggest that the leaves of evergreen species are tougher than deciduous species, and that there is a negative correlation between leaf toughness factors (physical defenses) and phytochemical-based defenses. These results are in agreement with predictions of the resource availability hypothesis. The phytochemistry of Aglaia odorata, A. odoratissima. and A. argentia was examined in detail. Compounds identified included the dammaranes, aglaiondiol and aglaitriol, and the bis-amides (S,S)-odorine, (S,R)-odorine (a new natural product), (S,S)-odorinol, and (S,R)-odorincT. Three dihydroflavanones were identified from the Meliaceae for the first time: 3-hydroxy-5,7,4'-trimethoxyflavanone (a new natural product), 5,7,4'-trimethoxyflavanone, and 5-hydroxy-7,4'-dimethoxyflavanone. All compounds were inactive against P. saucia. The inhibitory activity of A. odorata appeared to be due to a compound, tentatively identified as a limonoid, which may be acting in conjunction with a synergist. This compound inhibits P. saucia larval growth in the absence of antifeedant activity. The toxicology of limonoids, representing the major biosynthetic classes, was examined against P. saucia and the large milkweed bug, Oncopeltus fasciatus. Cedrelone and anthothecol inhibited P. saucia growth by 90%, but not feeding, when applied in diet at 0.5 μmol/g fwt. Cedrelone also inhibited O. fasciatus molting, with an MD₅₀ of 12.2 μg/nymph. In contrast, anthothecol, with an acetoxy function at C-11, was inactive against O. fasciatus. The D-seco compound gedunin, and the A,D-seco limonoids obacunone, nomilin, and pedonin were inactive in these assays; harrisonin initially inhibited feeding by neonate P. saucia but produced no long-term effects on growth rate. Bussein inhibited growth by 35% but entandrophragmin had no effect. Azadirachtin was the most toxic compound examined in this study. Peridroma saucia growth (EC₅₀ =0.4 nmol/g diet fwt), survivorship (LC₅₀ =5.2 nmol/g), pupation, pupal weight, and adult emergence were decreased in a dose-dependent manner. Chemosensory antifeedant activity was implicated in neonates but was much less marked with third instar larvae. Azadirachtin decreased relative growth and consumption rates at doses lower than those affecting nutritional efficiency, or feeding in the choice tests. This suggests an action directly on the gut or on the neural regulation of feeding. Bioactivity of other limonoids did not correlate with measures of skeletal oxidation or rearrangement, although these are dominant themes in the evolution of the limonoids. Melanoplus sanauinipes lacked an antifeedant response to azadirachtin, up to concentrations of 500 ppm. However, subsequent molting was markedly effected. Application of azadirachtin orally, topically, or by injection, allowed determination of the role of the gut and integument in limiting the bioavailability of this compound to putative target site(s) within the insect. The oral MD50, 10.8 μg/g insect fwt, was significantly higher than the injected MD₅₀, 3.01 μg/g, indicating a barrier to bioavailability in the gut. The oral activity of azadirachtin was synergised by coadministration of piperonyl butoxide, indicating that the barrier is due largely to oxidative metabolism. There was no significant difference between topical (3.8 μg/g) and injected activity, indicating that the integument does not pose a barrier to bioavailability. Azadirachtin decreased growth and consumption at doses which did not affect nutritional efficiency, again indicating an effect on the gut or neural regulation of feeding. No difference was seen in nutritional indices of nymphs treated with azadirachtin at 10 and 15 μq/q, although these doses produced markedly different effects on molting. This observation suggested that effects on endocrine events are not directly related to nutritional effects. The effects of azadirachtin treatment were not alleviated by dietary supplementation with cholesterol, and azadirachtin did not affect the hemolymph transport or metabolism of ¹⁴C-β-sitosterol, indicating that sterol metabolism is not the target for azadirachtin activity. Azadirachtin also did not form adducts with cysteine, suggesting that non-specific binding to sulfhydryl-rich protein is also unlikely as a mechanism of action.
Science, Faculty of
Botany, Department of
Zoology, Department of
Graduate

BACKGROUND: Alzheimer's disease (AD) is a complex, multifactorial neurological disorder characterized by an insidious onset with progressive symptoms of memory impairment, language deficit, behavioural problems including agitation, mood disturbances and poor judgment. AD involves multiple pathogenetic factors such as A~ production and aggregation, oxidative stress, tau protein aggregation, metal ions (e.g. Cu2+, Zn2+, Fe2+) and reduced levels of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) enzymes. All these factors play important roles in neurodegeneration associated with AD. These provide diverse multiple targets for examining AD-modifying drugs. In view of this, the study was focussed on finding natural plant extracts used traditionally for centuries to treat memory and cognition related disorders specifically AD. Attention was also focussed on certain phytochemicals to find their therapeutic values in AD. AIMS: To find the neuroprotective, anti-cholinesterase and antioxidant activities of an aqueous extract of Withania somnifera, Bacopa monniera, Salvia patens, Salvia elegans, Capsicum annum, Uncaria tomentosa, Melissa officinalis and Centella asiatica plants. Four phytochemicals namely allicin, ajoene, capsaicin and asiatic acid were also examined for these activities. The targets chosen in this study were AChE and BuChE, neurotoxic A~ fibrils and oxidative stress pathway using in vitro models. METHDOLOGY: The aqueous extracts were obtained by boiling dried powdered plant materials in deionized water for 25 min. Enzyme assay were performed to determine anti-ChE activity by Ellman's method, while kinetics (Km and Vmax) were analyzed using Lineweaver-Burk plot method. Antioxidant capacities of these plant extracts and phytochemicals were determined using FRAP and TEAC assays. The neuroprotective activity of plant extracts and phytochemicals were determined under in vitro condition in differentiated pheochromocytoma (PC12) cell culture against H202 and A~ induced toxicities. X111 The anti-aggregation property was determined by transmission electron microscopy (TEM) and the Thioflavin T (ThT) fluorescence assay. RESUL TS: An aqueous extract of W somnifera showed dual inhibitory activities against both AChE and BuChE. The ICso values for AChE and BuChE were O.l5±0.007 and 0.80±0.005mg/mL, respectively. The potency of inhibition was greater for AChE compared with BuChE. The mode of inhibition was non- competitive mixed inhibition. Other plant extracts displayed either weak or negligible enzyme inhibition activity. In contrast, allicin demonstrated potent dual enzyme inhibitory activity. The ICso values for AChE and BuChE inhibition were 0.01±0.009 and 0.05±0.018mg/mL, respectively, while ajoene demonstrated weak enzyme inhibitory activity compared with allicin. The ICso values for AChE and BuChE inhibition of ajoene were 0.55±0.012 and 0.07±0.015mg/mL, respectively. Most of the plant extracts possessed high antioxidant capacities; FRAP values: 4242±112 to 1364644±130J,!mol Fe2+E/g dried weight; TEAC values: 1.89±0.12 to 26.20±0.05 mmol Trolox E/g dried weight. In contrast, allicin, ajoene and capsaicin showed weak antioxidant capacities; FRAP values: 1615±90, 934±35 and 1423±75 umol Fe2+E/g dried weight, respectively, whereas asiatic acid showed no apparent antioxidant capacity. These results led to the second part of the study in which these aqueous extracts and phytochemicals were examined for their neuroprotective properties under in vitro conditions in differentiated PC12 cells against H202 and A~ induced toxicity. The neuroprotective effect varied from plant to plant. An aqueous extract of W somnifera, B. monniera, S. elegans, U tomentosa, C. asiatica and pure compounds asiatic acid and capsaicin showed up to 70-90% protection of PC 12 cells against H202 and A~ induced toxicity whereas, allicin and M ofjicinalis extract showed no protective effects. Anti-aggregation measured by TEM and ThT fluorescence assay showed that an aqueous extract of W somnifera and allicin strongly inhibited fibril formation compared with control samples. These results suggest that the aqueous extract of W somnifera root and allicin have an ability to inhibit formation of mature fibrils which leads to plaque formation. XIV CONCLUSION: These finding demonstrate that that an aqueous extract prepared from these medicinal plants and phytochemicals have significant neuroprotective activities against different targets including AChE, BuChE, ROS and A~ implicated in AD. Hence, these results provide evidence of the usefulness of these medicinal plants and phytochemicals which may be used in the future to develop new therapeutic strategies for the prevention of and treatment of AD.

