Estudos recentes mostram que a reação entre o complexo mer-[RuCl3(dppb)(H2O)] (dppb = 1,4- bis(difenilfosfino)butano) e a 1,2- fenilenodiamina produz os complexos amido- e imino-, com a coordenação ocorrendo nas posições ocupadas pelo cloro e pela água no complexo inicial. Baseando-se em procedimentos da literatura, com algumas modificações realizou-se a reação do complexo mer-[RuCl3(dppb)(H2O)] com catecol (1,2-dihidroxibenzeno) e 3,5-di-tercbutilcatecol, sendo estes compostos fenólicos simples com duas hidroxilas; também estendeu-se o estudo aos flavonoides com a realização da reação entre o complexo mer-[RuCl3(dppb)(H2O)] com a 3-hidroxiflavona. Os compostos foram caracterizados através de espectroscopia de absorção na região do infravermelho, ultravioleta-visível, análise elementar e voltametria cíclica. Foram comparados os comportamentos dos complexos formados e dos ligantes livres. Os comportamentos eletroquímicos dos ligantes 1,2-dihidroxibenzeno e 3,5- di-terc-butilcatecol têm perfis bastante interessantes em relação à influência dos substituintes no anel aromático; o 1,2-dihidroxibenzeno livre apresenta pico de redução e o 3,5-di-terc-butilcatecol não; o complexo com o primeiro ligante não apresenta pico de redução, enquanto que o pico de redução do complexo com o segundo ligante é bastante visível. Os espectros ultravioleta-visível dos complexos formados são semelhantes com bandas muito próximas caracteristicamente na região de 750 nm a 820 nm. O complexo formado com o ligante 3-hidroxiflavona mostra a influência de inúmeros fatores relacionados com a estrutura do flavonóide. / Recent studies have show that the reaction of the compound mer- [RuCl3(dppb)(H2O)] (dppb = 1,4- bis(diphenyphosphino)butane) and the 1,2- phenylenodiamine produces the amido- and imino- compounds, with the coordination occuring at chlorine and water positions in the initial complex. On Base of procedures of the literature with some modifications it was become the reactions of the compound mer-[RuCl3(dppb)(H2O)] with catechol (1,2- dihydroxybenzene) and 3,5-di-terc-butylcatechol, were carried out. The study extended was to the flavonoids with the reaction of the compound mer- [RuCl3(dppb)(H2O)] and to 3-hidroxiyflavon. There were difficulties in the syntheses, satisfactory results of yield of the products. The compounds were characterized by were obtained IR and uv-vis, elementare analysis and cyclic voltammetry. The electrochemical behavior of the free ligands 1, 2-dihydroxybenzene and 3,5-di-terc-butylcatechol presented quite interesting profile in relation to the influence of the substituent in the aromatic ring; 1,2-dihydroxybenzene shows reduction peak and the 3,5-di-terc-butylcatechol is not observed. The complex formed with the 3-hydroxyflavone shows influence of several factors related to the structure of the flavonoid.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / No município do Cantá-RR a produção de mel amargo é atribuída pelos apicultores da região ao período de floração das plantas do gênero Vochysia. Esta pesquisa tem como objetivo investigar a existência de marcadores químicos que possam confirmar a origem botânica deste mel, bem como verificar sua composição química e atividade antioxidante. Para tanto, foram elaborados extratos de mel amargo por XAD-2, C18, líquido-líquido e mel bruto liofilizado. Para comparação da constituição foram obtidos também os extratos etanólicos das flores sem anteras e das anteras das flores (ricos em pólen). Os extratos de mel por XAD-2 e etanólicos das flores foram fracionados, sendo obtidos extratos hexânico, clorofórmico, acetato de etila e metanólico. Análises por espectrometria de massas foram realizadas com os diversos extratos e verificou-se uma grande semelhança entre os espectros. Por meio de padrões autênticos, foram identificados a crisina e o ácido cafeico nos extratos do mel amargo. Nas análises dos extratos hexânicos, por meio de cromatografia a gás acoplada à espectrometria de massas (CG-EM), foram identificados diversos n-alcanos e n-alcenos com tempos de retenção e índices de retenção idênticos no mel e nas flores. Para determinar a posição da dupla ligação realizou-se uma derivatização com dimetildissulfeto. Os n-alcenos semelhantes nos extratos do mel e das flores apresentaram insaturação na posição 1. Análises espectrofotométricas foram realizadas para quantificar os compostos fenólicos e foram obtidos 94,64 mg ácido gálico/100g de mel e os flavonoides com 86,23 g quercetina/100g de mel. O mel apresentou uma coloração