Thesis (MSc (Biochemistry))--University of Stellenbosch, 2009. / Sutherlandia frutescens (S. frutescens), sub-species microphylla, is a member of the Fabacea family and is used as a herbal remedy for the treatment of several ailments which include influenza, diabetes, cancer, tuberculosis, chronic fatigue syndrome, rheumatoid arthritis, anxiety, clinical depression, and more recently, those living with human immunodeficiency virus/ acquired immune deficiency syndrome (HIV/AIDS) (1-4). Many of the symptoms of these ailments are associated with a perturbation of the stress response which may be associated with disorders of the endocrine system. Of all the traditional plants in South Africa, S. frutescens is regarded the most profound in that it is a multipurpose traditional remedy. The plant has enjoyed a long history of use and reports indicating its efficacy as a safe treatment for various health conditions have added to the popularity of this medicinal plant. The extracts of S. frutescens have been shown to exhibit anti-proliferative effects on cancer cells, antioxidant activity, and to possess anti-diabetic and anti-inflammatory potential (5, 6), providing scientific evidence for its therapeutic use in the treatment of cancer and diabetes. However, this study focuses on the potential use of this medicinal plant in the treatment of stress and stress related diseases. Chronic stress is characterized by elevated plasma levels of glucocorticoids. These steroid hormones are synthesized in the adrenal cortex in a series of reactions involving the steroidogenic enzymes. The major aim of this thesis was the determination of the influence of S. frutescens extracts on the adrenal cytochrome P450 enzymes. Aqueous, methanol and chloroform S. frutescens extracts were prepared and the interaction with the cytochrome P450 enzymes was investigated. The effect of these extracts towards progesterone (PROG), deoxycortisol and deoxycorticosterone (DOC) binding to the cytochrome P450 enzymes as well as their influence on the metabolism of these steroid substrates was investigated. A similar study (7) showed that compounds from the S. frutescens extracts could interact with these enzymes and possibly affect adrenal steroidogenesis. This study further investigates the bioactive properties of the plant material in terms of the influence of S. frutescens on the cytochrome P450 enzymes and the effect of the manufacturing process on the bioactivity of the plant.

Magister Pharmaceuticae - MPharm / Sutherlandia frutescens (tribe Galegeae, Fabaceae), is a popular medicinal plant traditionally used in South Africa. In 2000, a company called Phyto Nova (Pty) Ltd. initiated large-scale cultivation and contract manufacturing of tablets, made from the powdered herb (i.e. thin stems and leaves). Most of these commercial Sutherlandia solid dosage forms are made from the dried leaf powder but recently a new product, viz. Promune™ capsules, made from a freeze-dried aqueous extract, came on the market and was claimed to be “better” as it mimics the traditional tea. However, the pharmaceutical quality and stability of these preparations have not yet been investigated. The objectives of this study were firstly, to develop a validated stability-indicating HPLC assay for sutherlandioside B (SU-B); secondly, to compare the SU-B levels in the two commercially available Sutherlandia products viz, the Phyto Nova Sutherlandia SU1™ tablet and the Promune™ capsule, and, thirdly, to determine the effect of elevated temperature and humidity as well as acid hydrolysis on the SU-B levels in these two products. / South Africa

<p>Sutherlandia frutescens (tribe Galegeae, Fabaceae), is a popular medicinal plant traditionally used in South Africa. In 2000, a company called Phyto Nova (Pty) Ltd. initiated large-scale cultivation and contract manufacturing of tablets, made from the powdered herb (i.e. thin stems and leaves). Most of these commercial Sutherlandia solid dosage forms are made from the dried leaf powder but recently a new product, viz. Promune&trade / capsules, made from a freeze-dried aqueous extract, came on the market and was claimed to be &ldquo / better&rdquo / as it mimics the traditional tea. However, the pharmaceutical quality and stability of these preparations have not yet been investigated. The objectives of this study were firstly, to develop a validated stability-indicating HPLC assay for sutherlandioside B (SU-B) / secondly, to compare the SU-B levels in the two commercially available Sutherlandia products viz, the Phyto Nova Sutherlandia SU1&trade / tablet and the Promune&trade / capsule, and, thirdly, to determine the effect of elevated temperature and humidity as well as acid hydrolysis on the SU-B levels in these two products.</p>