Thesis MTech (Food Technology))--Cape Peninsula University of Technology, 2017.
Bambara groundnut (Vigna subterranea) an indigenous legume cultivated in Sub-Saharan Africa has been proclaimed to have medicinal properties from communities and in rural areas. However, there is not enough scientific information to validate these claims. Therefore, this study aimed to identify possible medicinal properties of Bambara groundnut (BGN), by analysing the phytochemical and antimicrobial properties of BGN seed and product extracts from Mpumalanga province within South Africa. The BGN extracts (70% methanol, 70% ethanol, milli-Q water) from seeds and products (milk and yoghurt) were screened for the presence of alkaloids, flavonoids, phenols, riboflavin and thiamine using analytical laboratory methods for basic screening, high-performance liquid chromatography (HPLC) and gas chromatography (GC) for quantification. The antimicrobial activity involved direct bioautography and minimum inhibitory concentration (MIC) against six antibiotic-resistant microorganisms, Acinetobacter baumannii ATCC 19606T, Enterococcus faecalis ATCC 29212, Klebsiella pneumoniae subsp. pneumoniae ATCC 700603, Pseudomonas aeruginosa ATCC 27853, Staphylococcus aureus subsp. aureus ATCC 33591 and Candida albicans ATCC 24433. For the seed extracts, flavonoids and phenols were highly concentrated in the red and brown hulls of BGN compared to whole and dehulled BGN. Organic solvents in comparison to water yielded the highest concentration of flavonoids, whilst water yielded the highest concentration for phenols. Flavonoid compounds that were detected at the highest concentrations were rutin (24.458 ± 0.234 mg.g-1, brown hull extracted with 70% methanol), quercetin (0.070 ± 0.043 mg.g-1, red hull extracted with 70% methanol), kaempferol (0.391 ± 0.161 mg.g-1; brown hull extracted with 70% ethanol) and myricetin (1.800 ± 0.771 mg.g-1; red hull extracted with 70% methanol). For phenol compounds, gallic acid (0.009 ± 0.004 mg.g-1; brown hull extracted with milli-Q water), catechin (0.026 ± 0.041 mg.g-1; brown hull extracted with milli-Q water), methyl gallate (0.008 ± 0.013 mg.g-1; brown whole extracted with milli-Q water), chlorogenic acid (0.115 ± 0.199 mg.g-1; brown hull extracted with milli-Q water) and ellagic acid (0.105 ± 0.082 mg.g-1; red hull extracted with milli-Q water) were detected. Vitamins B1 and B2 (riboflavin and thiamine) were mostly present in milli-Q water extracts. Black-eye hull had the highest concentration of thiamine (vitamin B1) and riboflavin (vitamin B2) consisting of 0.072 mg.g-1 (extracted with milli-Q water) and 0.002 mg.g-1 (extracted with 70% ethanol and 70% methanol). Red and brown hull extracts from organic solvents (70% ethanol and 70% methanol) showed the highest antimicrobial activity, whereas the whole, dehulled and hulls (black-eye and brown-eye) extracts had no antimicrobial activity. As for BGN products extracts, flavonoid compounds that were detected at the highest concentrations were rutin (5.694 mg.g-1, whole BGN milk, milli-Q water), quercetin (0.703 mg.g-1, whole BGN yoghurt, milli-Q water) and myricetin (0.987 mg.g-1, whole BGN yoghurt, 70% ethanol).

Thesis (MTech (Horticulture))--Cape Peninsula University of Technology, 2018.
Ticks and tick-borne diseases are important limiting factors to the attainment of sustainable animal and human health, affecting livelihood of resource poor farming communities in developing countries. Rhipicephalus appendiculatus and Amblyomma variegatum (Ixodidae) are among the most troubling tick species in Africa. While ticks can be controlled by applying chemical acaricides, these chemicals are quite expensive, especially, for small-scale famers in developing countries. Hence, the quest for alternative tick control over the years have revealed that entomopathogenic fungi (EPF) and plant extracts have huge prospects as sustainable alternatives for tick control. Beauveria bassiana (Hypocreales) is a fungal entomopathogen with the ability to colonize plants endophytically and induce secondary metabolite production in plants, and it has been found to be a potential biological control agent against a wide range of arthropods. Several plant species including plant species belonging to the family Alliaceae possess anti-tick activities (repellent and toxic); therefore, integrating the two strategies by inoculating Tulbaghia violacea with B. bassiana could enhance secondary metabolite contents in extracts obtained from the plant and increase medicinal materials. The primary purpose of the present study was to evaluate the effect of indigenous endophytic entomopathogens inoculation of T. violacea on the plant growth, tissue nutrient contents and secondary metabolites, and anti-tick activities of extracts of T. violacea. The specific objectives were: (i) to carry out a study on the effects of inoculating T. violacea with fungal strains of B. bassiana and Clonostachys rosea on plant growth and tissue nutrient contents in extracts of T. violacea with the view of selecting one of the two fungi for further investigation (ii) to compare secondary metabolite profiles of extracts obtained from plants exposed to fungus (EPF) inoculum and control treatment during cultivation, and (iii) to determine whether exposure of T. violacea to an endophytic fungus (B. bassiana) during cultivation affects the repellent and acaricidal activities of extracts of T. violacea on A. variegatum and R. appendiculatus. The study had two experiments, presented in chapters two and three. In the first experiment, eight weeks old potted seedlings of T. violacea were inoculated separately with B. bassiana (strain SM3) and C. rosea (strain SM8) conidia suspended at concentrations of 1 x 106 conidia mL-1. Plant growth parameters, such as number of leaves, plant height (aerial part), fresh weights of aerial parts, and tissue nutrient contents were assessed. Results indicated that B. bassiana induced higher growth of plants than C. rosea. Inoculation with B. bassiana did not significantly (P ˃ 0.05) influence most of the growth parameters, number of leaves, fresht weight of roots and of fresh weight aerial parts (leaves) of T. violacea assessed in the current study. However, mean plant heights and root lengths were significantly (P < 0.05) higher in favour of B. bassiana treated plants compared to those in C. rosea. Fe contents in the roots (1416.3 ± 305.10 mg/kg) were found to be positively influenced (P < 0.05) by the fungal inoculation. There was a significant difference in roots (P < 0.05) on the uptake of Mn in C. rosea treated plants (243 ± 19 mg/kg) compared to the control group (169 ± 16.37 mg/kg) and B. bassiana treated roots (161.3 ± 14.44 mg/kg). Macro nutrients up take did not differ significantly among treatments on both leaves and roots (P > 0.05).

