A atividade antioxidante (AOX) de vinhos é decorrente, dentre outros fatores, da presença de substâncias fenólicas, como as antocianinas que, além de contribuírem para a formação da cor, devido a sua capacidade redutora, são capazes de neutralizar moléculas de radicais-livres. No presente trabalho, 92 amostras de vinhos tintos da safra 2008/2009, provenientes de diversas regiões do país, agrupadas de acordo com a sua similaridade (UF e fisiografia), foram analisados quanto à cor, através de parâmetros de luz, saturação e tonalidade; à concentração de antocianinas totais (por espectrofotometria diferencial e por CLAE); à caracterização dos pigmentos (por CLAE); à concentração de polifenóis totais (IPT) (pelo método de Folin Ciocalteau’s); e à atividade antioxidante (por DPPH+), sendo que as duas últimas características foram avaliadas sob a forma de evolução temporal. Os resultados mostram que, em relação ao perfil evolutivo, pelo menos um dos grupos difere (p<0,05) nos tempos. A região Vale do Rio São Francisco (VSF) exibiu, tanto os mais elevados valores de IPT em todos os tempos de análise [(tzero=4.516; t25=3.516; t35=2.475; t45=2.027; t55=1.598) mg.L-1 GAE], quanto a mais alta AOX, nos quatro primeiros dos cinco momentos avaliados. A região Oeste do Paraná (OPR) apresentou os mais baixos resultados para ambos os parâmetros considerados em todos os tempos medidos, sendo que os valores de IPT corresponderam a 47, 52, 54, 58 e 63% e os valores de AOX a 49, 55, 57, 61 e 79% dos valores encontrados para VSF. No decorrer das 55 semanas avaliadas, as diferenças entre os grupos tende a diminuir. Em relação aos pigmentos, 12 padrões comerciais permitiram comparar os vinhos no que diz respeito as presença e a quantificação destes compostos. À exceção de dois, todos os vinhos analisados por CLAE confirmaram a malvidina-3-monoglicosídeo (Mv-3G), como o composto antociânico predominante. O Vale dos Vinhedos (Vvi) apresentou os maiores conteúdos, tanto do pigmento majoritário da uva (Mv-3G – 38,66 μ.L-1), quanto da cianidina-3-monoglicosídeo (Cn- 3G – 0,914 μ.L-1), composto antociânico mais relatado em alimentos. Correlações fortes e muito fortes foram observadas entre IPT e AOX e fortes entre antocianinas totais (expressas em Mv-3G por espectrofotometria diferencial) e Mv-3G (quantificada por CLAE). ANOVA mostrou que, em CLAE, não houve diferenças (p<0,1) entre as regiões, indicando que variações internas nos grupos, sobressaíram-se às evidenciadas pelas regiões, não sendo possível caracterizá-las conforme o tipo nem a concentração de antocianinas. / The antioxidant capacity (AOX) of wines is due, among other factors the phenolic substances, such as anthocyanins that, beyond to color contribution, present scavenging properties, having effect on radical oxygen species (ROS). In this study, 92 samples of 2008/2009 vintage Brazilian red wines, from dissimilar production areas, were grouped according to similarities (Federal State and physiographic attributes) and analyzed in terms of color, through light, saturation and hue parameters; in terms of total anthocyanin concentration (by pH-differential spectrophotometric methods and high performance liquid chromatography (HPLC)); in terms of pigment characterization (HPLC); in terms of total polyphenols concentration (IPT) (Folin Ciocalteau’s method); and in terms of its scavenging properties (DPPH+), being the last two attributes analyzed over time evolution profile. The results showed that for the evolution profile, at least one of the groups differed (p<0,05) in the evaluated times. The “Vale do Rio São Francisco” (VSF) region exhibited not only the most elevated values for IPT in all analyzed sampling [(tzero=4,516; t25=3,516; t35=2,475; t45=2,027; t55=1,598) mg.L-1 GAE], as the most AOX in the first four from the five evaluated moments. The “Oeste do Paraná” (OPR) presented the lowest results for both IPT and AOX parameters, in all measuring times, and the IPT values corresponded to 47, 52, 54, 58 e 63%, while the AOX values to 49, 55, 57, 61 e 79% from values found for VSF. Over the 55 weeks, the differences among groups tended to diminish. Considering the pigments, 12 commercial standards allowed to compare wines based on presence and concentration of these compounds. Except for two samples, all the HPLC-analyzed wines confirmed the malvidin-3-monoglucoside, as the anthocyanin most abounding in wines. The VVi exhibited the highest levels, even for the grape major pigment (Mv- 3G – 38.66 μ.L-1), as for cianidin-3-monoglucoside (Cn-3G – 0.914 μ.L-1), the most reported anthocyanin in food. Strong and very strong correlations were found among IPT and AOX, while total anthocyanins (in Mv-3G base, by pH-differential analysis) and Mv-3G concentration (by HPLC) exhibited strong linear associations. ANOVA showed that, in terms of chromatographic analysis, no differences (p<0,1) among regions were found, indicating that inside-groups variations were high enough to not allow the characterization according to qualitative and quantitative anthocyanin profile.

