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The effect of in vitro digestion on selected biological activities of Hypoxis sobolifera cormsVan Rooyen, Anzel January 2013 (has links)
In South Africa part of the cultural and religious beliefs of the African people is the use of traditional remedies to treat diseases. These remedies are obtained from medicinal plants (Steenkamp, 2003). One of the most frequently traded plants in the Eastern Cape is Hypoxis, commonly known as Afrika patat, or African potato. South African traditional healers instruct patients to brew the fresh Hypoxis corm as a tea and then ingest it (Steenkamp, 2006a). This prompted an investigation into the digestive stability of a traditionally prepared Hypoxis extract. The H. sobolifera extracts were digested using a simulated gastric/small intestinal digestion and their biological activity determined. The hot water H. sobolifera extract before digestion only showed cytotoxic activity against cancer cell lines at very high concentrations which are not likely to be achieved under normal ingestion circumstances. In Chang liver cells on the other hand, chronic exposure to the hot water H. sobolifera extract increased glucose uptake in amounts similar to that of metformin. On the negative side, the glucose utilization stimulation was lost due to the simulated digestion process. The significant inhibition of AGEs by hot water H. sobolifera extract (IC50 of 6.3 Ig/ml) is a very encouraging result as treatment in the management of diabetes. This activity was only slightly reduced by the in vitro digestion process. Also observed was enzyme inhibition activity by traditionally prepared H. sobolifera, with ∝-amylase being inhibited (IC50 of approximately 250 Ig/ml) and therefore preventing or limiting starch breakdown. From the DPPH results it was clear that H. sobolifera, even when digested, is a potent anti-oxidant (IC50 of 134.4 Ig/ml when undigested compared to 162.9 when digested with β-glucosidase added to stomach digestive step). HPLC and TLC experiments revealed that rooperol which has previously been thought to be the compound responsible for the anti-oxidant activity in Hypoxis extracts, was absent from the traditional extract of H. sobolifera and therefore cannot be the sole compound exhibiting anti-oxidant activity; other compounds such as phenolics may be contributing. The phenolic and flavonoid content results revealed very highconcentrations of these compounds in the traditionally prepared H. sobolifera extract. These compounds may therefore play major roles in all of the biological activities observed from treatment with Hypoxis spp. The ROS results yielded interesting and promising results. Using standard or traditionally prepared H. sobolifera extracts, activation of differentiated U937 cells with PMA was greatly enhanced by cotreatment with the extracts, while extracts on their own did not cause significant activation. Future studies should investigate this property of the extracts as a promising immune boosterThe HPLC results showed that hypoxoside was undetectable in the hot water traditional extract and the TLC anti-oxidant experiment proved that rooperol is not present in the hot water traditional extract after treatment with β-glucosidase. This indicates that neither one of the Hypoxis compounds previously believed to be responsible for the biological activities observed are present in the extract when prepared the traditional way. Therefore, the biological activities observed in this study can be attributed to other phytochemical compounds.
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The in vitro biological activities of three Hypoxis species and their active compoundsBoukes, Gerhardt Johannes January 2010 (has links)
The African potato is used as an African traditional medicine for its nutritional and medicinal properties. Most research has been carried out on H. hemerocallidea, with very little or nothing on other Hypoxis spp. The main aim of this project was to provide scientific data on the anticancer, anti-inflammatory and antioxidant properties of H. hemerocallidea, H. stellipilis and H. sobolifera chloroform extracts and their active compounds. The hypoxoside and phytosterol contents of the three Hypoxis spp. were determined using TLC, HPLC and GC. H. hemerocallidea and H. sobolifera chloroform extracts contained the highest amounts of hypoxoside and β-sitosterol, respectively. For the anticancer properties, cytotoxicity of the Hypoxis extracts and its purified compounds were determined against the HeLa, HT-29 and MCF-7 cancer cell lines (using MTT), and PBMCs (using CellTiter-Blue®). H. sobolifera had the best cytotoxicity against the three cancer cell lines, whereas H. stellipilis stimulated HeLa and HT-29 cancer cell growth. IC50 values of hypoxoside