Dissertação de Mestrado Integrado em Medicina Veterinária
Cytochrome P450 enzymes (CYP) are hemoproteins belonging to the group of monooxygenases and one of the main enzymatic systems responsible for drug metabolism. In the present study, in vitro approach was applied to evaluate the relation of CYP-catalyzed activities between human, rabbit, minipig and mouse, using single substrate assays (MultiCYP 7-ethoxycoumarin 0-deethylase (ECOD), CYP1A1/2 7-ethoxyresorufin 0- deethylase (EROD), CYP2A6 coumarin 7-hydroxylase (COH), CYP3A4 midazolam 1- hydroxylase (OH-MDZ), and CYP2E1 chlorzoxazone 6-hydroxylase (OH-CLZ)). It was also studied plant extracts (Pinus sylvestris, Angelica archangelica, Mentha sp., Citrus grandis) and phytochemicals (8-Hydroxybergapten, 5,6-dihydroxyangelicin, α, β-Thujone, α-Thujone, angelicin, bergamottin, bergapten, bergaptol, cnidilin, imperatorin, isobergapten, isopimpinellin, lanatin, phellopterin, psoralen, sphondin, xanthotoxin) as potential inhibitors in CYP-related activities of hepatic human microsomes (CYP1A1/2 (EROD), CYP2A6 (COH), CYP3A4 (OH-MDZ)). This study showed that the lowest ECOD activity was detected in humans and there was no similarity between other species. CYP1A1/2 showed equivalent activities. The highest CYP activities in humans were found for CYP2A6 and CYP3A4. In CYP2E1 activity, two similar groups were recognized: human and mouse versus rabbit and minipig. EROD reaction was the most inhibited CYP-mediated reaction. COH reaction was inhibited by few compounds. The highest inhibition was detected among angular furocoumarins. Linear furocoumarins group had the lower inhibitory concentration of CYP3A4. Thujone showed weak inhibition of CYP activities.
RESUMO - As enzimas do sistema citocromo P450 (CYP) são hemoproteinas pertencentes ao grupo das monoxigenases e um dos principais sistemas enzimáticos responsáveis pela metabolização de fármacos. Neste estudo foi avaliada a relação da actividade catalítica de diferentes CYPs entre humanos, coelhos, minipig e murganhos, recorrendo a substratos como sondas individuais in vitro para mensurar reacções especificas (MultiCYP 7-etoxicumarina 0-deetilase (ECOD), CYP1A1/2 7-etoxiresorufina 0-deetilase (EROD), CYP2A6 cumarina 7-hidroxilase (COH), CYP3A4 midazolam 1-hidroxilase (OH-MDZ), e CYP2E1 clorozoxazona 6-hidroxilase (OH-CLZ)). Também foram estudados extractos de plantas (Pinus sylvestris, Angelica archangelica, Mentha sp., Citrus grandis) e fitoquímicos (8-hidroxibergaptem, 5,6-dihidroxiangelicina, α, β-tujona, α-tujona, angelicina, bergamottin, bergapteno, bergaptol, cnidilina, imperatorina, isobergapteno, isopimpinelina, lanatin, felopterina, psoraleno, sphondin, xantotoxina) como potenciais inibidores da actividade catalítica dos CYPs microssomais hepáticos humanos (CYP1A1/2 (EROD), CYP2A6 (COH), CYP3A4 (OH-MDZ)). Neste estudo não foram detectadas actividades similires entre espécies na reacção ECOD e a actividade mais baixa foi detectada nos humanos. A reacção EROD dos CYP1A1/2 demonstrou actividades similares entre as diferentes espécies. As maiores actividades cataliticas verificadas nos humanos correspondem aos CYP2A6 e CYP3A4. No estudo da reacção do CYP2E1 foram determinados dois grupos distintos com actividades cataliticas similares: 1) humanos e murganhos, 2) coelhos e minipigs. No estudo de potenciais inibidores dos CYPs, a reacção EROD foi a mais inibida. Pelo contrário, a reacção COH foi inibida por poucos compostos. A maioria das inibições ocorreu por exposição a furocumarinas angulares. O grupo de furocumarinas lineares teve a menor concentração inibitória da reacção OH-MDZ do CYP3A4. Os fitoquímicos α, β-tujona e α- tujona demonstraram ter um fraco poder inibitório na actividade dos CYPs analisados.

