Cyclopia Intermedia Research Articles

Evaluation of chemical constituents of Rooibos (Aspalathus linearis) and Honeybush (Cyclopia intermedia) as adenosine A1/A2A receptor ligands

Rooibos (Aspalathus linearis) and Honeybush (Cyclopia intermedia) are popular tisanes in South Africa and are of growing interest due to the wide variety of flavonoids and other phytochemicals they contain. Despite their history as herbal teas and traditional medicines, the chemical constituents of these tisanes have yet to be studied for their effects on adenosine receptors. A series of 30 commercially available chemical constituents of Rooibos and Honeybush were investigated via radioligand binding studies to determine their adenosine A1 and A2A receptor affinity at both rat and human subtypes in order to establish structure-activity relationships and identify novel adenosine receptor ligands. In addition, in silico evaluations of the 30 test compounds were also performed to predict their physiochemical and pharmacokinetic properties. The most promising chemical constituent was kaempferol (28) which showed sub-micromolar affinity towards the rat A1 subtype (rA1Ki = 0.7287 μM; hA1Ki = 9.88 µM) and acted as an antagonist toward adenosine rA1 receptors. Additionally, quercetin (2), chrysoeriol (8), luteolin (9), eriodyctiol (12), and naringenin (27) also showed adenosine A1 and/or A2A receptor affinity. It was observed that a flavonol scaffold is preferred to flavone and flavanone scaffolds, and within the flavonols, C4’-OH substitution on ring B is preferred to C3’,4’-diOH substitution. These phytochemicals, specifically kaempferol (28), may be considered lead-like and valuable in designing novel ligands, based on in vitro and in silico evaluation.

The effect of Aspalathin linearis, Cyclopia intermedia and Sutherlandia frutescene on sperm functional parameters of healthy male wistar rats.

Introduction: Rooibos (Aspalathin linearis), honeybush (Cyclopia intermedia), and sutherlandia (Sutherlandia frutescene) are three Southern Africa indigenous plants, of which the extracts have become house-hold items and are consumed on a large scale. Although, they are known for their antioxidant properties, studies have highlighted danger in the excessive intake. Therefore, the current study investigated whether treatment with rooibos, honeybush, and sutherlandia will impact sperm functional parameters positively or otherwise, in healthy rats. Methods: Fourteen-week-old pathogen-free adult male Wistar rats (250-300g) were randomly divided into four groups of ten, including a control, rooibos (RF), honeybush (HB) and a sutherlandia (SL) group. After 7weeks of treatment, animals were sacrificed. Spermatozoa were retrieved from the cauda epididymis for motility, morphology and concentration analysis and the testis was used for all biochemical assays. Results: The infusion treated animals (RF, HB, and SL) presented with a non-significant decrease of -14.3%, -18.2%, -17.2% and -24.8%, -20.7%, -27.3% in total motility and progressive motility when compared to the control group, respectively. There was a significant increase in number of spermatozoa with slow speed (p = 0.03), especially in SL treated group compared to the control (p = 0.03). Additionally, there was an increase of 28.8%, 31.7%, 23% in superoxide dismutase (SOD) activity of RF, HB and SL compared to control, respectively. This was accompanied with a percentage decrease of -21.1%, -23.7%, 45.9% in malondialdehyde (MDA) levels compared to the control group. Conclusion: In summary, animals treated with the respective infusions presented with a percentage increase in SOD activity but have reduced sperm motility and decreased normal morphology. Paradoxically, they presented with increased sperm concentration. Hence, it is presumed that rooibos, honeybush and sutherlandia may enhance sperm quantity (concentration) but may impair sperm quality (motility morphology) when consumed by healthy animals.

Anti-Inflammatory, Anti-Diabetic, Anti-Oxidant and Cytotoxicity Assays of South African Herbal Teas and Bush Tea Blends.