A atividade antioxidante (AOX) de vinhos é decorrente, dentre outros fatores, da presença de substâncias fenólicas, como as antocianinas que, além de contribuírem para a formação da cor, devido a sua capacidade redutora, são capazes de neutralizar moléculas de radicais-livres. No presente trabalho, 92 amostras de vinhos tintos da safra 2008/2009, provenientes de diversas regiões do país, agrupadas de acordo com a sua similaridade (UF e fisiografia), foram analisados quanto à cor, através de parâmetros de luz, saturação e tonalidade; à concentração de antocianinas totais (por espectrofotometria diferencial e por CLAE); à caracterização dos pigmentos (por CLAE); à concentração de polifenóis totais (IPT) (pelo método de Folin Ciocalteau’s); e à atividade antioxidante (por DPPH+), sendo que as duas últimas características foram avaliadas sob a forma de evolução temporal. Os resultados mostram que, em relação ao perfil evolutivo, pelo menos um dos grupos difere (p<0,05) nos tempos. A região Vale do Rio São Francisco (VSF) exibiu, tanto os mais elevados valores de IPT em todos os tempos de análise [(tzero=4.516; t25=3.516; t35=2.475; t45=2.027; t55=1.598) mg.L-1 GAE], quanto a mais alta AOX, nos quatro primeiros dos cinco momentos avaliados. A região Oeste do Paraná (OPR) apresentou os mais baixos resultados para ambos os parâmetros considerados em todos os tempos medidos, sendo que os valores de IPT corresponderam a 47, 52, 54, 58 e 63% e os valores de AOX a 49, 55, 57, 61 e 79% dos valores encontrados para VSF. No decorrer das 55 semanas avaliadas, as diferenças entre os grupos tende a diminuir. Em relação aos pigmentos, 12 padrões comerciais permitiram comparar os vinhos no que diz respeito as presença e a quantificação destes compostos. À exceção de dois, todos os vinhos analisados por CLAE confirmaram a malvidina-3-monoglicosídeo (Mv-3G), como o composto antociânico predominante. O Vale dos Vinhedos (Vvi) apresentou os maiores conteúdos, tanto do pigmento majoritário da uva (Mv-3G – 38,66 μ.L-1), quanto da cianidina-3-monoglicosídeo (Cn- 3G – 0,914 μ.L-1), composto antociânico mais relatado em alimentos. Correlações fortes e muito fortes foram observadas entre IPT e AOX e fortes entre antocianinas totais (expressas em Mv-3G por espectrofotometria diferencial) e Mv-3G (quantificada por CLAE). ANOVA mostrou que, em CLAE, não houve diferenças (p<0,1) entre as regiões, indicando que variações internas nos grupos, sobressaíram-se às evidenciadas pelas regiões, não sendo possível caracterizá-las conforme o tipo nem a concentração de antocianinas. / The antioxidant capacity (AOX) of wines is due, among other factors the phenolic substances, such as anthocyanins that, beyond to color contribution, present scavenging properties, having effect on radical oxygen species (ROS). In this study, 92 samples of 2008/2009 vintage Brazilian red wines, from dissimilar production areas, were grouped according to similarities (Federal State and physiographic attributes) and analyzed in terms of color, through light, saturation and hue parameters; in terms of total anthocyanin concentration (by pH-differential spectrophotometric methods and high performance liquid chromatography (HPLC)); in terms of pigment characterization (HPLC); in terms of total polyphenols concentration (IPT) (Folin Ciocalteau’s method); and in terms of its scavenging properties (DPPH+), being the last two attributes analyzed over time evolution profile. The results showed that for the evolution profile, at least one of the groups differed (p<0,05) in the evaluated times. The “Vale do Rio São Francisco” (VSF) region exhibited not only the most elevated values for IPT in all analyzed sampling [(tzero=4,516; t25=3,516; t35=2,475; t45=2,027; t55=1,598) mg.L-1 GAE], as the most AOX in the first four from the five evaluated moments. The “Oeste do Paraná” (OPR) presented the lowest results for both IPT and AOX parameters, in all measuring times, and the IPT values corresponded to 47, 52, 54, 58 e 63%, while the AOX values to 49, 55, 57, 61 e 79% from values found for VSF. Over the 55 weeks, the differences among groups tended to diminish. Considering the pigments, 12 commercial standards allowed to compare wines based on presence and concentration of these compounds. Except for two samples, all the HPLC-analyzed wines confirmed the malvidin-3-monoglucoside, as the anthocyanin most abounding in wines. The VVi exhibited the highest levels, even for the grape major pigment (Mv- 3G – 38.66 μ.L-1), as for cianidin-3-monoglucoside (Cn-3G – 0.914 μ.L-1), the most reported anthocyanin in food. Strong and very strong correlations were found among IPT and AOX, while total anthocyanins (in Mv-3G base, by pH-differential analysis) and Mv-3G concentration (by HPLC) exhibited strong linear associations. ANOVA showed that, in terms of chromatographic analysis, no differences (p<0,1) among regions were found, indicating that inside-groups variations were high enough to not allow the characterization according to qualitative and quantitative anthocyanin profile.

