Search Results (24)

Pyranose oxidase (POx) is an FAD-dependent oxidoreductase and belongs to the glucose–methanol–choline (GMC) superfamily of oxidoreductases. As recently reported, POxs and FAD-dependent C-glycoside oxidases (CGOxs) share the same sequence space, and phylogenetic analysis of actinobacterial sequences belonging to this shared sequence space showed that it can be divided into four clades. Here, we report the biochemical characterization of a POx/CGOx from Microbacterium sp. 3H14 (MPOx), belonging to the hitherto unexplored clade II of actinobacterial POx/CGOx. Overall, MPOx demonstrates comparable features to POxs/CGOxs of clades III and IV, including the preference for glycosides over monosaccharides as electron donors. However, as MPOx efficiently oxidizes the C-glycoside aspalathin as well as the O-glycoside phlorizin, it shows activity with yet another set of glycoside structures compared to other POx/CGOx members. Full article

The excessive dietary intake of simple sugars and abnormal metabolism in certain diseases contribute to the increased production of α-dicarbonyls (α-DCs), such as methylglyoxal (MGO) and glyoxal (GO), the main precursors of the formation of advanced glycation end products (AGEs). AGEs play a vital role, for example, in the development of cardiovascular diseases and diabetes. Aspalathus linearis (Burman f.) R. Dahlgren (known as rooibos tea) exhibits a wide range of activities beneficial for cardio-metabolic health. Thus, the present study aims to investigate unfermented and fermented rooibos extracts and their constituents for the ability to trap MGO and GO. The individual compounds identified in extracts were tested for the capability to inhibit AGEs (with MGO or GO as a glycation agent). Ultra-high-performance liquid chromatography coupled with an electrospray ionization mass spectrometer (UHPLC–ESI–MS) was used to investigate α-DCs’ trapping capacities. To evaluate the antiglycation activity, fluorescence measurement was used. The extract from the unfermented rooibos showed a higher ability to capture MGO/GO and inhibit AGE formation than did the extract from fermented rooibos, and this effect was attributed to a higher content of dihydrochalcones. The compounds detected in the extracts, such as aspalathin, nothofagin, vitexin, isovitexin, and eriodictyol, as well as structurally related phloretin and phloroglucinol (formed by the biotransformation of certain flavonoids), trapped MGO, and some also trapped GO. AGE formation was inhibited the most by isovitexin. However, it was the high content of aspalathin and its higher efficiency than that of metformin that determined the antiglycation and trapping properties of green rooibos. Therefore, A. linearis, in addition to other health benefits, could potentially be used as an α-DC trapping agent and AGE inhibitor. Full article

Epigenetic mechanisms play an important role in the etiology of colorectal cancer (CRC) and other malignancies due, in part, to deregulated bromodomain (BRD) functions. Inhibitors of the bromodomain and extraterminal (BET) family have entered into clinical trials as anticancer agents, and interest has grown in other acetyl ‘reader’ proteins as therapeutic targets, including non-BET member bromodomain-containing protein 9 (BRD9). We report here that overexpression of BRD9 is associated with poor prognosis in CRC patients, and that siRNA-mediated knockdown of BRD9 decreased cell viability and activated apoptosis in human colon cancer cells, coincident with increased DNA damage. Seeking natural compounds as BRD9 antagonists, molecular docking in silico identified several polyphenols such as Epigallocatechin-3-gallate (EGCG), Equol, Quercetin, and Aspalathin, with favorable binding energies, supported by BROMOscan^® (DiscoverX) and isothermal titration calorimetry experiments. Polyphenols mimicked BRD9 knockdown and iBRD9 treatment in reducing colon cancer cell viability, inhibiting colony formation, and enhancing DNA damage and apoptosis. Normal colonic epithelial cells were unaffected, signifying cancer-specific effects. These findings suggest that natural polyphenols recognize and target BRD9 for inhibition, and might serve as useful lead compounds for bromodomain therapeutics in the clinical setting. Full article