South Africa is home to a variety of herbal teas, such as bush tea (Athrixia phylicoides DC.), honeybush tea (Cyclopia intermedia E. Mey and C. subternata Vogel), special tea (Monsonia burkeana Planch. ex Harv.), and rooibos tea (Aspalathus linearis (Burm.f.) R. Dahlgren) that are known to possess anti-oxidant, anti-inflammatory and anti-diabetic properties. The objective of this study was to determine the in vitro anti-oxidant activity of selected tea blends using 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) assays, as well as to assess their anti-inflammatory properties using the 15-lipoxygenase inhibitory assay. Furthermore, the study measured glucose utilisation in C2C12 myotubes. Lastly, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was used to test the safety of the tea extracts on Vero cells (African green monkey kidney cell line). Special tea and its blend with bush tea exhibited potent anti-oxidant and anti-inflammatory activity. The blending of bush tea with special tea at different ratios resulted in increased anti-oxidant activity. Although special tea had a level of cell toxicity, its toxicity was lowered during blending. All of the tea samples showed anti-diabetic effects, although with less potency as compared to insulin. The current investigation supports the use of blended herbal teas, and the positive anti-inflammatory effect of special tea warrants further research.

Development of HPLC method for quantification of phenolic compounds in Cyclopia intermedia (honeybush) herbal tea infusions

Cyclopia intermedia is the major source material for the production of honeybush tea, specifically the ‘fermented’ (oxidised) product. The phenolic composition of ‘cup-of-tea’ infusions from other Cyclopia species has been reported, but such information is lacking for C. intermedia. The HPLC method previously developed for extracts of green, unoxidised C. intermedia is not suitable due to the co-elution of unidentified compounds with mangiferin and isomangiferin. A method was developed and validated to quantify the major phenolic compounds in infusions of fermented C. intermedia at ‘cup-of-tea’ strength. Eighteen compounds were identified or tentatively identified and 14 compounds could be quantified with suitable intra- and inter-day repeatability (relative standard deviation < 8%). In a large sample set (n = 96) of fermented C. intermedia, hesperidin, 3-β-D-glucopyranosyl-4-O-β-D-glucopyranosyliriflophenone and mangiferin were present at the highest concentrations (mean > 5 mg/L) in the infusions, followed by isomangiferin, vicenin-2, 3-β-D-glucopyranosyliriflophenone, neoponcirin, hesperetin and protocatechuic acid (mean > 1 mg/L). Agglomerative hierarchical clustering analysis and principal component analysis showed the separation of samples into two groups mainly based on the presence (n = 32) or absence (n = 64) of neoponcirin. Based on information about the origin of the samples, the presence of chemotypes in different geographical areas is postulated.

Descriptive histomorphological evaluation of the testis and caudal epididymis following treatment with rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia) and sutherlandia (Lessertia frutescens) in healthy and streptozotocin-induced diabetic rats

Objective: To evaluate the testis and the cauda epididymis after treating both healthy and diabetic rats with rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia), and sutherlandia (Lessertia frutescens), respectively. Methods: Ninety male Wistar rats were randomly divided into nine groups (n=10 per group). Fifty rats were randomized to receive tap water, 0.1 M citrate buffer, 2.0% fermented rooibos, 4.0% fermented honeybush, or 0.2% unfermented sutherlandia. Diabetes was induced via a single intraperitoneal injection of streptozotocin (STZ) at 45 mg/kg in 40 rats. The diabetic rats were randomized to receive tap water, 2.0% fermented rooibos, 4.0% fermented honeybush, or 0.2% unfermented sutherlandia. Rats were sacrificed after seven weeks of diabetic induction. Blood was collected for hormonal assay, while the testes and caudal epididymis were retrieved for histomorphological evaluations. Results: Diabetic rats treated with 2.0% fermented rooibos exhibited a significant increase in fasting plasma glucose (FPG) versus the vehicle group (P=0.002), and a 3.6% decrease compared to the diabetic control group (P&gt;0.05). Diabetic rats treated with honeybush or sutherlandia showed 23.6% and 15.4% decrease in FPG when compared to the diabetic control group, respectively (both P&gt;0.05). The diabetic control rats had a significant decrease in the percentage of histologically normal seminiferous tubules compared to the control group (P=0.049). There was 21.0%, 14.0%, and 5.4% rise in the percentage of normal seminiferous tubules in diabetic rats receiving rooibos, honeybush or sutherlandia, respectively, compared to the diabetic control group. The infusion control groups (rooibos, honeybush and sutherlandia) showed normal seminiferous tubules, presence of spermatozoa in the epididymal lumen, and had normal overall architecture. Both testicular and epididymal morphology were altered in the diabetic control group, but these disruptions were mildly ameliorated by rooibos, honeybush, and sutherlandia. Conclusions: The detrimental effects of diabetes on the histomorphological architecture of the testis and epididymis are mildly ameliorated by the infusions (2.0% fermented rooibos, 4.0% fermented honeybush, and 0.2% unfermented sutherlandia).