A atividade antioxidante (AOX) de vinhos é decorrente, dentre outros fatores, da presença de substâncias fenólicas, como as antocianinas que, além de contribuírem para a formação da cor, devido a sua capacidade redutora, são capazes de neutralizar moléculas de radicais-livres. No presente trabalho, 92 amostras de vinhos tintos da safra 2008/2009, provenientes de diversas regiões do país, agrupadas de acordo com a sua similaridade (UF e fisiografia), foram analisados quanto à cor, através de parâmetros de luz, saturação e tonalidade; à concentração de antocianinas totais (por espectrofotometria diferencial e por CLAE); à caracterização dos pigmentos (por CLAE); à concentração de polifenóis totais (IPT) (pelo método de Folin Ciocalteau’s); e à atividade antioxidante (por DPPH+), sendo que as duas últimas características foram avaliadas sob a forma de evolução temporal. Os resultados mostram que, em relação ao perfil evolutivo, pelo menos um dos grupos difere (p<0,05) nos tempos. A região Vale do Rio São Francisco (VSF) exibiu, tanto os mais elevados valores de IPT em todos os tempos de análise [(tzero=4.516; t25=3.516; t35=2.475; t45=2.027; t55=1.598) mg.L-1 GAE], quanto a mais alta AOX, nos quatro primeiros dos cinco momentos avaliados. A região Oeste do Paraná (OPR) apresentou os mais baixos resultados para ambos os parâmetros considerados em todos os tempos medidos, sendo que os valores de IPT corresponderam a 47, 52, 54, 58 e 63% e os valores de AOX a 49, 55, 57, 61 e 79% dos valores encontrados para VSF. No decorrer das 55 semanas avaliadas, as diferenças entre os grupos tende a diminuir. Em relação aos pigmentos, 12 padrões comerciais permitiram comparar os vinhos no que diz respeito as presença e a quantificação destes compostos. À exceção de dois, todos os vinhos analisados por CLAE confirmaram a malvidina-3-monoglicosídeo (Mv-3G), como o composto antociânico predominante. O Vale dos Vinhedos (Vvi) apresentou os maiores conteúdos, tanto do pigmento majoritário da uva (Mv-3G – 38,66 μ.L-1), quanto da cianidina-3-monoglicosídeo (Cn- 3G – 0,914 μ.L-1), composto antociânico mais relatado em alimentos. Correlações fortes e muito fortes foram observadas entre IPT e AOX e fortes entre antocianinas totais (expressas em Mv-3G por espectrofotometria diferencial) e Mv-3G (quantificada por CLAE). ANOVA mostrou que, em CLAE, não houve diferenças (p<0,1) entre as regiões, indicando que variações internas nos grupos, sobressaíram-se às evidenciadas pelas regiões, não sendo possível caracterizá-las conforme o tipo nem a concentração de antocianinas. / The antioxidant capacity (AOX) of wines is due, among other factors the phenolic substances, such as anthocyanins that, beyond to color contribution, present scavenging properties, having effect on radical oxygen species (ROS). In this study, 92 samples of 2008/2009 vintage Brazilian red wines, from dissimilar production areas, were grouped according to similarities (Federal State and physiographic attributes) and analyzed in terms of color, through light, saturation and hue parameters; in terms of total anthocyanin concentration (by pH-differential spectrophotometric methods and high performance liquid chromatography (HPLC)); in terms of pigment characterization (HPLC); in terms of total polyphenols concentration (IPT) (Folin Ciocalteau’s method); and in terms of its scavenging properties (DPPH+), being the last two attributes analyzed over time evolution profile. The results showed that for the evolution profile, at least one of the groups differed (p<0,05) in the evaluated times. The “Vale do Rio São Francisco” (VSF) region exhibited not only the most elevated values for IPT in all analyzed sampling [(tzero=4,516; t25=3,516; t35=2,475; t45=2,027; t55=1,598) mg.L-1 GAE], as the most AOX in the first four from the five evaluated moments. The “Oeste do Paraná” (OPR) presented the lowest results for both IPT and AOX parameters, in all measuring times, and the IPT values corresponded to 47, 52, 54, 58 e 63%, while the AOX values to 49, 55, 57, 61 e 79% from values found for VSF. Over the 55 