published_or_final_version / Community Medicine / Master / Master of Public Health

Thesis (PhD(FoodSc))--Stellenbosch University, 2012. / ENGLISH ABSTRACT: Potential beneficial and/or adverse modulatory effects of polyphenol-enriched extracts of rooibos (Aspalathus linearis) and honeybush (Cyclopia spp.) on the antioxidant homeostasis in the liver were investigated. Phase II metabolism of aspalathin and mangiferin, the major polyphenols of rooibos and honeybush respectively, was assessed for potential glucuronidation and sulphation. Glucuronidation resulted in a loss of antioxidant activity for aspalathin and mangiferin in post-column HPLC-DAD-DPPH• and HPLC-DAD-ABTS•+ assays and also a decreased activity of iron chelating properties of mangiferin in the FRAP assay. Two independent studies for 28 and 90 days with polyphenol-enriched extracts (PEEs) of unfermented rooibos [Aspalathus linearis (PER)] and honeybush [Cyclopia. subternata (PECsub) and C. genistoides (PECgen)] in male Fischer rats were conducted to assess possible beneficial and/or adverse biological effects. PECgen was only included in the 28 day study. PEEs were characterised by in vitro antioxidant assays and HPLC analysis. The importance of detailed chemical characterization of rooibos and honeybush when investigating biological effects in vivo is clear as distinctive biological effects and major differences in compositions were evident. Biological parameters included were serum chemical parameters, activities of selected antioxidant enzymes, levels of glutathione and the modulation of expression of oxidative stress and antioxidant defense related genes in the liver. Sub-chronic (90 days) exposure of rats to PER and PECsub both adversely affected iron absorption significantly (P<0.05) and significantly (P<0.05) and markedly lowered levels of reduced glutathione (GSH) in the liver. The high levels of polyphenol intake were implicated and interaction with glutathione was postulated to occur via catechol o-quinone conjugations with GSH. This was also implicated in the significantly (P<0.05) increased activity of glutathione reductase (GR) following 28 days. These findings suggest that PEEs from rooibos and honeybush have the potential to alter the glutathione homeostasis, which could contribute to oxidative status in the liver. PECsub resulted in alterations in the liver biliary system which was manifested as significantly (P<0.05) increased serum total bilirubin (Tbili) and alkaline phosphatase (ALP), depending on the age of the rats, level of total polyphenols and duration of exposure. The expression of a number of oxidative stress and antioxidant defense related genes were differentially altered by the PEEs of rooibos and honeybush in rat liver and further indicated potential oxidative stress. Modulatory effects of PEEs on expression of 84 of these genes in rat liver were assessed with a quantitative real-time reverse transcription polymerase chain reaction (RT-PCR) array and provided additional insights into the possible adverse and protective effects of rooibos and honeybush. Further investigation on total polyphenol dose levels and time of exposure in the application of PEEs of rooibos and honeybush as dietary supplements and functional foods is recommended and will also be of value in anticipated regulatory requirements for future substantiation of safety and efficacy. / AFRIKAANSE OPSOMMING: Die moontlike voordelige en/of nadelige modulerende effekte van polifenol-verrykde ekstrakte van rooibos (Aspalathus linearis) en heuningbos (Cyclopia spp.) op die antioksidant homeostasis in die lewer is ondersoek. Die fase II metaboliete van aspalatien en mangiferin, die hoof verbindings in rooibos en heuningbos onderskeidelik, is ondersoek t.o.v. glukuronidering en sulfonering. Glukuronidasie het gelei tot n verlies in antioksidant aktiwiteit van aspalatien en mangiferin soos bepaal in post-kolom HPLC-DAD-DPPH• en HPLC-DADABTS•+ toetse, asook verminderde interaksie met yster van mangiferin in die FRAP toets. Twee onafhanklike studies van 28 en 90 dae is onderneem met polifenol-verrykde ekstrakte (PVEs) van ongefermenteerde rooibos [Aspalathus linearis (PVR)] en heuningbos [Cyclopia. subternata (PVCsub) and C. genistoides (PVCgen)] in manlike Fisher rotte om die moontlike voordelige en/of nadelige biologiese effekte te ondersoek. PECgen was slegs in die 28 dae studie ingesluit. PVEs is gekarrakteriseer deur in vitro antioksidant en HPLC analises. Die belang van chemiese karaktirisering van rooibos en heuningbos tydens ondersoeke na biologiese aktiwiteit is duidelik aangesien verskeie en variërende biologiese aktiwiteite en verskille in die komposisie in die huidige studie gesien is. Die biologiese parameters wat ondersoek is om die effek van die PVEs te bepaal het serum kliniese parameters, aktiwiteit van geselekteerde ensieme, glutatioon en evaluering van die ekspressie van oksidatiewe en antioksidant verwante gene in die lewer, ingesluit. Sub-kroniese (90 dae) blootstelling van rotte aan PVR en PVCgen het yster absorpsie negatief beïnvloed. Die beduidende (P<0.05) verlaagde vlak van gereduseerde glutatioon in die lewer was toegeskryf aan die hoë vlakke van polifenole ingeneem tydens die studie en word moontlik veroorsaak deur n spesifieke katekol o-konjugasie van GSH. Hierdie interaksie was ook moontlik die oorsaak van n beduidende (P<0.05) toename in die aktiwiteit van glutatioon reduktase. Dié bevindinge het moontlike implikasies t.o.v die glutatioon homeostase en is n moontlike indikase dat PVEs van rooibos kan bydra tot oksidatiewe stres. PVCsub het veranderinge in die lewer gal-sisteem tot gevolg gehad aangesien daar n beduidende (P<0.05) verhoging in die serum totale bilirubin en alkalien fosfaat was. Hierdie veranderinge is beïnvloed deur die ouderdom, vlakke van die totale polifenole en die periode van blootstelling. Die uitdrukking van oksidatiewe en antioksidant verwante gene is op verskillende maniere beïnvloed deur die PVEs van rooibos en heuningbos in rot lewer and dien as n verdere indikasie van onderliggende oksidatiewe stres. Die modulerende effekte van PVEs op geenuitdrukking het gelei tot additionele insig aangaande die moontlike skadelike of beskermende eienskappe van PVEs vir gebruik as kruie produkte of dieet aanvullings. Die indikasies van moontlike oksidatiewe stres was duidelik van biologiese parameters en modulering van geenuitdrukking in die lewer, en vereis verdere ondersoek na die polifenool dosis en periode van toediening voordat PVEs van rooibos en heuningbos as funksionele voedsel produkte gebruik word. Hierdie ondersoek sowel as toekomstige ondersoeke in hierdie verband sal van waarde wees vir regulatoriese vereistes omtrent die veiligheid en effektiwiteit van rooibos en heuningbos kruie produkte.

Thesis submitted in fulfillment of the requirements for the degree Master of Technology: Biomedical Technology In the Faculty of Health and Wellness Sciences At the Cape Peninsula University of Technology, 2012 / Human and experimental animal studies have shown that biomarkers of oxidative damage are elevated in subjects with certain diseases or risk factors. Consequently, it is hypothesized that oxidative stress plays an important role in the pathogenesis of these diseases and that dietary intake of, or supplementation with antioxidants may be protective or be useful therapeutic targets. This study was designed to investigate the modulatory effect of Camellia sinensis (Chinese green tea), Aspalathus linearis (rooibos herbal tea) and the two commercial supplements on the antioxidant status of the liver and kidney of tert-butyl hydroperoxide (t-BHP)-induced oxidative stress male Wistar rats. Rooibos and green tea are beverages well-known for their antioxidant content. Based on the specific beverage consumed, sixty male Wistar rats were randomly assigned into six groups, i.e. fermented rooibos (FRT), unfermented rooibos (URT), Chinese green tea (CGT), rooibos supplement (RTS), Chinese green tea supplement (GTS) and control (CTL). The animals had free access to the respective beverages and standard diet for 10 weeks, while oxidative stress was induced during the last 2 weeks via intraperitoneal injection of 30 μM of t-BHP per 100 g body weight. Among all the beverage and/or supplement preparations, the commercial rooibos supplement had the highest total polyphenol content and antioxidant activity while fermented rooibos, as previously shown, had a lower antioxidant content and potency when compared to its unfermented counterpart. The ability of these beverages and/or supplements to modulate the antioxidant status in tissues was organ specific and varied according to the assessment method. When considering the liver, the intake of unfermented rooibos, Chinese green tea and the commercial rooibos supplement significantly (P<0.05) restored the t-BHP-induced reduction and increased the antioxidant status with regards to oxygen radical absorbance capacity and trolox equivalent antioxidant capacity (TEAC) levels. All the beverages and/or supplements also significantly (P<0.05) enhanced the renal antioxidant capacity as assessed by the TEAC assay. In what may be an indication of decreased oxidative stress, all the beverages were associated with a general decline in activities of the antioxidant enzymes which reached significant levels in renal superoxidase dismutase activity. Generally, the beverages did not impact significantly on lipid peroxidation (LPO) although there were differing trends in the two LPO markers assessed. While thiobarbituric acid reactive substances levels showed a declining trend in both tissues, the conjugated dienes were generally elevated. In conclusion, this study confirms Camellia sinensis and Aspalathus linearis as well as their two supplements as good sources of dietary antioxidants and results demonstrated that rooibos and green tea improved the liver and kidney antioxidant capacity of oxidative stress-induced rats. Their impact on antioxidant status in rats was shown to vary between organs and according to the method of assessment. Hence multi-method, multi-organ assessment may be a more informative approach in in vivo antioxidant studies.