Testicular oxidative stress and apoptosis status in streptozotocin-induced diabetic rats after treatment with rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia), and sutherlandia (Lessertia frutescens) infusions

Objective: To investigate the testicular oxidative stress and apoptosis status, as well as the sperm functional parameters in streptozotocin (STZ) induced diabetic rats following treatment with rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia) and sutherlandia (Lessertia frutescens) infusions. Methods: Diabetes was induced by injecting fourteen-week-old adult male Wistar rats (250-300 g) with a single intraperitoneal injection of STZ (45 mg/kg body weight). Fifty rats were randomly divided into five groups: the vehicle group received 0.1 M citrate buffer, the diabetic control group received 45 mg/kg STZ, the diabetic+rooibos group received 45 mg/kg STZ + 2.0% rooibos, the diabetic+honeybush group received 45 mg/kg STZ + 4.0% honeybush, and the diabetic+sutherlandia group received 45 mg/kg STZ + 0.2% sutherlandia. Rats were sacrificed 7 weeks after induction of diabetes mellitus. The testes and epididymides were harvested and weighed after induction. Spermatozoa were retrieved from the cauda epididymis for motility, concentration, and morphology analysis, and the testis was used for all biochemical assays. Oxidative stress was determined by measuring malondialdehyde levels, catalase, and superoxide dismutase activities, while apoptotic biomarkers were evaluated by Western blotting assays. Results: After induction of diabetes, rats in the diabetic control group, diabetic+rooibos group, diabetic+honeybush group, and diabetic+sutherlandia group presented with significantly elevated blood glucose levels as compared with the vehicle group (P&lt;0.001). Rats in the diabetic control group had a reduction in sperm progressive motility, while rats in the diabetic+rooibos group and the diabetic+sutherlandia group displayed an increase in progressive motility as compared with the diabetic control group. The diabetic control animals showed a 40.0% decrease in sperm concentration when compared to the vehicle group, and there were no significant differences in sperm kinematic and speed parameters between the groups. In addition, the percentage of morphologically normal spermatozoa was increased by 13.0%, 16.0%, and 15.0% after treatment with rooibos, honeybush, and sutherlandia, respectively and the rats in the diabetic+infusion groups also displayed an increase in superoxide dismutase activity when compared to the diabetic control group. Conclusions: Rooibos, honeybush and sutherlandia infusions may partly alleviate diabetes-induced sperm function impairment by reducing oxidative stress.