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Sedum praealtum DC. (Crassulaceae) is one of 350 species pharmacologically active from the genus Sedum, whose actions in treatment of eyes (pain and swelling) and ulcer, inflammatory problems, as contraception and anti-fertilization, antinociceptive and anti-inflammatory have been reported. The objective was to evaluate the hydroethanolic extract of S. praealtum regarding their potential antimicrobial in vitro (some bacteria, yeasts and micobactéria strains), cytotoxic in vitro and genotoxic in vivo. A fast phytochemical screening of this extract was also performed. The antimicrobial activities were carried out by microdilution in broth and agar diffusion methods (CLSI). The genotoxic effects and systemic toxic and cytotoxicity were evaluated by micronucleus assay in mice bone marrow and cell cultures of Aedes albopictus, respectively. The selectivity index was also established (SI = CI50/MIC). Dosages of flavonoids and phenolic compounds were done by colorimetric and precipitation techniques. A high amount of phenolic compounds were identified in S. praealtum root. The S. praealtum leaves showed broad spectrum of action and variables MICs: Gram-negative bacteria (E. aerogenes, E. coli, P. aeruginosa, P. mirabilis, S. marcescens and S. typhimurium), gram-positive (B. cereus, B. subtilis, E. faecalis, M. luteus and S. aureus) and yeast (S. cerevisiae). The stem and root were restricted to gram-positive bacteria and S. cerevisiae, other than E. coli (stem) and P. mirabilis (root) microbicidal action microorganism- and anatomical part-dependent (leaf, stem or root). S. praealtum showed no inhibition against C. albicans, M. tuberculosis and M. bovis. The root showed acceptable SI (SI  1) for P. mirabilis; B. subtilis; B. cereus; M. luteus; E. faecalis; S. aureus and S. cerevisiae, whereas the sheet only for S. cerevisiae. The hydroalcoholic extract of S. praealtum leaves revealed no genotoxic effects (no clastogeny and/or aneugeny) and toxicity in bone marrow of mice, dose (0.5-2 g.Kg-1) and time-independent (24-48 hours), but sex-dependent (male and female). This was the first scientific study of this nature involving S. praealtum and partially the results provide a theoretical basis for comprehensive development and utilization of plant resources. However, advanced phytochemical characterization together with the various pharmacological and pharmacogenomic studies should be conducted in order to characterize their effects and, importantly, for the establishment of limits for human consumption, the delineation of potential risks to human health, and for rational strategies for implementing chemo-preventive measures.
Sedum praealtum DC. (Crassulaceae) é uma das 350 espécies farmacologicamente ativas do gênero Sedum, cujas ações no tratamento dos olhos (dores e inchaços) e úlcera, de problemas inflamatórios, como contraceptivo e antifertilização, antinociceptiva e anti-inflamatória foram relatadas. O objetivo foi avaliar o extrato hidroetanólico de S. praealtum quanto aos seus prováveis potenciais antimicrobiano in vitro de algumas cepas de bactérias, de leveduras e de micobactérias, citotóxico in vitro e genotóxico in vivo. Uma rápida triagem fitoquímica desse extrato também foi realizada. As atividades antimicrobianas foram realizadas empregando-se os métodos de microdiluição em caldo e em difusão em agar (CLSI). Os efeitos genotóxicos e tóxicos sistêmicos e a citotoxicidade foram avaliados pelo ensaio do micronúcleo na medula óssea de camundongos e pelas culturas celulares de Aedes albopictus, respectivamente. O índice de seletividade também foi estabelecido (IS = CI50/CIM). Dosagens de flavonoides e compostos fenólicos foram feitas usando técnicas colorimétricas e de precipitação. Uma elevada quantia de compostos fenólicos foi identificada na raiz de S. praealtum. As folhas de S. praealtum mostraram ação de amplo espectro e CIM variáveis: bactérias gram-negativas (E. aerogenes, E. coli, P. aeruginosa, P. mirabilis, S. marcescens e S. typhimurium), gram-positivas (B. cereus, B. subtilis, E. faecalis, M. luteus e S. aureus) e levedura (S. cerevisiae). O caule e a raiz foram restritos às bactérias gram-positivas e S. cerevisiae, exceto E. coli (caule) e P. mirabilis (raiz) ação microbicida micro-organismo-dependente e parte anatômica-dependente (folha, caule ou raiz). S. praealtum não apresentou ação contra C. albicans, M. tuberculosis e M. bovis. A raiz mostrou IS aceitável (IS  1) para P. mirabilis; B. subtilis; B. cereus; M. luteus; E. faecalis; S. aureus e S. cerevisae, enquanto que a folha apenas para S. cerevisae. O extrato hidroalcoólico das folhas de S. praealtum revelou efeitos não genotóxicos (ausência de clastogenia e/ou aneugênia) e efeitos tóxicos na medula óssea de camundongos, dose- (0,5-2 g.Kg-1) e tempo-independente (24-48h), porém sexo-dependente (macho e fêmea). Este foi o primeiro estudo científico dessa natureza envolvendo S. praealtum e, parcialmente, os resultados fornecem uma base para a utilização e para o desenvolvimento compreensivo de recursos vegetais. Todavia, a caracterização fitoquímica avançada aliada aos diversos estudos farmacológicos e farmacogenômicos deveriam ser conduzidos a fim de caracterizar os seus efeitos e, mais importante, estabelecer limites para o consumo popular, delinear os riscos potenciais à saúde humana, e implementar estratégias racionais e medidas quimio-preventivas.

O gênero Salmonella constitui um problema para a saúde animal e humana. Com a finalidade de evitar a ocorrência da transmissão deste microrganismo e a interrupção da doença provocada por eles, deve-se utilizar desinfetantes e antibióticos. A exposição continuada a estes agentes pode resultar no surgimento de resistência microbiana. Tendo em vista este fenômeno, a pesquisa por novos agentes antimicrobianos a partir de extratos vegetais se faz necessária, pois existe a possibilidade de se encontrar substâncias eficazes contra a resistência de micro-organismos já disseminados no ambiente. Os objetivos deste estudo foram: monitorar a resistência das 134 amostras de Salmonella isoladas em produtos de origem animal frente a antibióticos comumente utilizados na rotina veterinária; testar a atividade dos desinfetantes frente às Salmonellas resistentes; testar a hipótese de resistência cruzada entre grupos antibióticos e desinfetantes; avaliar a atividade bactericida de extratos de plantas medicinais nativas no Rio Grande do Sul sobre salmonelas padrão; avaliar a atividade antibacteriana do macerado hidroalcoólico de Achyrocline satureioides frente às salmonelas resistentes; fazer prospecção fitoquímica do macerado hidroalcoólico e do decocto de Achyrocline satureioides, Realizou-se o teste de sensibilidade a antibióticos frente a 134 amostras de Salmonella spp. isoladas em produtos de origem animal. Cinquenta e uma (51) salmonelas foram resistentes a pelo menos um antibiótico. Após, utilizou-se o teste de suspensão da avaliação quantitativa da atividade bactericida de desinfetantes. Foram testados os desinfetantes cloreto de cetil trimetilamônio (amônia quaternária), digluconato de clorexidina (clorexidina), hipoclorito de sódio e iodóforo, em quatro concentrações frente salmonelas resistentes, nos tempos 5, 15 e 60 minutos, densidade populacional 107 UFC/ mL. O iodóforo e o hipoclorito de sódio inativaram os isolados na maior parte dos experimentos. Porém, cloreto de cetil trimetilamônio (amônia quaternária) e digluconato de clorexidina (clorexidina) mesmo nas maiores concentrações do desinfetante, houveram micro-organismos resistentes. Pode-se perceber que não houveram relações concomitantes de resistência entre antibióticos e desinfetantes, levando-se em consideração todas as variáveis testadas. Na triagem inicial da atividade antibacteriana das plantas medicinais, utilizou-se a proporção de 10 g de planta para 100 mL do líquido extrator, em 8 e 24 horas, densidade populacional de Salmonella Choleraesuis (ATCC 10.708) 107, 106 e 105 UFC/ mL e formas decocção e maceração hidroalcoólica das plantas Achyrocline satureioides, Bidens pilosa, Conyza bonariensis, Jacaranda micrantha, Myrciaria cuspidata e Smilax cognata. A. satureioides, como macerado hidroalcoólico apresentou inativação nos tempos 8 e 24 horas, mostrando maior redução logarítmica. A. satureioides no macerado hidroalcoólico foi testada frente a 51 salmonelas resistentes. Observou-se que quanto maior o tempo de contato, nas menores densidades populacionais, maior foi o número de inativações. A atividade antibactericida do macerado hidroalcoólico de A. satureioides sugere o seu uso como ingrediente antisséptico em pomadas para uso animal ou humano, ou ser empregado na imersão de utensílios e partes removíveis de maquinários das indústrias alimentícias, frigoríficos, tendo como objetivo promover a desinfecção destes insumos. A prospecção fitoquímica realizada com o decocto e com o macerado hidroalcoólico de A. satureioides detectou a presença de compostos fenólicos, taninos hidrolisáveis e condensados, flavonóis e saponinas. O decocto de A. satureioides, diferentemente do macerado hidroalcoólico, não apresentou taninos hidrolisáveis e saponinas.