branca, com base na escala de Pfund e a atividade antioxidante, um IC50 de 16,94 mg/mL, determinado por meio do método do DPPH. Este trabalho permitiu confirmar a origem botânica do mel obtida por melissopalinologia e por dados químicos, além de caracterizar quimicamente o mel através da quantificação de algumas classes de produtos naturais e sua atividade antioxidante. / The bitter honey production in the city of Cantá-RR has been attribute by beekeepers in the region to the period of flowering plants of the genus Vochysia. This research aims to investigate the existence of chemical markers that can confirm the botanical origin of the honey, as well as verify their chemical composition and antioxidant activity. Thus, we prepared extracts of bitter honey by XAD-2, C18, liquid-liquid (dichloromethane) and lyophilized honey. For comparison of the constitution were also obtained the extracts ethanol of the flowers without anthers and of the anthers of the flowers, rich in pollen. The extracts of honey XAD-2 and ethanol flowers were fractionated, obtaining extracts with hexane, chloroform, ethyl acetate and methanol. Mass spectrometric analyzes were performed with the different extracts and there was a great similarity between the spectra. By using authentic standards, chrysin and caffeic acid were identified in the extracts of bitter honey. In hexanic extract analyzes, through gas chromatography coupled to mass spectrometry (GC-MS), several n-alkanes and n- alkenes were identified with times of retention and identical retention index in the honey and in the flowers. To determine the position of the double bond, a derivatization with dimethuldisulfide was performed. The similar n-alkenes in extracts of honey and flowers presented unsaturation at position 1. Spectrophotometric analyses were performed to quantify the phenolic compounds getting to result of 94,64 mg of gallic acid /100 g of honey and flavonoids with 86,23 g/100g honey. The honey presented a white coloration, based on the Pfund scale and antioxidant activity one IC50 of 16,94 mg / mL,determined by means of the DPPH method. This work confirmed the botanical origin of honey obtained by melissopalynology and chemical data, and also contributed to the chemical characterization of the honey through quantification of some classes of natural products and their antioxidant activity.

Orientadores: Maria Regina Wolf Maciel, Patrícia Fazzio Martins Martinez / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Química / Made available in DSpace on 2018-08-26T01:57:05Z (GMT). No. of bitstreams: 1 Medeiros_HeloisaHelenaBerredoReisde_D.pdf: 2926658 bytes, checksum: ecdad0019abf8c59be4ebbda4314ac06 (MD5) Previous issue date: 2014 / Resumo: O Brasil é o maior produtor mundial de laranjas, sendo também considerado um grande produtor e exportador mundial do óleo de laranja. Óleo de laranja doce, um importante subproduto gerado a partir da produção de suco de laranja, é constituído por aproximadamente 400 compostos, dentre os quais, destacam-se as classes de terpenos, oxigenados e flavonoides. Essas classes de compostos apresentam elevado potencial para as indústrias alimentícia, cosmética, farmacêutica e química. O fracionamento deste óleo tem recebido grande atenção da comunidade científica e industrial devido à instabilidade de determinados componentes. Neste trabalho, a separação e concentração dos componentes limoneno, valenceno, decanal, tangeritina e nobiletina foram realizadas utilizando-se um sistema híbrido de evaporação a pressões de 2 e 20 mbar. Três frações resultantes do sistema foram obtidas, são elas: destilado, lateral e resíduo. Através de um planejamento experimental do tipo composto central, observou-se que a temperatura do evaporador é a variável que tem maior influência nos resultados. As maiores concentrações de limoneno foram encontradas nas frações destilado e lateral, chegando-se à concentração de 99,5%. Os compostos valenceno e decanal não foram identificados na fração destilado e suas maiores concentrações estão presentes na fração resíduo. Comportamento semelhante foi observado para os flavonoides tangeritina e nobiletina, os quais foram concentrados na fração resíduo. De acordo com os resultados obtidos neste trabalho, é promissor separar e concentrar os compostos citados acima presentes no óleo de laranja doce utilizando um sistema híbrido de evaporação / Abstract: Brazil is the world¿s largest orange producer, and is also considered a major producer and exporter of orange oil. Sweet orange oil, an