Antimicrobial activity of honeybush (Cyclopia intermedia) ethanol extract against foodborne pathogens and its application in washing fresh-cut Swiss chard

This study aimed to determine the antimicrobial activities of honeybush ethanol extract (HBE) against Listeria monocytogenes and Escherichia coli O157:H7, and to define whether it could be used to wash fresh-cut Swiss chard. HPLC analysis showed that HBE contained the phenolic compounds, hesperidin, luteolin, kaempferol, mangiferin, and epicatechin, indicating potential antimicrobial activities. The results of disc diffusion tests, time-kill assays, and analyses of the released intracellular compounds revealed that HBE was more effective against L. monocytogenes than E. coli O157:H7. Moreover, scanning electron microscopy images showed that HBE damaged and shriveled the cells of both pathogens. Washing fresh-cut Swiss chard spiked with L. monocytogenes and E. coli O157:H7 with HBE (6 mg/mL) resulted in 2.31–2.67 log reductions in the populations of these bacteria compared with those in the controls. These results were similar to those of washing with sodium hypochlorite (2.20–2.58 log reduction). In addition, washing with HBE maintained the marketable qualities of fresh-cut Swiss chard, namely, color, texture, and electrolyte leakage rates that indicated tissue deterioration. In contrast, the electrolyte leakage rate of the samples washed with sodium hypochlorite increased. These results indicate that HBE could replace sodium hypochlorite and serve as a novel washing agent to ensure the safety and quality of fresh-cut leafy vegetables.

Pharmacological Activity of Honeybush (Cyclopia intermedia) in Boar Spermatozoa during Semen Storage and under Oxidative Stress.

Simple SummaryNowadays, pig breeding is mostly conducted by artificial insemination using diluted semen stored for 1 to 5 days. During semen handling and storage, sperm quality usually declines, mainly because of oxidative stress and bacterial contamination. As cheap and natural sources of antioxidants, medicinal plants have become an alternative to the most common additives used in semen extenders. In this regard, several indigenous plants from Southern Africa have shown pharmacological activity in different animal cell types, although their effects on sperm cells have not been explored extensively. In the present study, we tested the effects of honeybush (Cyclopia intermedia) aqueous extract as a preservative of boar semen during 5 days of storage and under induced oxidative stress. Overall, this plant extract enhanced several sperm quality parameters and did not show any toxic effects. Supplementation with honeybush extract was able to improve the preservative properties of a long-term semen extender, thus confirming the beneficial use of plant extracts as natural additives for boar sperm.In recent decades, an increasing number of ethnopharmacological studies have been dedicated to medicinal plants from South African fynbos. Among these plants, honeybush (Cyclopia spp.) has become a popular tea, mainly due to its healthy properties and caffeine-free status. The antioxidant, antimutagenic, and antimicrobial properties of this plant have been reported in several cell types, but its effects on reproductive function are still unknown. Here, we assessed the effects of honeybush (Cyclopia intermedia) on boar sperm parameters under induced oxidative stress (Fe2+/ascorbate) and during five days of semen storage at 17 °C without oxidative stress. In both experiments, four concentrations (200, 50, 12.5, and 3.125 µg/mL) of fermented honeybush were tested. Our results show that honeybush enhances sperm parameters, and no toxic effects were observed at any of the tested extract concentrations. Interestingly, honeybush (12.5 µg/mL) improved the sperm motility and kinetic parameters, preserved the plasma membrane integrity, and reduced the lipid peroxidation in the samples exposed to Fe2+/ascorbate (p < 0.05). In the stored samples, positive effects of honeybush on sperm parameters (motility, kinetics, acrosome, and mitochondria) were observed from 48 h until 120 h of semen storage (p < 0.05). Our results clearly show the protective effects of honeybush on sperm samples, thus promoting its use as a natural source of antioxidants for boar semen.

Hyperspectral Imaging as a Rapid Quality Control Method for Herbal Tea Blends

In South Africa, indigenous herbal teas are enjoyed due to their distinct taste and aroma. The acclaimed health benefits of herbal teas include the management of chronic diseases such as hypertension and diabetes. Quality control of herbal teas has become important due to the availability of different brands of varying quality and the production of tea blends. The potential of hyperspectral imaging as a rapid quality control method for herbal tea blends from rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia), buchu (Agathosma Betulina) and cancerbush (Sutherlandia frutescens) was investigated. Hyperspectral images of raw materials and intact tea bags were acquired using a sisuChema shortwave infrared (SWIR) hyperspectral pushbroom imaging system (920–2514 nm). Principal component analysis (PCA) plots showed clear discrimination between raw materials. Partial least squares discriminant analysis (PLS-DA) models correctly predicted the raw material constituents of each blend and accurately determined the relative proportions. The results were corroborated independently using ultra-high performance liquid chromatography coupled to mass spectrometry (UHPLC-MS). This study demonstrated the application of hyperspectral imaging coupled with chemometric modelling as a reliable, rapid and non-destructive quality control method for authenticating herbal tea blends and to determine relative proportions in a tea bag.

