Thesis (M.Sc. (Chemistry)) -- University of Limpopo, 2016 / In this study, antioxidant activity methodologies were evaluated in terms of analytical performances. The total antioxidant activity from Athrixia phylicoides leaves (Bush tea) determined using 2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH•) method, cupric ion reducing power (CUPRAC) method and cyclic voltammetry (CV). Folin-Ciocalteu method was used to quantify total phenolic content (TPC) in Athrixia phylicoides leaves. The influence of chemical and physical parameters on the total phenolic content and antioxidant activity determination were investigated. Results from direct sample and crude sample were compared. Antioxidant activity and phenolic content from Athrixia phylicoides leaves were compared with those from commercialised green tea, black tea and rooibos tea using two chosen antioxidant capacity method with acceptable characteristics. Results from the evaluation of the methods demonstrated excellent recoveries (99 to 103%) consistently, good linearity within the calibration concentration range (R2 = 0.997) and repeatable low coefficient of variation < 5% were indicative of good precision except for CV method. The average total antioxidant activity of various extracts of Athrixia phylicoides leaves ranged from 0.039 to 0.122 mg/mL (EC50), 0.031 to 0.233 mg/mL (EC50) and 339 to 429 mV (anodic potential) for DPPH method, CUPRAC method and CV method, respectively. The total antioxidant activity values for each Athrixia phylicoides samples determined by CUPRAC method were higher than the values produced by DPPH and CV methods. The highest antioxidant activities in the DPPH and CUPRAC methods were found in water extracts (direct sample). However, concentrated samples for DPPH method and CV gave a different trend with the methanol extract (crude sample) displaying the highest antioxidant capacity. Increasing the infusion time only increased total antioxidant activity determined by CUPRAC method, whilst DPPH and CV methods had the highest antioxidant activity in the lowest infusion time (3 min). Even though the results are inconclusive with regard to the effect of solid to solvent ratio effect on the total antioxidant activity, 1:150 ratio and 1:100 ratio extracts for both CUPRAC and DPPH methods and for CV gave the highest antioxidant capacities, respectively. The total antioxidant activities in pure antioxidant standards and in the teas were ranked in the following order by both CUPRAC and DPPH methods: Quercetin > catechin > Trolox and Chinese green tea > Joko black tea > Athrixia phylicoides leaves > Laager rooibos tea, respectively. Comparative study showed the necessity of employing more than one method, as each method for the same sample yielded different results. CUPRAC and DPPH methods displayed higher sensitivity and repeatability as compared to the CV method with poor precision.

Thesis (MSc. (Chemistry)) -- University of Limpopo, 2016 / In this study, antioxidant activity methodologies were evaluated in terms of analytical performances. The total antioxidant activity from Athrixia phylicoides leaves (Bush tea) determined using 2,2-diphenyl-1-picrylhydrazyl radical scavenging (DPPH•) method, cupric ion reducing power (CUPRAC) method and cyclic voltammetry (CV). Folin-Ciocalteu method was used to quantify total phenolic content (TPC) in Athrixia phylicoides leaves. The influence of chemical and physical parameters on the total phenolic content and antioxidant activity determination were investigated. Results from direct sample and crude sample were compared. Antioxidant activity and phenolic content from Athrixia phylicoides leaves were compared with those from commercialised green tea, black tea and rooibos tea using two chosen antioxidant capacity method with acceptable characteristics. Results from the evaluation of the methods demonstrated excellent recoveries (99 to 103%) consistently, good linearity within the calibration concentration range (R2 = 0.997) and repeatable low coefficient of variation < 5% were indicative of good precision except for CV method. The average total antioxidant activity of various extracts of Athrixia phylicoides leaves ranged from 0.039 to 0.122 mg/mL (EC50), 0.031 to 0.233 mg/mL (EC50) and 339 to 429 mV (anodic potential) for DPPH method, CUPRAC method and CV method, respectively. The total antioxidant activity values for each Athrixia phylicoides samples determined by CUPRAC method were higher than the values produced by DPPH and CV methods. The highest antioxidant activities in the DPPH and CUPRAC methods were found in water extracts (direct sample). However, concentrated samples for DPPH method and CV gave a different trend with the methanol extract (crude sample) displaying the highest antioxidant capacity. Increasing the infusion time only increased total antioxidant activity determined by CUPRAC method, whilst DPPH and CV methods had the highest antioxidant activity in the lowest infusion time (3 min). Even though the results are inconclusive with regard to the effect of solid to solvent ratio effect on the total antioxidant activity, 1:150 ratio and 1:100 ratio extracts for both CUPRAC and DPPH methods and for CV gave the highest antioxidant capacities, respectively. The total antioxidant activities in pure antioxidant standards and in the teas were ranked in the following order by both CUPRAC and DPPH methods: Quercetin > catechin > Trolox and Chinese green tea > Joko black tea > Athrixia phylicoides leaves > Laager rooibos tea, respectively. Comparative study showed the necessity of employing more than one method, as each method for the same sample yielded different results. CUPRAC and DPPH methods displayed higher sensitivity and repeatability as compared to the CV method with poor precision.