and rooperol were determined. DNA cell cycle arrest (using PI staining) occurred in the late G1/early S (confirmed by increased p21Waf1/Cip1 expression) and G2/M phases after 15 and 48 hrs, respectively, when treated with Hypoxis extracts and rooperol. H. sobolifera and rooperol activated caspase-3 and -7 (using fluorescently labelled antibodies) in HeLa and HT-29 cancer cells, and caspase-7 in MCF-7 cancer cells after 48 hrs. Annexin V binding to phosphatidylserines in rooperol treated U937 cells confirmed early apoptosis after 15 hrs. The TUNEL assay showed DNA fragmentation in the three cancer cell lines when treated with H. sobolifera and rooperol for 48 hrs. A shift pass the G2/M phase has led to the investigation of endoreduplication, which was confirmed by cell/nucleus size, and anti-apoptotic proteins (Akt, phospho-Akt, phospho-Bcl-2 and p21Waf1/Cip1). U937 cell differentiation to monocyte-macrophages was optimized using PMA and 1,25(OH)2D3, which was confirmed by morphological and biochemical changes. For the anti-inflammatory properties, Hypoxis extracts and rooperol significantly increased NO production in monocyte-macrophages (pre-loaded with DAF-2 DA) and phagocytosis of pHrodoTM E. coli BioParticles®. The treatments had no effect on COX-2 expression in monocyte-macrophages. The phytosterols significantly increased IL-1β and IL-6 secretion xv (using the FlowCytomix Multiplex human Th1/Th2 10plex Kit I) in the PBMCs of one donor. For the antioxidant properties, Hypoxis extracts and rooperol significantly increased ROS production in undifferentiated and differentiated U937 cells, which were pre-loaded with DCFH-DA. Hypoxis extracts and purified compounds had ferric reducing activities, but only rooperol had ferric reducing activities significantly greater than ascorbic acid. β-sitosterol, campesterol and cholesterol significantly increased SOD activity in Chang liver cells, while H. stellipilis, H. sobolifera and rooperol decreased SOD activity. Anticancer, anti-inflammatory and antioxidant properties of the Hypoxis extracts may be attributed to the β-sitosterol content, because Hypoxis chloroform extracts contained very little or no hypoxoside. Unidentified compounds, and synergistic and additive effects of the compounds may have contributed to the biological effects. This study confirms previous reports that rooperol is the active compound. Results provide scientific data on the medicinal properties of one of the most frequently used medicinal plants in South Africa.
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The biochemical effects of Hypoxis hemerocallidea in the kidney and liver of streptozotocin-induced diabetic male Wistar ratsGoboza, Mediline January 2015 (has links)
Thesis (MTech (Biomedical Technology))--Cape Peninsula University of Technology, 2015. / Diabetes mellitus (DM) is an endocrine disorder that is characterised not only by severe hyperglycemia but also altered metabolism of glucose and lipids. It is a major health problem worldwide and its impact is greatly noticed in developing countries due to the lack of adequate medical facilities. Oxidative stress remains the principal factor that actively plays major roles in the onset and progression of diabetes mellitus and its complications. The use of medicinal plants in the treatment of DM has undisputedly gained the attention and interest of researchers throughout the globe mainly because plants have established promising outcomes in the treatment of diabetes. It is evident that the plants’ constituents possess therapeutically potent metabolites that have beneficial effects such as antioxidant, antidiabetic, anticancer, anti-inflammatory and antibacterial activities. Hypoxis hemerocallidea is a native plant that grows in the Southern African regions. H. hemerocallidea is well known for its beneficial medicinal values. In South Africa it is known as the African potato. The main aim of this study was to investigate both the beneficial and also the possible toxic effects of H. hemerocallidea in the kidney and liver tissues of streptozotocin-induced diabetic male Wistar rats by assessing the antioxidant status and selected biochemical parameters in the two studied organs.
Diabetes was induced in overnight fasted rats by administration of a single intraperitoneal injection of STZ at a dosage of 50mg/kg in citrate buffer (0.1 M at 4.5 pH). Hyperglycemia was confirmed 72 hours after induction of diabetes using STZ in rats with glucose levels > 15 mmol/l. Treatment with the plants extract commenced on the fourth day after STZ administration via gastric gavage that was done once a day over a 6 week period. The effects of H. hemerocallidea on glucose, body weight, liver and kidney weights, liver function, kidney function and the oxidative status were evaluated after the feeding period.