Antimicrobial and antioxidant activities of different solvent extracts from fermented and green honeybush (Cyclopia intermedia) plant material

Ethnopharmacological relevanceCyclopia intermedia is indigenous to South Africa and used to prepare honeybush herbal tea. This aromatic herbal tea has been associated with numerous health benefits, mostly based on anecdotal evidence and with very few studies reporting on the antimicrobial activities. AimThe inhibitory effect on the growth of important nosocomial microorganisms and possible association with the antioxidant capacity/content of various solvent extracts of green/unfermented and fermented honeybush plant material were determined in the current study. Materials and methodsThe agar disk diffusion assay was used as a screening assay for the antimicrobial activity of the various honeybush extracts, while the minimal inhibitory concentration (MIC) values, using the broth micro-dilution method, were determined against Streptococcus pyogenes, Staphylococcus aureus and Candida albicans. Active antimicrobial compounds were then shown using thin layer chromatography bioautography. Total antioxidant capacities and -content were also determined for each extract, while the main phenolic compounds were quantified using HPLC. ResultsSix of the eight solvent extracts of honeybush showed antimicrobial activity, with the fermented and green methanol extracts being most effective against S. aureus and C. albicans respectively, whilst the green chloroform extract was most potent against S. pyogenes. Thin layer chromatography-bioautography acknowledged the existence of active antimicrobial fractions within these different solvent honeybush extracts. Regardless of the assay, the green honeybush extracts generally exhibited the highest antioxidant capacity when compared to the fermented honeybush. The total polyphenols were also observed to be highest in green extracts when using water and methanol as solvents. In general the mangiferin and hesperidin contents were higher in the green than the fermented extracts of honeybush plant material. ConclusionsThe inhibitory activity of the various extracts against specific microorganisms was observed to be linearly proportional to the extract concentration. Although the different solvent extracts can only be considered weak antimicrobial agents, three compounds showed specific activity and should be further elucidated in future. It appears the antioxidant capacity of the various solvent herbal extracts did not relate to the antimicrobial activities; however, further work will be required to ascertain this observation. The current data also suggested that the various extracts of honeybush can be considered a good source of a unique blend of natural phytochemical antioxidants and antimicrobial compounds and should be further elucidated.

Rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia) and cancer bush (Sutherlandia frutescens subsp. microphylla) protect against tobacco-specific mutagenesis in vitro

Antimutagenesis studies against the tobacco-specific mutagens 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N-oxide, 4 (methyl-nitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), were conducted using hot water aqueous extracts of rooibos (Aspalathus linearis), honeybush (Cyclopia intermedia), and cancer bush (Sutherlandia frutescens). Aqueous extracts of both “fermented” and “unfermented” (green) rooibos and honeybush were included, while extracts of green and black teas (Camellia sinensis) served as benchmarks. A polyphenol-enriched methanol extract of unfermented rooibos (RgM) was included to further elucidate the possible role of rooibos polyphenols. Studies were performed in the presence of the metabolic activation against Salmonella typhimurium tester strain TA1535, using the standard plate incorporation and micro-suspension, pre-incubation assays. The mutagenic effects of NNK against the strain TA1535 was best demonstrated using the standard plate incorporation assay, while a higher mutagenicity was demonstrated for NNAL using the micro-suspension, pre-incubation method. Black tea and RgM exhibited the highest protection against NNK-induced mutagenesis followed by the aqueous extracts of rooibos≥green tea≥honeybush≥cancer bush. Black tea, green tea, RgM and unfermented rooibos were the most effective against NNAL-induced mutagenesis, followed by fermented rooibos. The two honeybush extracts exhibited similar, but the weakest protective response. When considering the amount of total polyphenols (TPP) incorporated in the plate incorporation assay, cancer bush exhibited similar protection to that of fermented and unfermented honeybush against NNK mutagenesis. The involvement of specific polyphenol-cytochrome P450 (CYP450) interactions is likely to be involved in the protection against tobacco-related mutagenesis. Polyphenol constituents of rooibos, honeybush and cancer bush could play an important role in the protection against mutagenesis induced by the major tobacco-specific carcinogens.