The genus Salmonella constitutes a problem for animal and human health. In order to prevent the transmission of this organism and the interruption of the disease caused by them, you should use disinfectants and antibiotics. Continued exposure to these agents can result in the emergence of microbial resistance. In view of this phenomenon, the search for new antimicrobials from plant extracts is necessary, because there is a possibility to find effective substances against the resistance of microorganisms already disseminated in the environment. The objectives of this study were: to monitor the resistance of 134 samples of Salmonella isolated in products of animal origin in front of antibiotics commonly used in veterinary routine; test the activity of disinfectants against the salmonellas resistant; test the hypothesis of cross-resistance between antibiotics and disinfectants; evaluate the bactericidal activity of extracts of medicinal plants native to the Rio Grande do Sul on Salmonella standard; evaluate the antibacterial activity of hydroalcoholic macerate to Achyrocline satureioides to Salmonella resistant; do the hydroalcoholic macerate phytochemical prospecting and to Achyrocline satureioides vegetable water, antibiotic sensitivity testing in front of 134 samples of Salmonella spp. isolated in products of animal origin. Fifty-one (51) Salmonella were resistant to at least one antibiotic. After, we used the suspension test of quantitative evaluation of bactericidal activity of disinfectants. Have been tested the disinfectants cetyl trimetilamônio chloride (quaternary ammonia), chlorhexidine digluconate (chlorhexidine), sodium hypochlorite, iodophor in four concentrations Salmonella resistant front, 5 times, 15 and 60 minutes, population density 107 CFU/ mL. The iodophor and sodium hypochlorite inactivated isolates in most experiments. However, cetyl trimetilamônio chloride (quaternary ammonia) and chlorhexidine digluconate (chlorhexidine) even in the largest concentrations of disinfectant, there were resistant microorganisms. One can notice that there were no concurrent relationships of resistance between antibiotics and disinfectants, taking into account all the variables tested. In the initial screening of antibacterial activity of medicinal plants, the proportion of 10 g of plant for 100 mL of liquid Extractor, in 8 and 12:00 hour, population density of Salmonella Choleraesuis (ATCC 10,708) 107, 106 and 105 CFU/ mL decoction and hydroalcoholic maceration and the plants Achyrocline satureioides, Bidens pilosa, Conyza bonariensis, Jacaranda micrantha, Myrciaria cuspidata and Smilax cognate. A. satureioides hydroalcoholic macerate as presented 8 and 12 hour in inactivation, showing greater logarithmic reduction. A. satureioides in hydroalcoholic macerate has been tested in front of Salmonella resistant 51. It was observed that the longer the time of contact, the smallest population densities, higher number of inactivation. The antibacterial activity of hydroalcoholic macerate of A. satureioides suggests its use as an antiseptic ingredient in ointments for human or animal use, or be employed in the immersion of utensils and removable parts of machinery of food industries, refrigerators, aiming to promote the disinfection of these inputs. The phytochemical held prospecting with the vegetable water and hydroalcoholic macerate of A. satureioides detected the presence of phenolic compounds, hydrolysable and condensed tannins, flavonols and saponins. The vegetable water of A. satureioides, unlike the hydroalcoholic macerate, hydrolysable tannins and failed to provide saponins.

M.Sc. (Chemistry)
The extraction of the atropine and scopolamine alkaloids from both Datura ferox and Datura stramonium was carried out in this study to investigate potential of these plants as a source of these alkaloids. Tropane alkaloids such as atropine occur naturally in many members of several plant families, especially the Solanaceae, Erythroxylaceae, Proteaceae and Convolvulaceae families. Hyoscyamine or its racemic mixture atropine is used as an antidote to organophosphate insecticides, or as a nerve agent to dilate the pupil for retina examinations. Scopolamine or hyoscine is used to treat motion sickness, among other uses. Tropane alkaloids are often difficult to synthesise, and their extraction from commercial sources is therefore of some commercial interest. A systematic study was carried by comparing different extraction methods for the efficiencies of tropane alkaloid (TA) extraction at ambient pressure. The two plants selected are already widely spread as weeds in many regions in South Africa and are therefore well adapted to the local conditions. Among the extraction methods, TA’s were extracted in their acidic form using dilute aqueous acids with 5% sulfuric acid giving the highest yield of both atropine and scopolamine. Alternatively, the TA’s were extracted in their basic form from the plant material using basified organic solvents. The overall yields obtained were not significantly different from those obtained by using dilute aqueous acids, with D. ferox giving approximately 0.24 % and 0.0028 % scopolamine and atropine, respectively. D. stramonium on the other hand gave approximately 0.037 % and 0.002% atropine and scopolamine, respectively. Different plant parts were also tested in order to determine if there was an advantage in extracting the tropane alkaloids at a particular time in the growth, but concluded that the best option was to extract the seeds for various reasons such as the presence of relatively less chlorophyll...