Thesis (M.Sc.) --University of Limpopo, Turfloop Campus, 2010 / A study was conducted to determine the effect of fermentation temperature and duration on chemical composition of bush tea (Athrixia phylicoides DC.). Bush tea was fermented in incubators at different temperatures and for different lengths of time for quality improvement. Treatments for fermentation temperature consisted of control (24°C; room temperature), 30°C, 34°C, 38°C and 42°C where the tea leaves were fermented for 30 minutes. Treatments for fermentation time consisted of control (0), 60, 90, and 120 minutes at an incubator temperature of 22-26°C. A completely randomized design (CRD) was used with three replicates for both evaluations. The chemical analysis (polyphenols, tannins and antioxidants) were done using Waterman and Mole (1994) method. The results of this study demonstrated that fermentation temperature significantly increases polyphenols at 30, 34, and 38°C whereas tannin content showed a great reduction at 38 and 42°C. Increasing fermentation time achieved a significant increase in both polyphenols (60 and 90 minutes) and tannin contents (90 and 120 minutes). However, changes in either fermentation temperature or time did not give any significant influence on antioxidant content of bush tea. / National Research Foundation

Thesis (M.Sc)--University of Limpopo, 2010. / Black tea (Camellia sinensis) and black tea combined with bush tea (Athrixia phylicoides) were analyzed for their polyphenol content, antioxidant activity and tannin content. Solvent extraction methods were used for extracting polyphenols, antioxidant activity and tannin content. Total phenols were determined using the Folin-Ciocalteu method, antioxidant activity using Trolox Equivalent Antioxidant Capacity (TEAC) assay and tannin content using vanillin-HCl method. Polyphenol content, antioxidant activity and tannin content were calculated using regression equation Y=MX+C, where C=intercept, Y= Absorbance of the sample and M= Gradient. Results of black tea in this study showed that total polyphenols, antioxidant activity and tannin content decreased with decrease in temperature and time and this was attributed to the release of polysaccharides at 90°C. At 90°C for 3 minutes 7.68mg/100g of total polyphenol, 3.85μmol/g of antioxidants and 2.81mg/100g of tannin were obtained and this amount decreased to 5.50mg/100mg for total polyphenols, 1.31μmol/g for antioxidant activity and 0.72mg/100mg for tannin content at 30°C for 10 minutes. Combining the two tea types (50:50) resulted in a significant (p<0.005) decrease in total polyphenols, antioxidant and tannin content. Results of this study showed that at high temperature of 90°C for 3 minutes 2.64mg/100g for total polyphenol, 2.48μmol/g for antioxidants and 1.8mg/100g for tannin were obtained and these amounts decreased to 1.39mg/100mg for total polyphenols, 0.35μmol/g for antioxidant activity and 0.64mg/100mg for tannin content at 30°C for 10 minutes. Thus high brewing temperature causes increase in total polyphenols, antioxidant activity and tannin content of the combined teas. This study showed that black tea and combined black tea and bush tea solutions had large TEAC radical scavenging activity which can be related to total polyphenols and catechins. The antioxidant activity of the tea solution increased with increase in extraction temperature and duration of soaking. It is recommended that 3-min soaking temperature of 90 °C of black tea and combined bush tea and black tea can be used due to the highest total polyphenols and antioxidant activity obtained.