Polyphenols from Cyclopia genistoides and their xanthine oxidase inhibitory activity

The genus Cyclopia (Fabaceae), comprising four species, Cyclopia intermedia, genistoides, subternata, and sessiliflora has significant commercial value as the source of the popular caffeine free herbal tea, honeybush. Several studies investigated the chemical composition of Cyclopia, most of which are directed to the analysis of their polyphenolic substances [1]. The bioactivity of different species was also recorded, such as antimutagenic and estrogen-like effects, but anti-gout activity (xanthine oxidase inhibition) has not yet been investigated.

Sensory profiling of honeybush tea (Cyclopia species) and the development of a honeybush sensory wheel

Several Cyclopia species (Cyclopia sessiliflora, Cyclopia longifolia, Cyclopia genistoides, Cyclopia intermedia, Cyclopia subternata and Cyclopia maculata), used as honeybush herbal tea, were analyzed using descriptive sensory analysis in order to develop a generic honeybush sensory wheel. It was found that the “characteristic” sensory profile of honeybush could be described as “floral”, “sweet-associated”, “fruity”, “plant-like” and “woody” with a sweet taste and a slightly astringent mouthfeel, whereas other attributes defined differences in the sensory characteristics between the Cyclopia species. The species could be divided into three distinct groups: group A (C. sessiliflora, C. intermedia and C. genistoides) associated with “fynbos-floral”, “fynbos-sweet” and “plant-like” attributes, group B (C. longifolia and C. subternata) with “rose geranium” and “fruity-sweet” and group C (C. maculata) with “woody”, “boiled syrup” and “cassia/cinnamon”. The large sample set enabled the development of a generic honeybush sensory wheel, comprising of 30 attributes, organized into positive and negative attributes in primary and secondary tiers. Gas chromatographic-olfactometry of the aroma fraction of a sample of C. maculata with a prominent spicy aroma indicated a high level of eugenol, the only aroma-active compound that associated with a spicy aroma.

Leaf and stem anatomy of honeybush tea (Cyclopia species, Fabaceae)

The leaf and stem microstructure of commercialised Cyclopia species (Cyclopia intermedia, Cyclopia subternata, Cyclopia maculata, Cyclopia genistoides) is described and illustrated for the first time. Leaflet characters of diagnostic value include the shape, margin (flat or revolute), bundle sheath extensions, crystals (aggregates, solitary crystals and crystal sand), relative size of the upper and lower epidermal cells and thickness of the cuticle. The study revealed an abundance of phenolic compounds, not only in the leaf epidermal cells and leaf bundle sheaths, but also in the secondary phloem, xylem rays, xylem parenchyma and pith, showing that the inclusion of older stems may be acceptable and does not necessarily lower the quality of the product.