Aspalathus linearis (Burm.f.) R. Dahlgren, commonly known as rooibos tea, was consumed traditionally by the indigenous South African inhabitants as an herbal remedy. Beside antioxidant properties, it displays antiallergic, antispasmodic, and hypoglycemic activities. An ultra-high-performance liquid chromatography method coupled with photodiode array and mass spectrometry detectors were developed for the determination of 14 phenolic constituents from leaves and stems of A. linearis. The efficient separation was performed within 30 min at a temperature of 30 °C by using C-18 column as the stationary phase and water/acetonitrile with 0.05% formic acid as the mobile phase. Method validation for linearity, repeatability, limits of detection, and limits of quantification was achieved. The limits of detection from 0.2–1 μg/mL were reported for the standard compounds. Their total content varied substantially (1.50–9.85 mg/100 mg sample) in 21 dietary supplements. The presence of regioisomers and diastereomers which co-elute on a variety of stationary phases make separation for quantification purposes challenging. This method was found to be efficient in providing low retention times and excellent resolution for this type of phytochemicals. The established method is suitable for chemical fingerprint analysis of A. linearis and cost-effective for quality control of rooibos tea products. Full article

Antioxidant activity associated with green rooibos infusions is attributed to the activity of polyphenols, particularly aspalathin and nothofagin. This study aimed to optimise β-cyclodextrin (β-CD)-assisted extraction of crude green rooibos (CGRE) via total polyphenolic content (TPC) and antioxidant activity assays. Response surface methodology (RSM) permitted optimisation of β-CD concentration (0–15 mM), temperature (40–90 °C) and time (15–60 min). Optimal extraction conditions were: 15 mM β-CD: 40 °C: 60 min with a desirability of 0.985 yielding TPC of 398.25 mg GAE·g⁻¹, metal chelation (MTC) of 93%, 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging of 1689.7 µmol TE·g⁻¹, ferric reducing antioxidant power (FRAP) of 2097.53 µmol AAE·g⁻¹ and oxygen radical absorbance capacity (ORAC) of 11,162.82 TE·g⁻¹. Aspalathin, hyperoside and orientin were the major flavonoids, with quercetin, luteolin and chrysoeriol detected in trace quantities. Differences (p < 0.05) between aqueous and β-CD assisted CGRE was only observed for aspalathin reporting the highest content of 172.25 mg·g⁻¹ of dry matter for extracts produced at optimal extraction conditions. Positive, strong correlations between TPC and antioxidant assays were observed and exhibited regression coefficient (R²) between 0.929–0.978 at p < 0.001. These results demonstrated the capacity of β-CD in increasing polyphenol content of green rooibos. Full article

Rooibos tea (Apalathus linearis) extract, rich in glycosylated polyphenols (aspalathin and nothofagin), has been shown to improve glycaemic responses in individuals with prediabetes, a high-risk state for developing type 2 diabetes (T2D). However, evidence is scarce regarding its impact on Glucagon-like-peptide-1 (GLP-1). GLP-1 stimulates 50–70% of insulin production during a meal, also known as the incretin effect. Individuals with prediabetes may therefore benefit from an increase in GLP-1 concentration. On the other hand, a decrease in GLP-1 may indicate heightened incretin sensitivity, resulting in a reduced demand for GLP-1 secretion needed to improve glycaemic responses. We conducted an acute, single-blind, placebo-controlled, non-randomised, crossover study (GLARE study; ACTRN12617000837325) examining the impact of rooibos tea extract on GLP-1_total and GLP-1_active concentrations in participants with prediabetes. Nineteen participants (aged 65.0 ± 1.6 years, BMI 27.3 ± 1.1 kg/m², and HbA1c 42 ± 1 mmol/mol) were given a placebo or rooibos tea extract on separate occasions before an oral glucose tolerance test (OGTT). Blood samples were collected at 0, 30, 60, 90 and 120 min. Data were analysed using a linear mixed model for repeated measures. Secondary analysis was conducted by stratifying participants into either a healthier or less healthy prediabetes subgroup, with the less healthy group experiencing delayed postprandial glucose and/or insulin peaks. The study outcomes demonstrated that although prior to stratification there were no significant changes in the overall total incremental area under the curve (iAUC_total) of GLP-1_total and GLP-1_active in participants (p > 0.05), the healthier prediabetes subgroup exhibited a significant reduction in GLP-1_active compared to the control group (479.4 vs. 1046.7 pM.min, p = 0.038, effect size Cohen’s d = 0.6). This suggests that rooibos tea extract may reduce postprandial incretin demand in people with prediabetes. More study is warranted to confirm this observation. Full article