Bush tea (Athrixia phylicoides) is an herbaceous plant of the Asteraceae family used throughout history as medicinal herbal tea by the people of South Africa. Many studies stated that the plant has an ability to be commercialized as a medicinal herbal tea. But vegetative propagation of this type of plant by stem cutting, survival ability of the rooted cuttings, response to different hormone concentrations and the requirement of the seeds for germination has not been studied. In this investigation, different features aimed at effective propagation of bush tea were studied. These comprised: cutting position (apical vs. basal), media (pine bark vs. sand), hormone (Seradix No. 2), season (summer, autumn, winter and spring), transplanting survival of rooted apical and basal cuttings, response of basal cuttings to three hormone concentration levels (Seradix No. 1, 2 and 3) and light and temperature requirement for bush tea seed germination. In vegetative propagation, apical cuttings rooted to higher percentage and produced high root number as well as longer roots than basal cuttings. Pine bark improved the number of roots developed but had no effect on rooting percentage as well as root length. Application of rooting hormone (Seradix No. 2) increased root numbers but not rooting percentage or root length. Rooting of cuttings was improved when propagated in autumn (longer roots) and spring (more number of roots) than in summer or winter. There was higher survival percentage (67.5%), high root number as well as longer roots from apical cuttings than from basal cuttings (50%) two months after transplanting. Propagation in pine bark with hormone application increased root number after transplanting. Application of hormone also improved root and shoot length after transplanting. Apical cuttings propagated in pine bark with hormone developed more number of roots. Cuttings propagated in sand with hormone and in pine bark without hormone also produced longer shoots after transplanting. Regarding response of basal cuttings to hormone concentration, high number of roots was produced in pine bark with Seradix No. 2 at 10 days after planting (DAP) but at 15 DAP more roots were produced in pine bark with Seradix No. 1. With sand, more roots were produced with Seradix No. 3 than Seradix No. 1 and 2. Number of roots were also higher with 0.3% IBA concentration (Seradix No. 2) and 0.1% IBA concentration (Seradix No. 1). Similarly, cuttings with lower IBA concentration (0.1%, Seradix No. 1) rooted to higher percentage followed with 0.3% IBA concentration (Seradix No. 2). Germination percentage of bush tea seeds differed with the temperature treatments and the highest was 75.5% at 20 and 25°C followed by 15°C with 64.5% and low percentage at 30 and 10°C with 36 and 47% respectively. There was a high germination percentage in constant temperatures than alternate temperatures and in continuous light than alternate light: dark or continuous dark. Germination percentage was also higher in continuous light at constant temperatures than with alternated light: dark with constant temperatures. In addition, there was more differences in germination percentage with variation in light exposure than variation in temperatures. At low temperature (10°C), longer time was required to start germination and germination rate was high at 20°C continuous light and low at 30:30°C alternate light: dark. Based on this investigation, better vegetative propagation and survival of bush tea can be attained from apical cuttings with Seradix No. 2 but basal cuttings rooted better with Seradix No. 1 in pine bark. Seeds germinated to higher percentage and rate at 20°C constant temperature and continuous light. / Dissertation (MSc (Agric) Horticulture)--University of Pretoria, 2007. / Plant Production and Soil Science / unrestricted