Modulating Effects of Rooibos and Honeybush Herbal Teas on the Development of Esophageal Papillomas in Rats

Widespread consumption of herbal teas has stimulated interest in their role as cancer preventive agents. The present investigation monitored the modulation of methylbenzylnitrosamine (MBN)-induced esophageal squamous cell carcinogenesis by rooibos (Aspalathus linearis) and honeybush (Cyclopia intermedia) herbal and Camellia sinensis teas in male F344 rats. The tumor multiplicity was significantly (P < 0.05) inhibited by unfermented honeybush (45.5%), green (50%), and black (36%) teas, while the other teas exhibited weaker effects (<30% inhibition). The mean total papilloma size was reduced by unfermented rooibos (87%), unfermented honeybush (94%), and fermented honeybush (74%) due to the absence of large papillomas (>10 mm3). Reduction of the mean total papilloma number correlated with the total polyphenol (TPP) (r = 0.79; P < 0.02) and flavanol/proanthocyanidin (FLAVA) (r = 0.89; P < 0.008) intake (mg/100 g body weight) of the teas and the FLAVA (r = 0.89; P < 0.04) and flavonol/flavones/xanthones (r = 0.99; P < 0.002) intake when considering only the herbal teas. A daily TPP intake threshold of 7 mg/100 g body weight existed below where no inhibition of papilloma development was observed. Fermentation of herbal teas reduced the inhibitory effects on papilloma development associated with a reduction in the polyphenolic constituents. The inhibitory effect of herbal teas on papilloma development is associated with different flavonoid subgroups and/or combination thereof.

Development of HPLC method for Cyclopia subternata phenolic compound analysis and application to other Cyclopia spp.

Analysis of phenolic compounds in Cyclopia spp. (honeybush) is very important because of commercial interest in the plant material as an herbal tea and in extracts for the nutraceutical and cosmeceutical markets. An existing HPLC-DAD method was adapted and proved reproducible as evidenced by good intra- and inter-day precision. The method allows the quantification of several well-known as well as unidentified phenolic compounds. Its suitability for quantification of the major phenolic compounds present in the water extracts of unfermented and fermented Cyclopia subternata, Cyclopia intermedia, Cyclopia genistoides and Cyclopia sessiliflora was evaluated. Known compounds that were identified and quantified include mangiferin (0.1–11.8 g/100 g), isomangiferin (0.1–3.2 g/100 g), eriocitrin (not detected–0.4 g/100 g), hesperidin (0.2–1.2 g/100 g) and scolymoside (not detected–1.8 g/100 g), while narirutin and luteolin were detected in most samples, but could not be quantified due to low concentrations and co-elution. An unidentified compound (traces–4.8 g hesperidin equivalents/100 g), unidentified hydroxycinnamic acid derivative (not detected–0.3 g mangiferin equivalents/100 g), unidentified flavanone (not detected–2.0 g hesperidin equivalents/100 g) and an eriodictyol-glycoside (not detected–0.5 g hesperidin equivalents/100 g) could also be quantified. Large variation was observed within a sample series of the same extract type due to the nature of the plant material. Qualitative and quantitative differences were observed between species, while “fermentation”, a high-temperature process involving non-enzymatic oxidation, decreased the content of most phenolic compounds. Results using DAD as the only detection method should be handled with care, as the tentative identification of compounds 9 and 10, based on retention time and UV–Vis spectrum, could not be confirmed using liquid chromatography–electrospray ionisation-mass spectrometry.

Characterization of Flavor Modulating Effects in Complex Mixtures via High Temperature Liquid Chromatography

The identification of flavor modulating compounds, for example, bitter masking or sweet enhancing compounds, in complex mixtures such as botanical extracts or food preparations is difficult and time- and work-intensive. To accelerate this process, an improved screening method was developed on the basis of the separation of complex matrixes by the so-called LC Taste setup and subsequent comparative sensory analysis. The eluent containing only water and ethanol was diluted with a basic tastant solution (500 mg L(-1) caffeine and 5% sucrose, respectively) and evaluated by a trained panel by duo comparison tests. This novel method was applied to the known flavor and taste modulating substances homoeriodictyol (1), sterubin (2), hesperetin (3), and lactisol (9) as well as to simple mixtures of homoeriodictyol (1), sterubin (2), and hesperetin (3). To evaluate the potential of the method for more complex matrixes, the protocol was applied to plant extracts from Yerba Santa (Eriodictyon californicum) and honeybush tea (Cyclopia intermedia). The flavor modulating activities reported for homoeriodictyol (1), sterubin (2), and hesperetin (3) could be confirmed in these complex mixtures.