In the pursuit of bioactive phytochemicals as a therapeutic strategy to manage metabolic risk factors for type 2 diabetes (T2D), aspalathin, C-glucosyl dihydrochalcone from rooibos (Aspalathus linearis), has received much attention, along with its C-glucosyl flavone derivatives and phlorizin, the apple O-glucosyl dihydrochalcone well-known for its antidiabetic properties. We provided context for dietary exposure by highlighting dietary sources, compound stability during processing, bioavailability and microbial biotransformation. The review covered the role of these compounds in attenuating insulin resistance and enhancing glucose metabolism, alleviating gut dysbiosis and associated oxidative stress and inflammation, and hyperuricemia associated with T2D, focusing largely on the literature of the past 5 years. A key focus of this review was on emerging targets in the management of T2D, as highlighted in the recent literature, including enhancing of the insulin receptor and insulin receptor substrate 1 signaling via protein tyrosine phosphatase inhibition, increasing glycolysis with suppression of gluconeogenesis by sirtuin modulation, and reducing renal glucose reabsorption via sodium-glucose co-transporter 2. We conclude that biotransformation in the gut is most likely responsible for enhancing therapeutic effects observed for the C-glycosyl parent compounds, including aspalathin, and that these compounds and their derivatives have the potential to regulate multiple factors associated with the development and progression of T2D. Full article

The green chemistry approach has continuously been applied for the synthesis of functional nanomaterials to reduce waste, environmental hazards, and the use of toxic chemicals among other reasons. Bioactive natural compounds have been found great potential in this regard and are used to improve the stability, activity, and biodistribution of metal nanoparticles (MNPs). Aspalathin (ASP) from Aspalathus linearis (rooibos) has a well-defined pharmacological profile and functional groups capable of both reducing and capping agents in the synthesis of metallic nanoparticles (NP). This study provides the first report of the phytomediated synthesis of gold and silver nanoparticles (AuNPs/AgNPs) via ASP and the green rooibos (GR) extract. The study demonstrated a green chemistry approach to the biosynthesis of nanoparticles of GR-AuNPs, ASP-AuNPs, GR-AgNPs, and ASP-AgNPs. The results showed that GR and ASP could act both as reducing and stabilising agents in the formation of crystalline, with different shapes and dispersity of NPs in the ranges of 1.6–6.7 nm for AgNPs and 7.5–12.5 nm for the AuNPs. However, the ASP NPs were less stable in selected biogenic media compared to GR NPs and were later stabilised with polyethene glycol. The cytotoxicity studies showed that GR-AgNPs were the most cytotoxic against SH-SY5Y and HepG2 with IC₅₀ 108.8 and 183.4 μg/mL, respectively. The cellular uptake analysis showed a high uptake of AuNPs and indicated that AgNPs of rooibos at a lower dose (1.3–1.5 μg/mL) is favourable for its anticancer potential. This study is a contribution to plant-mediated metallic nanoparticles using a pure single compound that can be further developed for targeted drug delivery for cancer cells treatments in the coming years. Full article

The current study investigated the physiological effects of flavonoids found in daily consumed rooibos tea, aspalathin, isoorientin, and orientin on improving processes involved in mitochondrial function in C2C12 myotubes. To achieve this, C2C12 myotubes were exposed to a mitochondrial channel blocker, antimycin A (6.25 µM), for 12 h to induce mitochondrial dysfunction. Thereafter, cells were treated with aspalathin, isoorientin, and orientin (10 µM) for 4 h, while metformin (1 µM) and insulin (1 µM) were used as comparators. Relevant bioassays and real-time PCR were conducted to assess the impact of treatment compounds on some markers of mitochondrial function. Our results showed that antimycin A induced alterations in the mitochondrial respiration process and mRNA levels of genes involved in energy production. In fact, aspalathin, isoorientin, and orientin reversed such effects leading to the reduced production of intracellular reactive oxygen species. These flavonoids further enhanced the expression of genes involved in mitochondrial function, such as Ucp 2, Complex 1/3, Sirt 1, Nrf 1, and Tfam. Overall, the current study showed that dietary flavonoids, aspalathin, isoorientin, and orientin, have the potential to be as effective as established pharmacological drugs such as metformin and insulin in protecting against mitochondrial dysfunction in a preclinical setting; however, such information should be confirmed in well-established in vivo disease models. Full article