weeks, the differences among groups tended to diminish. Considering the pigments, 12 commercial standards allowed to compare wines based on presence and concentration of these compounds. Except for two samples, all the HPLC-analyzed wines confirmed the malvidin-3-monoglucoside, as the anthocyanin most abounding in wines. The VVi exhibited the highest levels, even for the grape major pigment (Mv- 3G – 38.66 μ.L-1), as for cianidin-3-monoglucoside (Cn-3G – 0.914 μ.L-1), the most reported anthocyanin in food. Strong and very strong correlations were found among IPT and AOX, while total anthocyanins (in Mv-3G base, by pH-differential analysis) and Mv-3G concentration (by HPLC) exhibited strong linear associations. ANOVA showed that, in terms of chromatographic analysis, no differences (p<0,1) among regions were found, indicating that inside-groups variations were high enough to not allow the characterization according to qualitative and quantitative anthocyanin profile.

Type 2 diabetes mellitus (T2DM) is an increasing global health concern, currently affecting an estimated 382 million individuals. There is no cure for T2DM and the search for new and improved treatments is ongoing. Presently, various pharmacological regimens are available to treat T2DM, but with varied success. Thousands of traditional herbs are also used to treat T2DM, but mainly without scientific validation. The aim of this study was to assess the polyphenolic content, antioxidant capacity, as well as in vitro toxicity and hypoglycaemic activity of a commercial ‘antidiabetic’ tea mixture (Diabetea) and its individual constituents: Achillea millefolium L. (Yarrow), Agathosma betulina Bartl. & Wendl. (Buchu), Salvia officinalis L. (Sage), Taraxacum officinalis L. (Dandelion), Thymus vulgaris L. (Thyme), Trigonella foenum-graecum L. (Fenugreek) and Urtica urens L. (Nettle). All herbs were tested as crude extracts, prepared using hot water (HW) and dichloromethane (DCM). The total polyphenolic content of each extract was determined using the Folin-Ciocalteau and aluminium trichloride methods. The non-cellular antioxidant activity was assessed using 2,2-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) methods. The cell-based antioxidant activity was measured against p-chloranil-induced generation of reactive oxygen species (ROS) in Ea.hy926 cells, using the fluorescent dye, 2',7'-dichlorfluorescein-diacetate (DCFH-DA). The effect of each extract on the viability of C2C12 myotubes, Ea.hy926 endothelial cells and human lymphocytes (HL) was determined using sulforhodamine B (SRB). The in vitro hypoglycaemic activity was assessed against α-amylase and α-glucosidase activity using 3,5-dinitrosalicylic acid (DNSA) and p-nitrophenyl-α-D-glucopyranoside (p-NPG), respectively. The type of inhibition exerted on these enzymes was determined using the Michaelis-Menten enzyme kinetics model, expressed as mixed, competitive, non-competitive and uncompetitive. Glucose uptake activity was measured using the 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-D-glucose (2-NBDG) fluorescent analogue. T. vulgaris and S. officinalis had the highest amount of polyphenols of all extracts tested. The HW extracts of T. vulgaris and S. officinalis showed significant (p < 0.05) cell-free antioxidant activity and cell-based radical scavenging activity. In addition, U. urens (HW) also limited cell-based ROS generation (p < 0.05). The Diabetea extracts presented with poor antioxidant activities, of which some had a pro-oxidant effect on Ea.hy926 cells. The positive linear relationship between antioxidant activity and polyphenolic content was shown to be dependent on the solvent type used. All of the DCM extracts had low antioxidant activity and polyphenolic content. None of the extracts produced < 50% cell density at the concentrations tested (1.3 - 20 μg/mℓ). In general, the DCM extracts showed a greater decrease in cell density than the HW extracts. The Ea.hy926 cells were the least affected by the extracts in terms of decreased cell density. The DCM extract of U. urens inhibited α-amylase