<p>Endocrine&ndash / disrupting compounds (EDCs) are exogenous compounds/chemicals which interfere with, or have adverse effects on the production, distribution and function of natural hormones, thereby affecting normal endocrine activity, health and quality of life of both humans and wildlife. The reproductive system is highly susceptible to EDCs due to it being controlled by an array of hormonal signals. The effects of EDCs on the male reproductive system include infertility, decreased sperm count, function and morphology, abnormal development of secondary sex characteristics, reproductive function and sexual behaviour as well as decreased libido. There are various sources by which EDCs enter the environment which include effluents from several industries (mining, agriculture, smelting, hazardous waste sites, manufacturing industries, etc.), sewage treatment effluents, urban and agricultural runoff and effluents which include natural and pharmaceutical chemicals excreted in the urine of humans and domestic livestock, pesticides, polychlorinated biphenyls, dioxins, plasticizers, surfactants, etc. Humans and animals can also be affected by EDCs by consuming food containing endocrine active substances. The growing concern regarding adverse effects due to EDC exposure of humans and wildlife, as well as the increased incidence of EDC contamination has prompted extensive research into the development and validation of screening tests to detect and monitor known EDCs and new substances with endocrine-disrupting capability. These screening tests involve assessing the effect of known and potential EDCs on reproductive function and development as well as&nbsp / hormone production. To assess the effect of EDCs on the reproductive system different methods are employed which include in vitro, in vivo and ex vivo methods. In vitro methods have been suggested as a suitable screening tool for EDC monitoring due to low costs, reduced animal usage, the use of standard and basic equipment as well as the ability to screen a large number of samples with multiple endpoints. Of the available in vitro methods, the minced testes method has been suggested as the most suitable method for screening EDCs and for this reason has been employed in this study. The aim of this study was thus to employ a minced testes method to screen samples for male reproductive toxicity using cell viability and hormone production (testosterone and estradiol) as endpoints.The first objective of this study was to optimize an in vitro testicular cell culture assay by determining both optimal luteinizing hormone (LH)&nbsp / concentration and incubation time needed for testosterone production. Testicular cell cultures were prepared and cells were treated with varying concentrations of LH (10, 1, 0.1, 0.01 and 0 mu/ml) and incubated for 4 hours and 20 hours. Testosterone production was evaluated for each incubation period. Testosterone production was significantly increased for both incubation periods at all LH concentrations tested as compared to the control. For both incubation periods, there was no significant difference in testosterone production between the different LH concentrations tested. From the data obtained, the 4 hour incubation period as well as the LH concentration of 10 mu/ml were selected as optimal for the testicular cell culture assay. The second objective of this study was to determine the effect of Tulbaghia violacea Harv. on the male reproductive system. T. violacea is a plant species indigenous to southern Africa and is used locally as a herbal remedy/medicine to treat several ailments. Cells were treated with varying concentrations of the T. violacea ethanol extract (with/without LH-treatment) and incubated for 4 hours. Hormone production and cell viability were evaluated. The results obtained from this pilot in vitro study demonstrated that the ethanol extract of T.violacea has androgenic properties by significantly increasing LH-induced testosterone production in mouse testes with no significant change in cell viability. The third objective of this study was to assess the effect of Sutherlandia frutescens(L.) R.Br and Artemisia afra Jacq. Ex Willd. on the male reproductive system. S. frutescens and A. afra are also plant species indigenous to southern Africa and used locally as a herbal remedy/medicine to treat several ailments. Ethanol extracts of each plant was prepared and cells were treated with varying concentrations of each extract (0, 156.25, 312.5, 625, 1250,2500 and 5000 &mu / g/ml) with or without LH-treatment and incubated for 4 hours. Cytotoxicity by LDH measurement and hormone production (testosterone and estradiol) were endpoints that were evaluated. The results obtained showed that the ethanol extracts of both plants are not cytotoxic to testicular cells and that A. afra decreases testosterone production at high concentrations. The fourth and final objective of this study was to assess the acute effect of four heavy metals, namely manganese, copper, cadmium and magnesium on the male reproductive system. These heavy metals are used extensively in manufacturing and mining industries. Cells were treated with varying concentrations of each metal salt (200, 100, 50, 25, 12.5, and 6.25&nbsp / &mu / M) with or without LH-treatment and incubated for 4 hours. Endpoints evaluated included cell viability, testosterone and estradiol production. The results obtained showed that manganese, cadmium and copper are highly toxic to testicular cells in vitro and therefore may potentially cause reproductive toxicity.</p>