Eranthis longistipitata Regel is an endemic plant of Central Asia. The flavonoid profile of E. longistipitata leaves was studied by mass spectrometry for the first time (natural populations of Kyrgyzstan and Uzbekistan, in 70% aqueous–ethanol extracts by liquid chromatography coupled with high-resolution mass spectrometry). Mass spectrometry revealed 18 flavonoid compounds. Flavonols featured the highest diversity, and 10 such substances were identified: 2 free aglycones (quercetin and kaempferol), 6 quercetin glycosides (peltatoside, hyperoside, reynoutrin, quercetin 3-sambubioside, rutin, and isoquercitrin), and 2 kaempferol glycosides (juglalin and trifolin). Two flavans (cianidanol and auriculoside), two hydroxyflavanones (6-methoxytaxifolin and aromadendrin), and one C-glycoside flavone—carlinoside—were identified. Dihydroxychalcones aspalathin, phloridzin, and phloretin were found too. Levels of rutin, quercetin, kaempferol, and hyperoside were confirmed by means of standards and high-performance liquid chromatography. Rutin concentration was the highest among all other identified flavonoid compounds: in the leaf samples from Kyrgyzstan, it ranged from 2.46 to 3.20 mg/g, and in those from Uzbekistan, from 1.50 to 3.01 mg/g. The diversity of flavonoid compounds in E. longistipitata leaves is probably due to external ecological and geographic factors and adaptive mechanisms. Full article

Skin cells suffer continuous damage from chronic exposure to ultraviolet light (UV) that may result in UV-induced oxidative stress and skin thinning. This has necessitated the formulation of cosmeceutical products rich in natural antioxidants and free radical scavengers. Aspalathus linearis (rooibos) is an endemic South African fynbos plant growing naturally in the Western Cape region. The plant is rich in phenolics and other bioactives with a wide spectrum of health benefits. The chemical study of an acetonic extract of green A. linearis afforded a novel compound named linearthin (1) and two known dihydrochalcones, aspalathin (2) and nothofagin (3). The chemical structure of the novel compound was elucidated based on spectroscopic data analysis. The bio-evaluation of the isolated chalcones in vitro for protection against UVB-induced oxidative stress were systematically assessed by examining cell viability, metabolic activity, apoptosis, and cytotoxicity using HaCaT and SK-MEL-1 skin cells models. It was observed that pre-treatment with tested samples for 4- and 24 h at low concentrations were sufficient to protect skin cells from UVB-induced damage in vitro as evidenced by higher cell viability and improved metabolic activity in both keratinocytes (HaCaT) and melanocytes (SK-MEL-1). The results further show that the pre-treatment regimen employed by this study involved some degree of cellular adaptation as evidenced by higher levels of reduced glutathione with a concomitant decrease in lipid peroxidation and lowered caspase 3 activity. Furthermore, compound 1 was most cytoprotective against UVB irradiation of HaCaT cell line (over 24 h) with an IC₅₀ of 282 µg/mL and SK-MEL-1 cell line with IC₅₀ values of 248.3 and 142.6 µg/mL over 4 and 24 h, respectively. On the other hand, HaCaT cells exposed to 2 over 4 h before UVB irradiation showed the highest degree of cytoprotection with an IC₅₀ of 398.9 µg/mL among the four studied samples. These results show that linearthin (1) and the two glycoside dihydrochalcone of A. linearis have the potential to be further developed as antioxidant cosmeceutical ingredients that may protect skin against UVB-induced damage. Full article

Green rooibos extract (GRE), shown to improve hyperglycemia and HDL/LDL blood cholesterol, has potential as a nutraceutical beverage ingredient. The main bioactive compound of the extract is aspalathin, a C-glucosyl dihydrochalcone. The study aimed to determine the effect of common iced tea ingredients (citric acid, ascorbic acid, and xylitol) on the stability of GRE, microencapsulated with inulin for production of a powdered beverage. The stability of the powder mixtures stored in semi-permeable (5 months) and impermeable (12 months) single-serve packaging at 30 °C and 40 °C/65% relative humidity was assessed. More pronounced clumping and darkening of the powders, in combination with higher first order reaction rate constants for dihydrochalcone degradation, indicated the negative effect of higher storage temperature and an increase in moisture content when stored in the semi-permeable packaging. These changes were further increased by the addition of crystalline ingredients, especially citric acid monohydrate. The sensory profile of the powders (reconstituted to beverage strength iced tea solutions) changed with storage from a predominant green-vegetal aroma to a fruity-sweet aroma, especially when stored at 40 °C/65% RH in the semi-permeable packaging. The change in the sensory profile of the powder mixtures could be attributed to a decrease in volatile compounds such as 2-hexenal, (Z)-2-heptenal, (E)-2-octenal, (E)-2-nonenal, (E,Z)-2,6-nonadienal and (E)-2-decenal associated with “green-like” aromas, rather than an increase in fruity and sweet aroma-impact compounds. Green rooibos extract powders would require storage at temperatures ≤ 30 °C and protection against moisture uptake to be chemically and physically shelf-stable and maintain their sensory profiles. Full article