important by-product generated from the production of orange juice, consists of approximately 400 compounds, among which stand out the classes of terpenes, oxygenated and flavonoids. These classes of compounds have high potential for the food, cosmetic, pharmaceutical and chemical industries. Fractionation of this oil has received great attention of the scientific and industrial community due to the intability of certain components. In this work, the separation and concentration of the components limonene, valencene, decanal, tangeritin and nobiletin were performed using a hybrid system of evaporation at pressures from 2 and 20 mbar. Three fractions of the resulting system were obtained, they are: distillate, side and residue. Through an experimental design of the central composite type, it is observed that the evaporator temperature is the variable that has the greatest influence on the results. The highest concentrations of limonene were found in the distillate and side fractions, reaching to the concentration of 99.5%. The valencene and decanal compounds were not identified in the distillate fraction and its highest concentrations are present in the residue fraction. Similar behavior eas observed for tangeritin and nobiletin flavonoids, which are concentrated in the residue fraction. According to the results obtained in this study it is promising to separate and concentrate the compounds mentioned above present in sweet orange oil using a hybrid system of evaporation / Doutorado / Desenvolvimento de Processos Químicos / Doutora em Engenharia Quimica

Orientador: Gabriela Alves Macedo / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos / Made available in DSpace on 2018-08-20T21:21:56Z (GMT). No. of bitstreams: 1 Ferreira_LiviaRosas_M.pdf: 6389418 bytes, checksum: 474ea71f0f4b85a4fb59fa346c31f972 (MD5) Previous issue date: 2012 / Resumo: A tanino acil hidrolase (EC 3.1.1.20), conhecida como tanase, é uma enzima com habilidade de atuar em ligações éster e depsídicas de taninos hidrolisáveis e também foi descrita como capaz de hidrolisar polifenóis como ácido clorogênico, epicatequina galato e epigalocatequina galato. Os polifenóis, ou compostos fenólicos, estão distribuídos em uma ampla variedade de fontes vegetais e estão relacionados à prevenção de câncer e doenças cardiovasculares. No entanto, os polifenóis ocorrem frequentemente como glicosídeos ou outros conjugados, o que pode comprometer a biodisponibilidade e os efeitos benéficos destes compostos à saúde, sendo necessária a biotransformação destes conjugados no trato gastrointestinal. Como alternativa tecnológica, a biotransformação enzimática dos polifenóis ou de suas fontes vegetais também pode liberar estes compostos de seus conjugados e consequentemente, melhorar a atividade funcional de tais antioxidantes. O objetivo do presente trabalho foi estudar a especificidade do extrato semipurificado de tanase de Paecilomyces variotii frente a diferentes padrões comerciais de polifenóis e avaliar sua atuação na matriz alimentar suco de laranja quanto a modificações em características físico-químicas, perfil fenólico e atividade antioxidante. O estudo da especificidade do extrato semipurificado de tanase foi realizado por CLAE-DAD e ESI-MS e o suco de laranja foi avaliado quanto aos parâmetros acidez total titulável, sólidos totais, pH, teor de vitamina C e fenólicos totais. As alterações no perfil fenólico do suco de laranja foram avaliadas por CLAE-DAD e a atividade antioxidante pelos métodos in vitro ORAC e de sequestro de radicais DPPH. Os resultados obtidos por CLAE-DAD e ESI-MS mostraram que não houve hidrólise dos padrões ácido clorogênico, ácido ferúlico, ácido elágico, resveratrol, quercetina e hesperetina pelo extrato semipurificado de tanase nas condições empregadas para o teste. No entanto, foi observado a hidrólise da ligação entre a aglicona e o dissacarídeo dos flavonoides rutina, naringina e hesperidina, gerando respectivamente os produtos quercetina, naringenina e hesperetina, e indicando uma atividade diglicosídica do extrato semipurificado de tanase. Para o suco de laranja, os resultados de acidez titulável, pH, sólidos solúveis e vitamina C não apresentaram diferença significativa após o tratamento enzimático do suco com extrato semipurificado de tanase, no entanto, houve um aumento de 16,8% no teor de fenólicos totais. Além disso, foi verificada uma modificação no perfil polifenólico do suco de laranja obtido por CLAE-DAD, resultando em um aumento na atividade antioxidante do suco biotransformado