<p>Sutherlandia frutescens (tribe Galegeae, Fabaceae), is a popular medicinal plant traditionally used in South Africa. In 2000, a company called Phyto Nova (Pty) Ltd. initiated large-scale cultivation and contract manufacturing of tablets, made from the powdered herb (i.e. thin stems and leaves). Most of these commercial Sutherlandia solid dosage forms are made from the dried leaf powder but recently a new product, viz. Promune&trade / capsules, made from a freeze-dried aqueous extract, came on the market and was claimed to be &ldquo / better&rdquo / as it mimics the traditional tea. However, the pharmaceutical quality and stability of these preparations have not yet been investigated. The objectives of this study were firstly, to develop a validated stability-indicating HPLC assay for sutherlandioside B (SU-B) / secondly, to compare the SU-B levels in the two commercially available Sutherlandia products viz, the Phyto Nova Sutherlandia SU1&trade / tablet and the Promune&trade / capsule, and, thirdly, to determine the effect of elevated temperature and humidity as well as acid hydrolysis on the SU-B levels in these two products.</p>

<p>Endocrine&ndash / disrupting compounds (EDCs) are exogenous compounds/chemicals which interfere with, or have adverse effects on the production, distribution and function of natural hormones, thereby affecting normal endocrine activity, health and quality of life of both humans and wildlife. The reproductive system is highly susceptible to EDCs due to it being controlled by an array of hormonal signals. The effects of EDCs on the male reproductive system include infertility, decreased sperm count, function and morphology, abnormal development of secondary sex characteristics, reproductive function and sexual behaviour as well as decreased libido. There are various sources by which EDCs enter the environment which include effluents from several industries (mining, agriculture, smelting, hazardous waste sites, manufacturing industries, etc.), sewage treatment effluents, urban and agricultural runoff and effluents which include natural and pharmaceutical chemicals excreted in the urine of humans and domestic livestock, pesticides, polychlorinated biphenyls, dioxins, plasticizers, surfactants, etc. Humans and animals can also be affected by EDCs by consuming food containing endocrine active substances. The growing concern regarding adverse effects due to EDC exposure of humans and wildlife, as well as the increased incidence of EDC contamination has prompted extensive research into the development and validation of screening tests to detect and monitor known EDCs and new substances with endocrine-disrupting capability. These screening tests involve assessing the effect of known and potential EDCs on reproductive function and development as well as&nbsp / hormone production. To assess the effect of EDCs on the reproductive system different methods are employed which include in vitro, in vivo and ex vivo methods. In vitro methods have been suggested as a suitable screening tool for EDC monitoring due to low costs, reduced animal usage, the use of standard and basic equipment as well as the ability to screen a large number of samples with multiple endpoints. Of the available in vitro methods, the minced testes method has been suggested as the most suitable method for screening EDCs and for this reason has been employed in this study. The aim of this study was thus to employ a minced testes method to screen samples for male reproductive toxicity using cell viability and hormone production (testosterone and estradiol) as endpoints.The first objective of this study was to optimize an in vitro testicular cell culture assay by determining both optimal luteinizing hormone (LH)&nbsp / concentration and incubation time needed for testosterone production. Testicular cell cultures were prepared and cells were treated with varying concentrations of LH (10, 1, 0.1, 0.01 and 0 mu/ml) and incubated for 4 hours and 20 hours. Testosterone production was evaluated for each incubation period. Testosterone production was significantly increased for both incubation periods at all LH concentrations tested as compared to the control. For both incubation periods, there was no significant difference in testosterone production between the different LH concentrations tested. From the data obtained, the 4 hour incubation period as well as the LH concentration of 10 mu/ml were selected as optimal for the testicular cell culture assay. The second objective of this study was to determine the effect of Tulbaghia violacea Harv. on the male reproductive system. T. violacea is a plant species indigenous to southern Africa and is used locally as a herbal remedy/medicine to treat several ailments. Cells were treated with varying concentrations of the T. violacea ethanol extract (with/without LH-treatment) and incubated for 4 hours. Hormone production and cell viability were evaluated. The results obtained from this pilot in vitro study demonstrated that the ethanol extract of T.violacea has androgenic properties by significantly increasing LH-induced testosterone production in mouse testes with no significant change in cell viability. The third objective of this study was to assess the effect of Sutherlandia frutescens(L.) R.Br and Artemisia afra Jacq. Ex Willd. on the male reproductive system. S. frutescens and A. afra are also plant species indigenous to southern Africa and used locally as a herbal remedy/medicine to treat several ailments. Ethanol extracts of each plant was prepared and cells were treated with varying concentrations of each extract (0, 156.25, 312.5, 625, 1250,2500 and 5000 &mu / g/ml) with or without LH-treatment and incubated for 4 hours. Cytotoxicity by LDH measurement and hormone production (testosterone and estradiol) were endpoints that were evaluated. The results obtained showed that the ethanol extracts of both plants are not cytotoxic to testicular cells and that A. afra decreases testosterone production at high concentrations. The fourth and final objective of this study was to assess the acute effect of four heavy metals, namely manganese, copper, cadmium and magnesium on the male reproductive system. These heavy metals are used extensively in manufacturing and mining industries. Cells were treated with varying concentrations of each metal salt (200, 100, 50, 25, 12.5, and 6.25&nbsp / &mu / M) with or without LH-treatment and incubated for 4 hours. Endpoints evaluated included cell viability, testosterone and estradiol production. The results obtained showed that manganese, cadmium and copper are highly toxic to testicular cells in vitro and therefore may potentially cause reproductive toxicity.</p>