Herbal medicines are an important part of the African culture and tradition and about 80% of Africa’s population relies on traditional remedies for their primary health care needs (WHO, 2008). Athrixia phylicoides is widely used as a traditional remedy, but despite its substantial use, literature on its chemical composition and biological activities is limited. In this study, the chemical composition of the essential oil was determined using gas chromatography combined with mass spectroscopy (GC/MS) and analysis resulted in the identification of 182 compounds. The major compounds identified in selected samples were α-pinene, β-pinene, caryophyllene oxide, β-caryophyllene, myrcene and spathulenol. Based on the traditional uses of A. phylicoides, various in vitro biological activities were investigated. The extensive use of A. phylicoides for the treatment of boils, sores, bad acne, infected wounds and cuts, prompted the antimicrobial (bacterial and fungal) study using the minimum inhibitory concentration (MIC) assay. The methanol extract and essential oil were more selective for the Grampositive bacteria than the Gram-negative bacteria. The extract exhibited stronger activity against all micro-organisms tested compared to the essential oil, with the highest activity against S. aureus, B. cereus and B. subtilis (MIC = 1 mg/ml). The bioactive compound, (4-hydroxyphenyl) propyl coumaroate was isolated from the extract and proved to be most active against S. aureus (MIC = 19.5 μg/ml). Using the DPPH• assay, a comparative antioxidant study was performed. The anti-oxidant activity of the aqueous extract of A. phylicoides (IC50 = 14.01 ± 2.68 μg/ml) was greater than rooibos and Ceylon (black) tea (IC50 > 25.00 μg/ml); comparable to green rooibos and honeybush tea (IC50 = 18.01 ± 4.06 μg/ml; 18.02 ± 4.27 μg/ml, respectively), but less active than green tea (IC50 = 9.64 ± 0.96 μg/ml). The antimalarial activity was determined using the [3H] hypoxanthine incorporation method. The essential oil exhibited greater antimalarial activity against the chloroquine-resistant Plasmodium falciparum (FCR-3) strain (IC50 = 1.006 ± 0.06 μg/ml) than the methanol extract. The 5-lipoxygenase assay was used to assess the anti-inflammatory activity of the methanol extract and essential oil. Only the essential oil displayed anti-inflammatory activity (IC50 = 25.68 μg/ml). Low safety indices were reported for both the methanol extract (SI = 2.28) and essential oil (SI = 1.77). The in vitro biological activities may validate the use of A. phylicoides in traditional medicine.

Thesis (M.Sc. (Horticulture)) --University of Limpopo, 2012 / Refer to document / the National Research Foundation

Thesis (M.Sc. (Horticulture)) --University of Limpopo, 2007 / Bush tea (Athrixia phylicoides DC) has been used for many years by the people of South Africa for medicinal purposes. The herbs was only harvested from the wild, therefore data on the effects of mineral nutrition on total antioxidant activity had not been established. The objective of this study was to determine the seasonal effect of nitrogen (N), phosphorus (P) and potassium (K) nutrition on total antioxidant activity (TAA) in cultivated bush tea leaves. Treatments consisted of 0, 100, 200, 300, 400 or 500 kgha-1 N, P or K in a randomized complete block design with four replications under 50 % shade nets. Three (N, P and K) parallel trials were conducted per season (autumn, winter, spring and summer). TAA was determined using 2,2-diphenyl-1-picrylhydrazyl-hydrate (DPPH) method and analyzed in a spectrophotometer. Results of this study demonstrated that regardless of season, the application of N, P and K fertilizers increased quadratically levels of TAA in bush tea with most of the increase occurring between 0 and 300 N, 300 P and 200 K kgha-1. Therefore, for improved total antioxidant content in bush tea leaves, 300 N, 300 P and 200 K kgha-1 N are recommended. / National Research Foundation (NRF), Agricultural Research Council(ARC-LNR)

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