em aproximadamente 50% pelo método ORAC e 70% pelo método DPPH. Os padrões comerciais hesperidina e naringina biotransformados pelo extrato semipurificado de tanase também apresentaram maior atividade antioxidante do que o controle sem reação. Pelo que se tem conhecimento, a hidrólise de flavonoides glicosilados por tanase é um relato inédito na literatura e, portanto este trabalho fornece novos substratos para o extrato semipurificado de tanase de P. variotii e confirma que a biotransformação é uma boa estratégia para melhorar a atividade antioxidante in vitro de polifenóis e de suco de laranja / Abstract: Tannin acyl hydrolases (EC 3.1.1.20), commonly named as tannases, are enzymes able to act on ester and depside bonds of hydrolysable tannins and have also been described to hydrolyse polyphenols such as chlorogenic acid, epicatechin gallate and epigallocatechin gallate. Polyphenols, or phenolic compounds, are widely distributed in the plant kingdom and are related to the prevention of cancer and cardiovascular diseases. However, polyphenols are often found as glycosides or other conjugates, which may affect their bioavailability and health benefits, being necessary the biotransformation of these compounds in the gastrointestinal tract. As a technological alternative, the enzymatic biotransformation of the polyphenols or their plant sources can also release these compounds from their conjugates and therefore improve the functional activity of these antioxidants. The objectives of the present study were to investigate the specificity of the crude extract of tannase from Paecilomyces variotii in reaction with differents comercial standards of phenolic compounds and to evaluate the effects on the physico-chemical properties, phenolic profile and antioxidant activity of the orange juice reacted with the crude extract of tannase. The specificity study of the crude extract of tannase was performed by HPLC-DAD and ESI-MS. The orange juice was subject to analysis of titratable acidity, pH, and total solids, vitamin C and total phenolics contents. Furthermore, changes in the phenolic profile of orange juice were analyzed by HPLC-DAD and the antioxidant capacity of the samples was tested by in vitro DPPH and ORAC assays. The results obtained by HPLC-DAD and ESI-MS did not show hydrolysis of the standards chlorogenic acid, ferulic acid, ellagic acid, resveratrol, quercetin and hesperetin at test conditions. However, the crude extract of tannase was able to hydrolyse the glucosidic bond between disaccharide and aglycone of the flavonoids rutin, hesperidin and naringin, yielding as products quercetin, hesperetin and naringenin, respectively, and indicating a diglycosidase activity of the crude extract of tannase. For orange juice, the results of acidity, pH, soluble solids and vitamin C showed no significant difference after enzymatic treatment with the crude extract of tannase, but there was an increase of 16.8% in total phenolic content. In addition, there was an extensive modification in the polyphenolic profile from the biotransformed orange juice obtained by HPLC-DAD, which resulted about 50% and 70% increase in orange juice antioxidant activity by ORAC and DPPH methods, respectively. The standards hesperidin and naringin biotransformed by crude extract of tannase also showed higher antioxidant activities than the control. To the best of our known, this is the first report about tannase which could hydrolyze flavonoid glycosides and, therefore this work provides new substrates for crude extract of tannase from P. variotii and confirms that biotransformation is a good strategy to improve in vitro antioxidant activity of polyphenols and orange juice / Mestrado / Ciência de Alimentos / Mestra em Ciência de Alimentos

Orientador: Paulo Cesar Rodrigues Palma / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-04T10:03:35Z (GMT). No. of bitstreams: 1 Dambros_Miriam_D.pdf: 5436149 bytes, checksum: adcf62040b0e93aa24e7479ee60c4ebb (MD5) Previous issue date: 2005 / Resumo: Os agentes antimuscarínicos permanecem como drogas de primeira linha para o tratamento da Síndrome da Bexiga Hiperativa (SBH), a despeito das dúvidas ainda existentes sobre a significância clínica e sua eficácia. Poucos fármacos que atuam por meio de mecanismos distintos dos antimuscarínicos, têm demonstrado eficácia no tratamento da SBH. Ravonóides são um grupo de compostos polifenólicos que têm recebido especial atenção dentro da pesquisa básica e clínica devido ao seu amplo espectro de atividade