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Pharmaceutical analysis and drug interaction studies : African potato (Hypoxis hemerocallidea)Purushothaman Nair, Vipin Devi Prasad January 2006 (has links)
In order for a medicinal product to produce a consistent and reliable therapeutic response, it is essential that the final composition of the product is invariable and that the active ingredient/s is/are present in appropriate, non-toxic amounts. However, due to the complexity involved in the standardization of natural products, quality control (QC) criteria and procedures for the registration and market approval of such products are conspicuously absent in most countries around the world. African Potato (AP) is of great medical interest and this particular plant has gained tremendous popularity following the endorsement by the South African Minister of Health as a remedy for HIV/ AIDS patients. Very little information has appeared in the literature to describe methods for the quantitative analysis of hypoxoside, an important component in AP. It has also been claimed that sterols and sterolins present in AP are responsible for its medicinal property but is yet to be proven scientifically. To-date, no QC methods have been reported for the simultaneous quantitative analysis of the combination, β- sitosterol (BSS)/ stigmasterol (STG)/ stigmastanol (STN), purported to be present in preparations containing AP. The effect of concomitant administration of AP and other herbal medicines on the safety and efficacy of conventional medicines has not yet been fully determined. Amongst the objectives of this study was to develop and validate quantitative analytical methods that are suitable for the assay and quality control of plant material, extracts and commercial formulations containing AP. Hypoxoside was isolated from AP and characterized for use as a reference standard for the quality control of AP products and a stability-indicating HPLC/ UV assay method for the quantitative determination of hypoxoside was developed. In addition, a quantitative capillary zone electrophoretic (CZE) method was developed to determine hypoxoside, specifically for its advantages over HPLC. A HPLC method was also developed and validated for the quantitative analysis of BSS, STG and STN in commercially available oral dosage forms containing AP material or extracts thereof. The antioxidant activity of an aqueous extract of lyophilized corms of AP along with hypoxoside and rooperol were investigated. In comparison with the AP extracts and also with hypoxoside, rooperol showed significant antioxidant activity. The capacity of AP, (extracts, formulations, hypoxoside and rooperol as well as sterols to inhibit in vitro metabolism of drug substrates by human cytochrome P450 (CYP) enzymes such as CYP 3A4, 3A5 and CYP19 were investigated. Samples were also assessed for their effect on drug transport proteins such as P-glycoprotein (P-gp). Various extracts of AP, AP formulations, stigmasterol and the norlignans, in particular the aglycone rooperol, exhibited inhibitory effects on CYP 3A4, 3A5 and CYP19 mediated metabolism.These results suggest that concurrent therapy with AP and other medicines, in particular antiretroviral drugs, can have important implications for safety and efficacy. Large discrepancies in marker content between AP products were found. Dissolution testing of AP products was investigated as a QC tool and the results also revealed inconsistencies between different AP products.
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The effects of the wild african potato (hypoxis hemerocallidea) supplementation on streptozotocin-induced diabetic wistar rats reproductive functionJordaan, Audrey Emmerentia January 2015 (has links)
Thesis (MTech (Biomedical Technology))--Cape Peninsula University of Technology, 2015. / Diabetes mellitus (DM) has been reported to be one of the greatest global public health threats. Statistics of the fertility status of modern society has linked increased DM to a decrease in fertility rates. Hyperglycaemia is characteristic of DM that results in a disturbance of proteins, lipids and carbohydrate metabolism leading to an increase production of reactive oxygen species (ROS). In the case where ROS overwhelms antioxidant mechanisms, the body goes into state of oxidative stress (OS). OS plays a vital role in the progression of DM which leads to dysfunction and damage of various organs including that of the reproductive system. Os has shown to cause damage to the sperm membraneby oxidation of polyunsaturated fatty acids (PUFA’s) as the sperm membrane are rich in PUFA’s. This damage contributes to reduced sperm motility, concentration, morphological abnormalities and the sperms ability to fuse with the ZP of the oocyte. DM has been observed to cause testicular degeneration by interrupting sertoli cell production and maintenance thus resulting in a disturbance of the normal functioning of the reproductive system. Experimental studies have targeted more natural sources for treating DM and its complications of the reproductive system. Plants and natural dietary substances have shown to have high antioxidant contents that combat DM induced oxidative stress. This study explored the effect the Hypoxis hemerocallidea (H. hemerocallidea) supplementation on testicular and epididymal tissue, sperm motility and reproductive hormones in male wistar rats. The experiment were conducted for 6 weeks and the rats (230-260 grams) were randomly divided into 5 groups (n=12 per group). Diabetes was induced in 3 of the 5 groups. The first group was the normal control group (A), second the diabetic control group (B), third was the diabetic group treated with 800mg/kg H. hemerocallidea (group C), fourth the diabetic group treated with 200mg/kg H. hemerocallidea (group D) and fifth the non-diabetic group supplemented with 800mg/kg H. hemerocallidea (group E). Blood glucose showed a significant increase in the diabetic group when compared to the normal control and treated groups. H. hemerocallidea showed improvement in sperm motility and sperm morphology more at 800mg/kg when compared to diabetic group and diabetic group treated with 200mg/kg. Body, testicular and epipidymal weights of diabetic control were significantly lower when compared to the other groups. Testicular and epididymal Malondialdehyde levels were decreased in normal control, diabetic groups treated with different doses of H. hemerocallidea and the non-diabetic group supplemented with H. hemerocallideaon comparing with the diabetic control group. Antioxidants such as Superoxide dismutase, Catalase and total Glutathione activity was observed to be dosage dependent in certin groups but most showed a significant increase when compared to the diabetic control group. The total antioxidant capacity was measured using Oxygen radical absorbance capacity (ORAC) and Ferric ion reducing antioxidant power (FRAP); increase was observed when normal control group and treated groups were compared to the diabetic group. Testosterone and estradiol levels were also increased when the normal control group and treated groups were compared to the diabetic control group.