In vitro Cultures of Cyclopia Plants (Honeybush) as a Source of Bioactive Xanthones and Flavanones

In vitro shoot and callus cultures of the endemic South-African shrubs: Cyclopia intermedia E. Mey., Cyclopia subternata Vogel, and Cyclopia genistoides (L.) Vent. (Fabaceae) were established and examined for the presence of polyphenolic compounds. The xanthones mangiferin and isomangiferin, as well as the flavanones hesperidin and eriocitrin were identified by LC-ESI-MS and LC-DAD, and analyzed quantitatively by HPLC. The respective intact plants were analyzed for comparison. From all in vitro cultures, the highest levels of mangiferin (1.55%) and isomangiferin (0.56%) were recorded in C. subternata microshoots, compared to 1.31% and 0.49% found in the intact plant. Callus cultures of all species synthesized only trace amounts of mangiferin and isomangiferin. Hesperidin and eriocitrin contents were significanly lower in all in vitro cultures, in comparison to the respective intact plants. Among the obtained in vitro biomasses, the highest hesperidin content was recorded in C. intermedia (0.9%) and C. subternata (0.87%) microshoots, whereas C. subternata callus was characterized by the best growth parameters and highest hesperidin content (0.69%) from all examined Cyclopia calli.

Chemoprotective properties of rooibos ( Aspalathus linearis), honeybush ( Cyclopia intermedia) herbal and green and black ( Camellia sinensis) teas against cancer promotion induced by fumonisin B 1 in rat liver

The chemoprotective properties of unfermented and fermented rooibos ( Aspalathus linearis) and honeybush ( Cyclopia intermedia) herbal teas, and green and black teas ( Camellia sinensis) were investigated against fumonisin B 1 (FB 1) promotion in rat liver utilizing diethylnitrosamine (DEN) as cancer initiator. The various teas differently affected the clinical chemical parameters associated with liver and kidney damage associated with FB 1 suggesting specific FB 1/iron/polyphenolic interactions. Green tea enhanced ( P < 0.05) the FB 1-induced reduction of the oxygen radical absorbance capacity, while fermented herbal teas and unfermented honeybush significantly ( P < 0.05) decreased FB 1-induced lipid peroxidation in the liver. The teas exhibited varying effects on FB 1-induced changes in the activities of catalase, glutathione peroxidase (GPx) glutathione reductase (GR) as well as the glutathione (GSH) status. Unfermented rooibos and honeybush significantly ( P < 0.05) to marginally ( P < 0.1) reduced the total number of foci (>10 μm), respectively, while all the teas reduced the relative amount of the larger foci. Fermentation seems to reduce the protective effect of the herbal teas. Differences in the major polyphenolic components and certain FB 1/polyphenolic/tissue interactions may explain the varying effects of the different teas on the oxidative parameters, hepatotoxic effects and cancer promotion in rat liver.

A review of the bioactivity of south African herbal teas: rooibos (Aspalathus linearis) and honeybush (Cyclopia intermedia)

Rooibos (Aspalathus linearis) and honeybush (Cyclopia intermedia) are popular tisanes in their native South Africa and have a growing worldwide market. Both herbal teas are used traditionally for medicinal purposes and are rich in polyphenols with rooibos a rare source of the dietary dihydrochalcones, aspalathin and nothofagin. The principal polyphenols in honeybush include the xanthone mangiferin and the flavonones hesperitin and isokuranetin. Despite their divergent phytochemical and nutrient compositions, rooibos and honeybush share potent antioxidant and antimutagenic activities in vitro. Animal model studies indicate both herbal teas possess potent antioxidant, immune-modulating and chemopreventive actions. However, human studies of rooibos are limited and of honeybush are absent. No adverse effects of rooibos or honeybush consumption as tisanes have been reported.