farmacológica. Recentemente, um estudo in vitro demonstrou que o fIavonóide galangina exerce uma atividade inibitória sobre a contratilidade da bexiga de ratos. Esta tese foi desenvolvida a fim de examinar os efeitos da galangina sobre a resposta contrátil da bexiga de suínos e testar a hipótese de um efeito sobre a mobilização do cálcio. Além disso, investigou-se a eficácia da galangina em promover uma ação protetora sobre o dano da musculatura lisa da bexiga causada por um período de estimulação elétrica repetitiva, o qual é empregado como um modelo, in vitro, de hiperrefIexia. Em um grupo de experimentos, fragmentos do detrusor de suínos foram submetidos a um período de 90 mÍn de estimulação elétrica de repetição. Galangina, em diferentes concentrações, foi adicionada ao banho do órgão a fim de determinar seu possível efeito protetor durante a estimulação elétrica. Em outro grupo de experimentos, a resposta contrátil ao carbacol, cafeína, potássio e eletroestímulo foi determinada antes e após a adição de galangina (3xl0-sM) ao meio. Os efeitos do fIavonóide foram também avaliados após a administração de diferentes antagonistas no banho de órgão. Galangina (1O-7M) evitou o decréscimo da resposta contrátil do músculo liso causada por um período de estimulação elétrica de repetição. vesical Galangina inibiu a amplitude da contração detrusora induzi da pelo eletro-estímulo, carbacol, cafeína e potássio (p<0.05). O efeito inibitório do fIavonóide foi deixado inalterado após a introdução de diferentes antagonistas: propranolol, fentolamina e capsazepina, no banho de órgão (p>0,05). Entretanto, quando verapamil foi adicionado ao meio de estudo, a inibição da contração foi parcialmente abolida. Concluindo, os experimentos acima descritos demonstraram que a galangina, em concentração nanomolar, exerceu um efeito protetor sobre a musculatura detrusora. Entretanto, quando empregado em uma concentração micromolar, o flavonóide promoveu einibição da musculatura lisa vesical, o qual resultou de uma ação sobre a mobilização do cálcio extra e intracelular / Abstract: Antimuscarinjc agents remain first-line therapy for the overactive bladder syndrome (OAB), despite doubts about the clinjcal signifjcance of their effectiveness. Few drugs acting through other mechanisms have been found to be efficacious for treatment of GAB. Flavonoids are a group of polyphenolic compounds and have recently gained tremendous interest, due to their broad pharmacologjcal activity. Recently, the inhibitory effect of galangin, a member of the flavonol class of flavonoid, on rat bladder contractility has been investigated. This thesis was undertaken to further examine the actions of galangin on the contractile responses of bladder strips and to test the hypothesis that an effect on calcium mobilization in smooth muscle cells is involved. Furthermore, it was investigated the efficacy of galangin to counteract the detrusor damage caused by the smooth muscle of the urinary bladder being exposed to repetitive field stimulation (RFS), as a model of hyperreflexia. In one experiment, pig detrusor strips were mounted for tension recording in organ baths and were subjected to RFS for 90min. Galangin, at different concentrations, was added to the organ bath in order to determine its protective effect on RFS. In another experiment the contractile response to carbachol, potassium and electrical field stimulation - EFS were determined before and after the addition of galangin (3xlO-sM). Furthermore, the effect of galangin was also evaluated after the administration in the bath of a number of antagonists. Galangin (1O-7M)avoided the decrease in contractile smooth muscle response of strips to EFS during RFS. Galangin inhibited the maximal contractile response to EFS, carbachol, caffeine media and potassium (pO,O5).However, when verapamil was added to the medium, the inhibitory effects of galangin were partially blocked. In summary, the current experiments have shown that galangin, in subllmol concentrations, exerted a protective effect on bladder smooth muscle contractility. Furthermore, galangin, in !lIDO!concentrations inhibited pig bladder contractility, which probably resulted from an action on Ca2+mobilization from intracellu!ar stores as well as influx of calcium. At the present, there is no proof that our results are reproducible in functional or dysfunctional human bladders. Further study is needed to investigate the potential of ga!anginto be used to inhibited overactive bladder syndrome in vivo / Doutorado / Cirurgia / Doutor em Cirurgia