Recent evidence shows that rooibos compounds, aspalathin and phenylpyruvic acid-2-O-β-d-glucoside (PPAG), can independently protect cardiomyocytes from hyperglycemia-related reactive oxygen species (ROS). While aspalathin shows more potency by enhancing intracellular antioxidant defenses, PPAG acts more as an anti-apoptotic agent. Thus, to further understand the protective capabilities of these compounds against hyperglycemia-induced cardiac damage, their combinatory effect was investigated and compared to metformin. An in vitro model of H9c2 cardiomyocytes exposed to chronic glucose concentrations was employed to study the impact of such compounds on hyperglycemia-induced damage. Here, high glucose exposure impaired myocardial substrate utilization by abnormally enhancing free fatty acid oxidation while concomitantly suppressing glucose oxidation. This was paralleled by altered expression of genes involved in energy metabolism including acetyl-CoA carboxylase (ACC), 5′ AMP-activated protein kinase (AMPK), and peroxisome proliferator-activated receptor-alpha (PPARα). The combination treatment improved myocardial substrate metabolism, maintained mitochondrial membrane potential, and attenuated various markers for oxidative stress including nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and glutathione content. It also showed a much-improved effect by ameliorating DNA damage when compared to metformin. The current study demonstrates that rooibos compounds offer unique cardioprotective properties against hyperglycemia-induced and potentially against diabetes-induced cardiac damage. These data also support further exploration of rooibos compounds to better assess the cardioprotective effects of different bioactive compound combinations. Full article

Type 2 diabetic patients possess a two to four-fold-increased risk for cardiovascular diseases (CVD). Hyperglycemia, oxidative stress associated with endothelial dysfunction and dyslipidemia are regarded as pro-atherogenic mechanisms of CVD. In this study, high-fat diet-induced diabetic and non-diabetic vervet monkeys were treated with 90 mg/kg of aspalathin-rich green rooibos extract (Afriplex GRT) for 28 days, followed by a 1-month wash-out period. Supplementation showed improvements in both the intravenous glucose tolerance test (IVGTT) glycemic area under curve (AUC) and total cholesterol (due to a decrease of the low-density lipoprotein [LDL]) values in diabetics, while non-diabetic monkeys benefited from an increase in high-density lipoprotein (HDL) levels. No variation of plasma coenzyme Q10 (CoQ₁₀) were found, suggesting that the LDL-lowering effect of Afriplex GRT could be related to its ability to modulate the mevalonate pathway differently from statins. Concerning the plasma oxidative status, a decrease in percentage of oxidized CoQ₁₀ and circulating oxidized LDL (ox-LDL) levels after supplementation was observed in diabetics. Finally, the direct correlation between the amount of oxidized LDL and total LDL concentration, and the inverse correlation between ox-LDL and plasma CoQ₁₀ levels, detected in the diabetic monkeys highlighted the potential cardiovascular protective role of green rooibos extract. Taken together, these findings suggest that Afriplex GRT could counteract hyperglycemia, oxidative stress and dyslipidemia, thereby lowering fundamental cardiovascular risk factors associated with diabetes. Full article

Aspalathin, the main polyphenol of rooibos (Aspalathus linearis), is associated with diverse health promoting properties of the tea. During fermentation, aspalathin is oxidized and concentrations are significantly reduced. Standardized methods for quality control of rooibos products do not investigate aspalathin, since current techniques of aspalathin detection require expensive equipment and expertise. Here, we describe a simple and fast thin-layer chromatography (TLC) method that can reproducibly visualize aspalathin in rooibos herbal tea and plant extracts at a limit of detection (LOD) equal to 178.7 ng and a limit of quantification (LOQ) equal to 541.6 ng. Aspalathin is a rare compound, so far only found in A. linearis and its (rare) sister species A. pendula. Therefore, aspalathin could serve as a marker compound for authentication and quality control of rooibos products, and the described TLC method represents a cost-effective approach for high-throughput screening of plant and herbal tea extracts. Full article