Thesis (MTech (Biomedical Technology))--Cape Peninsula University of Technology, 2014. / Over the past few decades, it has been shown that various critical diseases including heart disease, cancer, and diabetes associated with free radical generation and low endogenous antioxidant capacity, lead to oxidative stress and cell injury. In recent years, numerous studies have also reported that antioxidants, present in various beverages, vegetables and some foods have attracted a significant research interest due to their potential benefits to human health. However, epidemiological evidence shows a correlation between the intake of food rich in antioxidants and the reduced incidence of some mortality of chronic diseases, certain cancers and coronary heart disease. The aims of this study were to determine the effects of rooibos teas (fermented and unfermented) and green tea as a comparison on the biochemical parameters and the trace element absorption in a rat model. In this study 4 groups of experimental animals were used. All groups had ad libitum access to standard rat chow. Group A, the controls (11 animals), were fed with tap water; group B (11 animals) were fed with the liquid extract of fermented rooibos tea; group C (9 animals) were fed with the liquid extracts of unfermented rooibos and group 0 (9 animals) were fed with the liquid extract of green tea. All groups were fed for a period of 10 weeks. After the feeding period, the animals were sacrificed by euthanization with intraperitoneal injections of pentobarbital. Blood was sampled by cardiac puncture and centrifuged to obtain the serum. Some elemental analyses were performed with X-ray emission and backscattering. ICP-OES was used to determine the magnesium content. For X-ray emission, backscattering and ICP-OES analyses, 100 µL of each serum sample in a group were added to 2 ml freeze-drying tube. Of the combined specimen, 100 µL was used for the magnesium determination by ICP-OES. The remainder of the combined serum specimens for each group were freeze-dried at -80°C and then pressed into a pellet. The pellet was coated with carbon and analyzed using X-ray emission and backscattering. The elemental X-rays of P, S, Ca, Mn, Fe, Cu, Co, Zn, Mo, Ca and Se emitted were quantified to obtain the respective concentrations. Biochemical chemistry analyses were performed on each serum sample of each animal. The biochemical parameters tested for were total protein, albumin, globulin, total bilirubin, lactate dehydrogenase, blood urea nitrogen, uric acid, total cholesterol, aspartate aminotransferase, alanine aminotransferase, creatine phosphokinase and creatinine.

Indiana University-Purdue University Indianapolis (IUPUI) / Down syndrome (DS), a genetic disorder that affects ~1 in 700 live births, is caused by trisomy of human chromosome 21 (Hsa21). Individuals with DS are affected by a wide spectrum of phenotypes which vary in severity and penetrance. However, cognitive and skeletal impairments can be commonly observed in all individuals with DS. To study these phenotypes, we utilized the Ts65Dn mouse model that carries three copies of approximately half the gene orthologs found on Hsa21 and exhibit similar phenotypes as observed in humans with DS. Individuals with DS and Ts65Dn mice have deficits in bone mineral density (BMD), bone architecture, bone strength, learning and memory. Over-expression of DYRK1A, a serine-threonine kinase encoded on Hsa21, has been linked to deficiencies in DS bone homeostasis and cognition. Epigallocatechin-3-gallate (EGCG), an aromatic polyphenol found in high concentrations in green tea, is a selective inhibitor of DYRK1A activity. Normalization of DYRK1A activity by EGCG therefore may have the potential to ameliorate skeletal and cognitive deficits. We hypothesized that supplements containing EGCG obtained from health food stores/ online vendors will not be as effective as EGCG from a chemical company in correcting bone deficits associated with DS. Our results suggest that EGCG improves the bone mineral density of trisomic femurs significantly better than the supplements while the EGCgNOW supplement from NOW FOODS improves trabecular and cortical bone structure. The results from HPLC analysis of supplements showed the presence of other catechins in EGCgNOW and degradation analysis revealed the rapid degradation of supplements. Therefore we hypothesize that the presence of EGCG degradation products and other green tea catechins in supplements may play a role in the differential skeletal effects we observed. We further hypothesized that a three week treatment of adolescent mice with EGCG will lead to an improvement in the learning and memory deficits that are observed in trisomic animals in comparison to control mice. However, our results indicate that three weeks of low-dose EGCG treatment during adolescence is insufficient to improve hippocampal dependent learning and memory deficits of Ts65Dn mice. The possibility remains that a higher dose of EGCG that begins at three weeks but lasts throughout the behavioral test period may result in improvement in learning and memory deficit of Ts65Dn mice.