The general aim of the thesis was to deduce whether selected naturally occurring flavonoids (genistein, epicatechin gallate (EC), epigallocatechin gallate (EGCG), baicalin) attenuate various secondary insults that may cause death of ganglion cells in primary open angle glaucoma (POAG). An ischemic insult to the rat retina significantly causes the inner retina to degenerate indexed by changes of various antigens, proteins and mRNAs located to amacrine and ganglion cells. These changes are blunted in animals treated with genistein as has been shown for ECGC. Studies conducted on cells (RGC-5 cells) in culture showed that hydrogen peroxide, L-buthionine sulfoximine (BSO)/glutamate and serum deprivation (mimicking oxidative stress), rotenone, sodium azide (affecting mitochondria function in specific ways) and light (where the mitochondria are generally affected) all generated reactive oxygen species and caused death of RGC-5 cells. EGCG was able to attenuate cell death caused by hydrogen peroxide, sodium azide and rotenone. Only EC was able to attenuate BSO/glutamate-induced cell death, in addition to cell death caused by hydrogen peroxide and rotenone. Genistein had no positive effect on cell death in experiments carried out on RGC-5 cells. Exposure of RGC-5 cells to flavonoids showed that EC and EGCG increased the mRNA expression of endogenous antioxidants such as HO-1 (heme oxygenase 1) and Nrf-2 (nuclear erythroid factor-2-related factor 2). Light insult, rotenone and sodium azide activate the p38 (protein kinase 38) pathway, while only light and rotenone activate the JNK (c-Jun amino-terminal kinase) pathway. Serum deprivation affects mitochondrial apoptotic proteins causing an increase in the ratio of Bax/Bcl2 (Bax: Bcl-2-associated X protein; Bcl-2: B-cell lymphoma 2). An insult of light to RGC-5 cells, unlike that induced by sodium azide, is inhibited by necrostatin-1 and causes an activation of AIF (apoptosis-inducing factor) with alpha-fodrin being unaffected. These studies suggest that ganglion cell death caused by insults as may occur in POAG involves various cellular signaling pathways. The selected flavonoids have diverse actions in increasing cellular defense mechanisms, and in negating the effects of ischemia and specific types of oxidative stress. The results argue for the possible use of flavonoids in the treatment of POAG to slow down ganglion cell death.

Abstract Increased amounts of epicuticular waxes and UV-absorbing compounds, such as flavonoids, and smaller leaf/needle surface area are plant defence mechanisms against UV-B radiation. The response of the needle epicuticular waxes of Scots pine (Pinus sylvestris L.) and Norway spruce (Picea abies Karst.) seedlings to increased UV-B were investigated in short-term and long-term greenhouse experiments. In a more realistic long-term field experiment with mature Scots pines, the methanol-extractable UV-absorbing compounds were also analysed. Some significant changes were observed in the wax tube distribution (WTD, %) and the amount of waxes in Norway spruce seedlings in the short-term Belgian greenhouse experiment (UV-BBE 0, 11.3 and 22.6 kJ m-2 d-1), but no changes were detected in Scots pine seedlings. No changes in waxes were observed in the long-term Finnish greenhouse experiment (UV-BBE 0, 2.2–6.6 and 5.6–16.8 kJ m-2 d-1), where both the Norway spruce and the Scots pine seedlings seemed to respond by having smaller needle surface areas. A field experiment (UV-BBE 0.5–2.4 kJ m-2 d-1 and 0.7–5.1 kJ m-2 d-1) with mature Scots pines revealed no significant changes in WTD during the three growing seasons or the amount of waxes during the third growing season. In the long-term field experiment the amount of UV-absorbing compounds varied significantly between seasons and/or needle age classes. Elevated amounts of these compounds were already observed in the three-day-old needles and also in the oldest (c + 2) needles when the waxes were still undeveloped or already somewhateroded. No significant differences in the amount of UV-absorbing compounds were observed between the treatments during the first and second growing seasons. During the third growing season, needles of all ages contained significantly or slightly less UV-absorbing compounds in supplemental UV-B than in the ambient treatment, possibly due to cumulative effects of UV-B in already inhibited pigment synthesis. This suggests that these defence mechanisms are not efficient enough to prevent the UV-B-induced damage in the long term.