Based on our findings it can be concluded that H. hemerocallidea supplementation can potentially be used to counteract deleterious effects of DM on the male reproductive system.
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African traditional medicines-antiretroviral drug interactions: the effect of African potato (Hypoxis hemerocallidea) on the pharmacokinetics of efavirenz in humansMogatle, Seloi January 2009 (has links)
African Potato (Hypoxis hemerocallidea), (AP) is an African traditional medicine (TM) that is commonly used for various nutritional/medicinal purposes and also by people infected with the human immuno deficiency virus HIV and AIDS patients as an immune booster. The use of AP has also been recommended by the former Minister of Health of South Africa for use by HIV positive people. The main phytochemical component of AP is a norlignan glucoside, hypoxoside, and other relatively minor components have also been reported. A recent in vitro study reported the effects of AP extracts, hypoxoside and rooperol (the metabolite of hypoxoside) on human metabolic enzymes such as the cytochrome P450 (CYP450) group of enzymes and also on the transporter protein, p-glycoprotein (P-gp). This research focussed on investigating the clinical significance of those in vitro effects on the pharmacokinetics of efavirenz (EFV) in humans. EFV was chosen as the substrate drug because it is in first-line regimen of treatment of HIV/AIDS in South Africa, and also has been reported to be a substrate for the specific CYP isozymes, 3A4 and 2B6, in common with APs metabolic involvement with 3A4. A high performance liquid chromatography method with ultra-violet detection (HPLC-UV) for the quantitative determination of EFV in plasma was developed and successfully validated according to international standards with good reproducibility, accuracy, recovery, linear response and requisite sensitivity. The preparation of the plasma samples for analysis was effected by using a simple and rapid precipitation method, and the mobile phase consisted of readily available solvents. EFV in plasma samples was found to be stable under the relevant storage conditions studied. The oral dose of AP, administered as a freshly prepared traditional decoction, was standardised based on the hypoxoside content, and the quality of all the AP decoctions was analysed immediately prior to administration, using a validated HPLC-UV method. A single dose, two-phase sequential study was conducted over a period of 31 days in 10 healthy volunteers. The clinical study was approved by the Rhodes University Ethical Standards Committee, and all the participants agreed to the conditions of the study by giving their informed consent. On day 1 of the study, human subjects were administered a 600 mg EFV tablet and blood samples were collected before dosing and at various intervals over a period of 48 hr post dosing. From day 16, a traditionally prepared AP decoction was administered daily at a standardized dose of 15 mg/kg/day per subject until day 30. On day 29, volunteers were administered a single 600 mg dose of EFV as was done on day 1. Plasma samples were harvested immediately after blood sample collection and frozen at -80 ºC until assayed. Geometric mean ratios of relevant pharmacokinetic parameters, Cmax (maximum plasma concentration achieved following dosing) and AUC0-48 (area under the curve of a plot of drug plasma concentrations versus time representing the extent of absorption) of EFV before and after co-administration of 14 successive daily doses of AP were compared and evaluated to determine whether an interaction had occurred. All subjects completed the study and the geometric mean ratios of Cmax and AUC0-48 were 97.30 and 102.82 with corresponding 90% confidence intervals (CIs) of 78.81-120.14% and 89.04-118.80%, respectively. Whereas the acceptance criteria for the ratios of the AUCs fell within the preset 90% CIs indicating no interaction, the Cmax ratios fell outside the limits. Although the protocol was developed in accordance with the United States of America Food & Drug Administration’s Guidance for Drug Interactions, a priori stating that both criteria need to fall within the acceptance limits to indicate no interaction, an argument is presented to waive the Cmax requirement for the declaration of an interaction. As a result, the pharmacokinetic data generated during this study indicated that the effect of AP on the pharmacokinetics of EFV is not clinically significant. Hence, co-administration of AP is unlikely to affect the clinical use of EFV. In summary the objectives of this project were: 1. To develop and validate a suitable HPLC-UV method for the quantitative determination of EFV in plasma. 2. To perform a mini-validation of the determination of hypoxoside for use as a marker in the quality control and standardisation of AP decoctions. 3. To conduct a clinical interaction study in order to determine whether AP affects the pharmacokinetics of EFV following concurrent administration. 4. To apply the validated HPLC-UV method to determine plasma concentrations of EFV in plasma of human subjects. 5. To use appropriate statistical methods and treatments such as a non-compartmental pharmacokinetic analysis to determine the occurrence of an interaction.
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