MSc (Microbiology)
Department of Microbiology
Background: The medicinal value of plants lies in some chemical substances that produce a definite physiological action in the human body. The secondary metabolites help the plants to survive hash conditions and could be used by humans as supplements of their health, as foods additives or for medicinal purposes. This bioactive compounds are not always beneficial to human beings, and some of this plants bioactive compounds can be toxic or genotoxic to human cells. This study used several methods to evaluate of phytochemical constituents and mutagenic properties of Coccinia rehmanni and Jatropha zeyheri plant extracts. Methodology: Methanol was used for extraction of the bioactive compounds from the two selected plants, filtered with Whatman filter paper and evaporated with rotary evaporator. The extracts were fractionated using open column chromatography. Chemical and TLC methods were used to determine phytochemicals of the study plants extracts and fractions. The plants extracts and fractions were tested against Vero cell lines in order to evaluate cytotoxicity and genotoxicity of the plants. NucRed and LTR Hoechst 33342 dyes were used for cytotoxicity and genotoxicity respectively. For the evaluation of cytotoxicity and genotoxicity Quantification of live and dead cells for the screening assay was performed using the ImageXpress Micro XLS Widefield Microscope and acquired images analyses using the MetaXpress software and Multi-Wavelength Cell Scoring Application Module. Antimutagenicity of plants extracts was observed using PARP universal colorimetric assay kit. Acquired data was transferred to an EXCEL spreadsheet and data was analyzed. Results and discussion: C. rehmanni (12.03%) yielded more extract than J. Zeyheri (8.20%). the two plants had different compound composition and were in different stages of maturity. The study revealed the domination of Terpenoids, Cardiac glycosides, Phenolic and tannis. With an exception of two fraction fractions all the fractions was found to be toxic to an extent were genotoxicity of such fraction could not be concluded. The reason for such extreme toxicity could be due to the influence of the retained alcohol during rotary evaporation. xvi | P a g e Conclusion: this study provides and add to existing knowledge on the phytochemicals mutagenicity and anti-mutagenicity of C. rehmanni and J. Zeyheri medicinal plants. The study serves as scientific proof that extensive use of this plant in traditional medicine for treatment of various ailments may lead to some irreversible damages.
NRF

Dissertação de Mestrado em Biologia
As doenças neurodegenerativas têm sido um foco da ciência com o objetivo de desenvolver novas estratégias neuroprotetoras capazes de prevenir a morte neuronal associada a estas patologias. A identificação de plantas medicinais tem sido alvo de vários investigadores com a finalidade de encontrar novas fontes de compostos com propriedades farmacológicas. O Brasil, nomeadamente o bioma Caatinga, é rico em plantas medicinais sendo estas utilizadas pelos habitantes locais na medicina tradicional. Tendo isto em conta, os objetivos deste trabalho foram: 1) caracterizar o perfil fitoquímico e conteúdo em compostos fenólicos, de extratos provenientes de plantas presentes neste bioma; 2) avaliar as propriedades antioxidantes in vitro destes extratos; 3) explorar o potencial antioxidante e neuroprotetor destes extratos em modelo celular. Os extratos das folhas das plantas selecionadas para este estudo mostraram (HPLC-DAD) elevados níveis de compostos fenólicos, nomeadamente ácido gálico, caempferol, quercetina e seus derivados, catequinas entre outros. Extratos provenientes de Sp 3 (Caesalpinia ferrea), 4 (Anadenanthera peregrina), 6 (Mimosa tenuiflora) e 9 (Schinopsis brasiliensis) revelaram o maior conteúdo em compostos fenólicos. Tendo em conta os EC50 obtidos para todos os ensaios, os extratos provenientes de Sp 3 e 9 mostraram a maior capacidade scavenging de DPPH; 4 e 18 (Cleome spinosa) mostraram a maior capacidade quelante de ferro; 4, 6 e 14 (Capparis flexuosa) revelaram a maior capacidade de inibição de produção de NO; e 6 e 3 mostraram a melhor capacidade scavenging de superóxido. Portanto, Sp 3, 4, 6, 9, 14 e 18 parecem ser os mais promissores. Assim, observou-se que as propriedades antioxidantes demonstradas por estes extratos poderão estar relacionadas com o seu conteúdo em compostos fenólicos. Revelou-se importante estudar as propriedades neuroprotetras destes extratos usando o modelo neuronal U251, exposto ao agente oxidante t-BHP, de modo a induzir stress oxidativo nestas células. Os resultados revelaram que os extratos que se mostraram ser mais promissores nos ensaios anteriores, nomeadamente no conteúdo em compostos fenólicos e potencial antioxidante demonstrado, Sp 3, 4, 6 e 9, induziram maior protecção nas células, em condições de stress oxidativo. Estes resultados sugerem assim que, provavelmente, o elevado conteúdo em compostos fenólicos poderá induzir, por algum mecanismo, a proteção celular. O passo seguinte consistiu em avaliar quais as condições de incubação que poderiam potenciar o efeito neuroprotetor dos extratos. Os resultados mostraram que, para a maioria dos extratos, nomeadamente, Sp 3, 4 e 6, a pré-incubação destes (em que o extrato atua sozinho) seguido da co-incubação do agente oxidante t-BHP, potencia o efeito neuroprotetor, o que sugere uma ação profilática, isto é, o extrato atua como medida preventiva contra o dano. Portanto, a proteção celular é aumentada quando fornecemos às células os compostos bioativos, que provavelmente induzem o aumento de defesas nas células, antes do dano oxidativo ocorrer. Assim, os nossos estudos suportam a ideia de que as plantas do bioma Caatinga têm diversas potencialidades podendo assim ser usadas como fonte de compostos com propriedades farmacológicas, sendo que os extratos estudados parecem possuir propriedades antioxidantes e neuroprotetoras contra processos neurodegenerativos associados a stress oxidativo.