Ph.D. / Please refer to full text to view abstract

Magister Scientiae - MSc / The overall aims of this project were to assess the pharmaceutical quality and consistency of commercially available solid dose Sutherlandia frutescens containing products (viz. tablets & capsules) by exploring the use of monitoring the pharmaceutical presentation, flavonoid profile and antioxidant activity levels and to develop/or adapt methods and specifications that may be used for the quality control of such products.Stability tests were conducted on all of the selected SCP. The products were stored under elevated temperatures and environmental humidity conditions and total phenol, antioxidant and chromatographic analysis was conducted on these samples. Samples of each of the SCP were hydrolyzed using HCL and then analyzed using HPLC to test the stability of the flavonoids present in each product. The SCP investigated in this study physically appeared to be of quite good “pharmaceutical” quality, but generally lacked information on the date of manufacture and lacked package inserts, or when these were present they contained insufficient information. Based on the results obtained, it is recommended that, the manufacturers of SCP pay more attention to the information provided on the package inserts and the storage conditions for their products. Further the levels of antioxidant activity, total phenols and flavonoid (sutherlandins A to D) be used as specifications to control the quality of commercially available solid dose Sutherlandia frutescens containing preparations on an individual basis. / South Africa

Philosophiae Doctor - PhD / Various formulations of the popular South African medicinal plant, Sutherlandia frutescens,are commercially available, with no documented specifications for quality assessment. With plans already underway for a clinical trial to assess its efficacy in HIV patients, there is a need for scientifically validated tests for the quality control of products of this plant. Chemical constituents of the plant are many and varied but it is still unclear which might be the most appropriate ones to monitor for activity or to describe the quality of the plant’s products. For quality control and regulatory purposes, the content and dissolution of flavonoids in the plant products can be assessed. However, these compounds are not monitored for regulation and there are as yet no HPLC or dissolution methods that can be employed for quality control of herbals like S. frutescens. Therefore, the objectives of this study were to assess the suitability of its flavonoid constituents as quality control (QC) marker compounds, and the suitability of content levels and dissolution tests of flavonoids as QC tools for S. frutescens products. To realise the afore-mentioned objectives, non-commercially available flavonoid compounds (sutherlandins) that could be used as marker compounds were isolated from S. frutescens. An HPLC assay was developed and validated for determination of flavonoid content in solution. Five S. frutescens materials viz leaf powder (LP), spray-dried aqueous extract (SDAE) and freeze-dried aqueous extracts (FDAE) were analysed for flavonoid content and dissolution. Dissolution tests were conducted for different S. frutescens materials and dissolution profiles of flavonoids in capsules containing these materials were compared using Q-release values, the similarity factor (f2) and mathematical models. To predict in vivo bioavailability of the flavonoids, in silico assessment of in vivo bioavailability of flavonoids (glycosides and aglycones) that may be contained in different S. frutescens materials was conducted. Sutherlandins A, B, C and D were successfully isolated (percentage purity approximately99 % for sutherlandins A, C and D, and 90 % for sutherlandin B) and identified, and used, along with other flavonoid compounds, for the development of a simple and robust HPLC method. Content of sutherlandins A, B, C and D, quercetin and kaempferol in different plant materials were 0.4 ± 0.3, 0.8 ± 0.2, 1.3 ± 0.2, 0.6 ± 0.1, 0.01 ± 0.02 and 0.08 ±0.1 %,respectively, and differed significantly (p < 0.001). In vitro dissolution showed faster dissolution of flavoniod glycosides compared to aglycones. The flavonoids from the LP and SDAE materials showed characteristics of immediate release with Q75 in ≤ 45 minutes, and delayed release from the FDAE material, i.e. Q75 > 45 minutes. The dissolution profiles of each flavonoid compared from different S. frutescens materials were different as signified by their f2 values which were all below 50. The mathematical models describing release were also different for each flavonoid from the different S. frutescens materials. For in vivo bioavailability modelling and prediction studies, the flavonoid aglycones met the conditions for oral bioavailability while the flavonoid glycosides did not. In conclusion, the sutherlandins isolated from S. frutescens proved to be good markers for HPLC assay and dissolution tests of S. frutescens materials. The HPLC method was suitable for assessing flavonoid levels in S. frutescens materials, and also showed differences in flavonoid content in these materials. The dissolution method was simple and reproducible, and Q-release values, the f2 and mathematical models proved to be good tools for differentiating between S. frutescens materials. In silico modelling showed that the flavonoid glycosides and aglycones differed in oral bioavailability. Although not presently required by the Medicines Control Council (MCC), quantification, release and dissolution studies and specifications may be employed as tools for routine analysis and for quality control of herbal drug formulations containing S. frutescens.