activity in a mixed manner, which was comparable to the percentage inhibition exerted by the commercial drug, acarbose. Both the HW and DCM extracts of U. urens caused a significant (p < 0.05) increase in glucose uptake into C2C12 myotubes. The HW extract of T. vulgaris had a significant (p < 0.05) inhibitory activity against α-glucosidase (mixed). It also caused the uptake of glucose into C2C12 myotubes, which was significantly (p <0.05) more active than insulin. S. officinalis (DCM extract) also inhibited α-glucosidase activity (p < 0.05) in a mixed manner. Its HW extract displayed potent hypoglycaemic potential by causing glucose uptake into C2C12 myotubes, which was more significant (p < 0.05) than the activity of the positive control, insulin. The DCM extract of A. betulina was active against α-glucosidase (non-competitive), which was comparable to the activity of acarbose. Its HW extract also showed a significant (p < 0.05) glucose uptake activity. Furthermore, the DCM extracts of T. officinalis, A. millefolium, Diabetea and HW extracts of T. foenum-graecum and T. officinalis also caused a significant (p < 0.05) increase in glucose uptake into C2C12 myotubes. This study provides evidence for the antidiabetic potential of T. vulgaris and S. officianlis, in terms of antioxidant capacity and potential to prevent of post-prandial hyperglycaemia and alleviate hyperglycaemia by mimicking the action of insulin. In addition, the organic preparation of U. urens is also a potent α-amylase inhibitor. All herbs tested in this study exerted some form of in vitro antidiabetic activity. The Diabetea mixture, as a traditional preparation, did not have a significant antidiabetic capacity. In vitro observations from this study do not support the use of Diabetea as an antidiabetic preparation and reveal that some of the individual extracts prove more efficacious than the herb mixture. / Dissertation (MSc)--University of Pretoria, 2014. / lk2014 / Pharmacology / MSc / Unrestricted

MSc (Chemistry) / Department of Chemistry / Rauvolfia caffra Sond, a species of evergreen trees and shrubs in the dogbane family, (Apocynaceae), is used as a medicinal plant among traditional communities in many countries for the treatment of malaria, diabetes, coughs, gastrointestinal disturbances, skin infections, impotence, insomnia, diarrhoea, dysentery, scabies, worm infections, and both parasitic and microbial infections. Phytochemical studies have revealed that indole alkaloids are the major constituents of the stem bark. However, there are limited studies linking the compounds with the ethnomedicinal uses. The aim of this study is to isolate and characterize bioactive compounds from Rauvolfia caffra Sond. The highest phenolic content found in a fraction was 16.06±0.125 mg GAE/g, while the highest flavonoid content measured was 9.453±0.081 mg QE/g. In the DPPH free radical scavenging activity and reducing power tests, a lowest IC50 value of 0.022±0.003 μg/mL and IC0.5 value 0.518±0.044 μg/mL, respectively, was found. Six compounds were isolated from the stem bark, including lupeol, a pentacyclic tri-terpenoid isolated for the first time from the genus Rauvolfia; raucaffricine, a rare glycoalkaloid of the monoterpenoid indole class; N-methylsarpagine, an indole alkaloid isolated for the time from R. caffra and spegatrine, an indole alkaloid isolated for the first time from R. caffra, respectively. Concerning antimicrobial activity, the highest activity of a fraction was against B. cereus with MIC values as low as 12.5 mg/mL. One fraction at the tested concentration (250 μg/mL) decreased the viability of Plasmodium falciparum (4.149±6.979 %) with an IC50 value of 6.533 μg/mL. The crude extract and some fractions affected the viability of the Trypanosomes at the tested concentration (250 μg/mL), giving -0.133 ± 0.206 %, 11.334 ± 2.692 %, 1.026 ± 0.143 % and 20.769 ± 9.054 % with IC50 values of 18.50 μg/mL, 14.15 μg/mL, 15.58 μg/mL and 34.71 μg/mL, respectively. Furthermore, the fractions did not show significant cytotoxic effects at a concentration of 50 μg/mL. / NRF