Neurodegenerative diseases have been a focus of science with the aim of developing new neuroprotective strategies capable of preventing neuronal death associated with these pathologies. The identification of medicinal plants has been the target of various researchers in order to find new sources of compounds with pharmacological properties. Brazil, namely Caatinga biome, is rich in medicinal plants and these are used by locals in traditional medicine. Concerning this, the objectives of this study were: 1) characterize the phytochemical profile and content in phenolic compounds, from plant extracts present in this biome; 2) evaluate the in vitro antioxidant properties of these extracts; 3) explore the antioxidant and neuroprotective potential of these extracts on cellular model. The plant leaves extracts selected for this study showed (HPLC-DAD) high levels of phenolic compounds, including gallic acid, kaempferol, quercetin and derivatives, catechins and others. Extracts from Sp 3 (Caesalpinia ferrea), 4 (Anadenanthera peregrina), 6 (Mimosa tenuiflora) and 9 (Schinopsis brasiliensis) revealed the highest content in phenolic compounds. Given the EC50 obtained for all asays, extracts from Sp 3 and 9 showed the greatest ability to DPPH scavenging; 4 and 18 (Cleome spinosa) showed the greatest ability to chelate iron; 4, 6 and 14 (Capparis flexuosa) showed the greatest capacity of inhibiting NO production; and 6 and 3 showed better superoxide scavenging capacity. So, Sp 3, 4, 6, 9, 14 and 18 seems to be the most promising. It was found that the antioxidant properties shown by these extracts may be related to its content in phenolic compounds. It has important to access the neuropropertive properties of these extracts using a neuronal model U251 exposed to the oxidant t-BHP, to induce oxidative stress in these cells. The results revealed that the extracts that proved to be most promising in previous studies, including the content of phenolics and antioxidant activity demonstrated, namely Sp 3, 4, 6 and 9, induced protection in conditions of oxidative stress. These results suggest that, probably, the high content of phenolic compounds may induce, by some mechanism, cell protection. The next step was to evaluate which incubation conditions could potentiate the neuroprotective effect of the extracts. The results showed that for the most of the extracts, namely, Sp 3, 4 and 6, the pre-incubation of these (where the extract acts alone) followed by co-incubation of t-BHP, oxidizing agent, enhances the neuroprotective effect, suggesting a prophylactic action, i.e., the extract serves as a preventative measure against damage. Therefore, the cell protection is increased when we provide the bioactive compounds to the cells, which probably induce increased the cell defenses, before oxidative damage occurs. Thus, our studies support the idea that Caatinga biome plants have several different potentialities that can be used as a source of compounds with pharmacological properties, and these extracts seems to possess antioxidant and neuroprotective properties against neurodegenerative diseases associated with oxidative stress.

Medicinal plants are the focus of intense study, in particular whether their traditional uses are supported by real pharmacological effects, or merely based on folklore. Piper capense L.f. (Piperaceae) is used traditionally for the treatment of infectious diseases, and has the potential to be a source of novel antimicrobial compound(s). Crude solvent extracts (water, methanol, hexane and acetone) and sequentially extracted subfractions of the root-bark of P. capense were prepared, of which the hexane-soluble subfraction MsAsHs was identified as the most promising antimicrobial subfraction. Phytochemical analyses of the various extracts and subfractions using TLC with numerous mobile phases and compound selective visualising reagents revealed the presence of quinones in all of the crude solvent extracts. Alkaloids, lipids/sterols/steroids, phenolic compounds and amino acids/peptides were detected in select subfractions. Gradient reverse phase HPLC analyses using 0.1% formic acid and methanol indicated three major peaks in MsAsHs. IR spectroscopy indicated that carbonyl and hydroxyl functional groups, and aromatic characteristics were present in the major compound present in MsAsHs. Further analysis using targeted LC-MS Q-TOF and quadrupole LC-MS/MS analyses indicated an empirical formula of C11H8O3. This formula was confirmed for the isolated compound by GC-MS (HP5-MS column) that identified the compound as 5-hydroxy-2-methyl-1,4-naphthoquinone (C11H8O3 MW: 188.18) with 98% certainty using the database. Although 5-hydroxy-2-methyl-1,4-naphthoquinone (also known as plumbagin) is well-known, this is the first time that the presence of this compound is reported in the Piper genus. Antimicrobial activities of P. capense root-bark extracts and the subfractions were determined against Gram-negative and Gram-positive bacteria and a yeast strain using the disk diffusion and broth micro-dilution assays. Antimicrobial activity was observed against Gram-positive bacteria, Gramnegative bacteria as well as a yeast strain, indicating broad spectrum activity. The antimicrobial activities of the crude solvent extracts decreased in the order: acetone > methanol > hexane > water. The MsAsHs subfraction demonstrated the highest antimicrobial activity with an MIC of 29 μg/ml against both Staphylococcus aureus (ATCC 12600) and Candida albicans (ATCC 10231). HPLC eluents of this subfraction that were collected in a drop-wise fashion onto silica TLC plates and assayed by bioautography, indicated that the major compound eluting at 13.6 minutes accounted for most of the antimicrobial activity. Antioxidant activity was observed for the crude water extract, crude methanol extract, crude acetone extract, MsAsAs subfraction as well as the MsAsHs subfraction. Cytotoxicity against mammalian cells in culture was observed for the crude methanol extract, crude acetone extract, crude hexane extract and the MsAsHs subfraction when determined using C2C12 cells as well as resting and PHA stimulated lymphocytes. Stability testing of the MsAsHs subfraction revealed that the antimicrobial compounds found in this subfraction appear to be stable up to 30 days at both 25°C and 40°C when assayed against S. aureus. However, when assayed against C. albicans, there was an increase in antifungal activity from 29 μg/ml to < 7 μg/ml after 30 days at both temperatures tested. This study provides scientific support for the ethnomedical use of the rootbark of P. capense as an antimicrobial. To date, the presence of plumbagin has not been reported in any other plant in the Piper genus. Due to the significant cytotoxic activity against mammalian cells reported in the current study and the mechanism of action of plumbagin, the therapeutic potential of P. capense extracts is very limited due to non-selective cytotoxicity, despite its marked antimicrobial activity.
Dissertation (MSc)--University of Pretoria, 2011.
Pharmacology
unrestricted

The use of plants for cosmeceutical applications is becoming more important since “safer” and more “natural” skin products are gaining popularity. The effect of different drying methods and storage conditions on metabolite changes and biological activity of two species with cosmeceutical application namely Greyia flanaganii and Leucosidea sericea were investigated using 1H-NMR metabolomics. The multivariate analysis (PCA and OPLS-DA), and 1H-NMR sample spectra were used to analyse the significant differences (P<0.05) resulting from the different treatments. The effect of these treatments on anti-tyrosinase and anti-bacterial (against Staphylococcus aureus) activity of G.flanaganii and L.sericea ethanol leaf extracts respectively, was further investigated to assess the quality. Four different drying methods adopted were freeze drying, oven drying at 50°C, air drying at room temperature and sun drying in a greenhouse. The dried leaf extracts were stored in three different conditions of fridge, freezer and shade conditions and samples from each storage condition taken for analysis at three and six months of storage. The chemical constituents of the leaf extracts of both species were not affected by the drying method and the storage condition, but the concentrations of the metabolites changed. The treatments did not show a significant difference (P<0.05) on the biological activity of the extracts. However, G.flanaganii plant material harvested from the University of Pretoria exhibited a higher anti-tyrosinase activity than material harvested from Mothong heritage site. In G. flanaganii freshly freeze and oven dried extracts exhibited a higher anti-tyrosinase activity with fifty percent inhibitory (IC50) activity of 16.8±0.69 μg/ml and 15.73±0.85 μg/ml respectively than the activity of sun and air dried with IC50 values of 33.08±0.78 μg/ml and 36.86±2.01 μg/ml respectively. The metabolite concentrations and anti-tyrosinase activity dropped significantly after storage. Leucosidea sericea oven and freeze dried extracts, exhibited good anti-bacterial activity with a minimum inhibitory concentration (MIC) value of 0.25 μg/ml and 0.125 μg/ml respectively as compared to sun and air dried extracts with the same MIC value of 0.5 μg/ml. Freeze dried samples showed the best anti-bacterial activity (MIC 0.125 μg/ml) compared to other drying methods. Fridge and freezer storage conditions enhanced the activity of stored sample.