Magister Pharmaceuticae - MPharm

Since ancient times, indigenous plants have been used by traditional healers for treating various ailments. Sutherlandia frutescens is one of the most commonly used medicinal plants of southern Africa. This widely distributed plant has been traditionally used to treat cancer and HIV patients; however scientific validation is still in high demand. This research aimed to phytochemically characterise the various extracts prepared and to determine if any chemotypes were present. Subsequent biological characterisation was carried out to preliminary ascertain whether this medicinal plant could have anti-cancer and/or immunemodulating properties and which compounds might be responsible for these actions. Various traditional and organic extracts were prepared. Extracts, fractions and compounds generated were analysed and chemical profiles obtained. Column chromatographic techniques were used to isolate and purify compounds and structure elucidation was carried out using various analytical techniques. Sulforhodamine B and cytometric bead array assays were performed to determine the biological activities of samples generated. / Life and Consumer Sciences / (M. Sc. (Life Sciences)

Since ancient times, indigenous plants have been used by traditional healers for treating various ailments. Sutherlandia frutescens is one of the most commonly used medicinal plants of southern Africa. This widely distributed plant has been traditionally used to treat cancer and HIV patients; however scientific validation is still in high demand. This research aimed to phytochemically characterise the various extracts prepared and to determine if any chemotypes were present. Subsequent biological characterisation was carried out to preliminary ascertain whether this medicinal plant could have anti-cancer and/or immunemodulating properties and which compounds might be responsible for these actions. Various traditional and organic extracts were prepared. Extracts, fractions and compounds generated were analysed and chemical profiles obtained. Column chromatographic techniques were used to isolate and purify compounds and structure elucidation was carried out using various analytical techniques. Sulforhodamine B and cytometric bead array assays were performed to determine the biological activities of samples generated. / Life and Consumer Sciences / (M. Sc. (Life Sciences)

Traditional medicines prepared from Sutherlandia frutescens are used to manage diseases including HIV and cancer. This study aimed at isolating and identifying biologically active compounds isolated from S. frutescens. Sutherlandia frutescens plants were collected in Petrusburg and Paarl. Powdered plant material was extracted using ethanol or water and their metabolite composition was compared using UPLC-MS. A novel cycloartane, an acetylated variant of this compound as well as a Sutherlandioside B triterpenoid was isolated and characterised using chromatographic and analytical techniques such as NMR and UPLC-MS. Preliminary biological studies were conducted to assess the activity of plant extracts on cell toxicity, herpes virus replication and cytokine expression. The results of this study suggest that aqueous extracts from S. frutescens do not appear to be cytotoxic or show anti-herpetic activity, but may activate the immune system by increasing expression of IL-6, IL-10 and TNFα. Further research should be conducted to confirm and optimise these results. / Life and Consumer Sciences / M. Sc. (Life Science)