College of Agriculture and Environmental Sciences
M. Sc. (Agriculture)

A breast cancer cell line stably transfected with the CYP19 gene had been employed for aromatase inhibition. Among the phytochemicals tested, the major dietary flavonoids, such as genistein and daidzein, produced very weak inhibition. On the other hand, the red clover isoflavone biochanin A, the hydroxychalcone butein and the red grape phytoalexin resveratrol were found to be effective aromatase inhibitors. Cell proliferation assay had shown that they could inhibit ER-positive cell proliferation induced by testosterone, and the inhibitory effect was specifically attributed to the reduction of estrogen synthesis. In another breast cancer cell line SK-BR-3, resveratrol, biochanin A and genistein inhibited CYP19 both in enzyme and promoter I.3/II transcriptional levels. The element responsible for the inhibition of aromatase by these phytoestrogens should fall within the region between -556 to -446 by upstream of exon II.
Breast cancer is one of the most common cancers affecting women. Estrogen plays an important role in breast cancer initiation and development. The majority of breast tumors are initially dependent upon estrogen to support their growth. Most breast cancers occur in the postmenopausal period. However, the intra-tumoral estradiol (E2) is maintained at a high level equivalent to the pre-menopausal status. High intra-tumoral E2 level in postmenopausal women is sustained by the biosynthesis of estrogens in the tumorous tissue.
Genistein and Biochanin A, ranged from 0.1 to 10 muM, might act as estrogen agonist and induced aromatase activity and promoter I.1 transactivation in ERalpha-transfected SK-BR-3 cells. (Abstract shortened by UMI.)
The aromatase enzyme, CYP19, belongs to a family of P450 enzyme. As a final rate-limiting step in estrogen biosynthesis, it catalyzes the conversion of C 19 steroids to estrogens. The expression of CYP19 is tissue-specific, and is regulated by alternate promoter usage. The use of aromatase inhibitors for breast cancer treatment has become a major therapeutic approach.
The consumption of some phytochemicals protects against breast cancer. Yet the mechanisms are far from clear. In my present study, various phytochemicals, including phytoestrogens, monoterpenes and carotenoids, were evaluated for their effect on aromatase.
Wang Yun.
"July 2005."
Adviser: Lai-Kwok Leung.
Source: Dissertation Abstracts International, Volume: 67-07, Section: B, page: 3716.
Thesis (Ph.D.)--Chinese University of Hong Kong, 2005.
Includes bibliographical references (p. 145-169).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Abstract in English and Chinese.
School code: 1307.

Throughout the Mediterranean region, V. vinifera are subjected to a wide range of environmental stresses, especially during the summer, when prolonged dry periods are combined with high radiation load and high temperatures. Furthermore, given the strong influence of the atmospheric factors on this species, seasonal weather pattern can significantly affect grapevine physiology and wine quality under future climate change projections. Although grapevines have multiple acclimation strategies to environmental constrains, the evidence for significant climate change in the upcoming decades urges adaptation and mitigation measures to be taken by the whole viticulture and winemaking sectors. Short‐term adaptation measures can be considered as a first protection strategy and should be focused at specific threats, mostly changes in agronomical practices, such as the application of biostimulants to enhance nutrition efficiency, abiotic stress tolerance and quality traits of plants. Seaweed-based extracts have been recently employed as sustainable tools to improve abiotic stress tolerance and increase grape quality. However, the effect of these extracts on secondary metabolism compounds, that are fundamental for grape and wine quality, is still scarce. Under this scenario, the objectives of the present PhD project were: 1) to investigate the physiological performances of V.vinifera and the biochemical adjustments induced in grape berry skins by A. nodosum foliar treatments under field conditions; 2) to evaluate if these treatments affect grapevine water relations and berry phenylpropanoid metabolism, mitigating the effects of a post-véraison drought in potted vines. An enhancement in the biosynthesis of secondary metabolites in berry skins and in leaves in response to these treatments, effective to improve grape quality and help vines to cope with abiotic stresses, was hypothesized. In order to achieve these goals, two experiments were arranged. The comparison between vines treated with A. nodosum extract and non-treated ones was set up in a field experiment carried out in 2016 and 2017 in a commercial vineyard in the Chianti Classico area (Tuscany, Italy), on 18-year-old vines of the red cv. Sangiovese (V. vinifera). Furthermore, in order to deeply investigate if A. nodosum treatments are effective in help vines to cope with drought stress, a second study was performed in 2017 on potted vines (cv. Pinot noir), subjected to two irrigation regimes (well watered and water stressed) associated with A. nodosum foliar treatments, performed with the same protocol as the field experiment. In both experiments, gas exchanges, chlorophyll fluorescence and water potentials on leaves treated with A. nodosum extract and non-treated control leaves, were monitored at three phenological stages. In addition, at the same stages, anthocyanins, flavonols and hydroxycinnamic acids were quantified in berry skins. In general, this PhD thesis provides evidence of A. nodosum treatments-induced changes in eco-physiological traits and berry skin metabolism of V. vinifera, resulting in different physiological adjustments to counteract environmental stress. Furthermore, in both experiments, the biochemical berry skin analyses revealed that the A. nodosum extract likely acted selectively in the phenylpropanoid pathway. In particular A. nodosum extract may influence anthocyanin biosynthesis by affecting the activity of enzymes involved in the phenylpropanoid pathway, thus resulting in changes in the content of methoxylated compounds in treated vines. Moreover, the increases in the phenolic and flavonoid profiles have a significant effect on enhancing multiple stress tolerance as well as improving plant water status and photosynthetic performances. Overall, these results support previous findings on the beneficial effects of A. nodosum treatments on plant acclimation to stressful environmental condition. For the first time, this study shows the potential effect of seaweed extracts in promoting higher tolerance to stress in vines. Considering the challenges posed by climate change in the Mediterranean basin, the use of seaweed extracts might represent a sustainable tool to mitigate the negative effects caused by increasing severity of drought events, often associated to heat-waves, in the viticulture sector.