Food-grade phytosome vesicles for nanoencapsulation of labile C-glucosylated xanthones and dihydrochalcones present in a plant extract matrix-Effect of process conditions and stability assessment.

Phytosomes consist of a phytochemical bound to the hydrophilic choline head of a phospholipid. Their use in food products is gaining interest. However, literature on the use of food-grade solvents, crude plant extracts as opposed to pure compounds, and unrefined phospholipids to prepare phytosomes is limited. Furthermore, studies on compound stability are lacking. This study aimed to develop nano-phytosome vesicles prepared from inexpensive food-grade ingredients to improve the stability of polyphenolic compounds. Cyclopia subternata extract (CSE) was selected as a source of phenolic compounds. It contains substantial quantities of C-glucosyl xanthones, benzophenones, and dihydrochalcones, compounds largely neglected to date. The effect of process conditions on the complexation of CSE polyphenols with minimally refined food-grade fat-free soybean lecithin (PC) was studied. The PC:CSE ratio, sonication time, and reaction temperature were varied. This resulted in phytosomes ranging in vesicle size (113.7-312.7 nm), polydispersity index (0.31-0.48), and zeta potential (-55.0 to -38.9 mV). Variation was also observed in the yield (93.5%-96.0%), encapsulation efficiency (3.7%-79.0%), and loading capacity (LC, 1.3%-14.7%). Vesicle size and LC could be tailored by adjusting the sonication time and PC:CSE ratio, respectively. Chemical interaction between the lipid and the phenolic compounds was confirmed with nuclear magnetic resonance. Phytosomal formulation protected the compounds against degradation when freeze-dried samples were stored at 25 and 40°C for 6 months at low relative humidity. The study provided valuable information on the importance of specific process parameters in producing food-grade phytosomes with improved phenolic stability.

Stability of labile xanthones and dihydrochalcones in a ready-to-drink honeybush beverage during storage.

BACKGROUND: The shelf-life of a functional herbal tea-based beverage is important not only for consumer acceptability, but also for the retention of bioactive compounds. The present study aimed to clarify the role of common iced tea beverage ingredients (citric and ascorbic acids) on the shelf-life stability of an herbal tea-based beverage. A hot water extract of green Cyclopia subternata, also used as honeybush tea, was selected as the main ingredient because it provides different types of phenolic compounds associated with bioactive properties (i.e. xanthones, benzophenones, flavanones, flavones and dihydrochalcones). RESULTS: The model solutions were stored for 180 and 90 days at 25 and 40 °C, respectively. Changes in their volatile profiles and color were also quantified as they contribute to product quality. 3',5'-Di-ß-d-glucopyranosyl-3-hydroxyphloretin (HPDG; dihydrochalcone) and, to a lesser extent, mangiferin (xanthone), were the most labile compounds. Both compounds were thus identified as critical quality indicators to determine shelf-life. The stability-enhancing activity of the acids depended on the compound; ascorbic acid and citric acid enhanced the stability of HPDG and mangiferin, respectively. However, when considering all the major phenolic compounds, the base solution without acids was the most stable. This was also observed for the color and major volatile aroma-active compounds [α-terpineol, (E)-ß-damascenone, 1-p-menthen-9-al and trans-ocimenol]. CONCLUSION: The addition of acids, added for stability and taste in ready-to-drink iced tea beverages, could thus have unwanted consequences in that they could accelerate compositional changes and shorten the shelf-life of polyphenol-rich herbal tea beverages. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Class-modelling of overlapping classes. A two-step authentication approach.

Honeybush is an indigenous herbal tea highly valued for its aroma, flavour and medicinal properties. It is protected as Geographical Indication (GI) since it is produced from a number of Cyclopia species that are endemic to South Africa. Most commonly used for honeybush tea production are C. intermedia, C. subternata and C. genistoides, differing slightly, but distinctly in flavour. Demand for species-specific honeybush tea instead of mixtures have increased, meriting a strategy for authentication of C. intermedia, C. subternata and C. genistoides. Samples of these three species were analysed, using hyperspectral imaging (HSI) in the near-infrared spectral range. The data were pre-processed and used for class-modelling, a general approach well suited for authentication purposes. Unfortunately, since the HSI data of Cyclopia species studied are very similar, the classification results obtained with individual class-models are unsatisfactory, e.g., class-models constructed for C. genistoides and C. subternata yielded correct classification rate (CCR) values of 76.4 and 83.1%, respectively. On the other hand, discriminant modelling, which is another type of classification technique, led to good classification outcomes (CCR 98.9%). However, the classical discriminant model cannot be applied for authentication purposes since it always assigns a new sample to one of the classes studied, even if in reality, it belongs to none of them. Counterfeits or non-representative samples would be incorrectly assigned by the discriminant model to one of the authentic classes. Therefore, in this study, a two-step authentication of overlapping classes is proposed, which combines the advantages of class-modelling and discriminant methods. When applied to the authentication of Cyclopia species studied, the two-step approach yielded a CCR of 97.4%, which is a significant improvement compared to results obtained with the individual class-models. The proposed approach is general and can be applied when classes studied are very similar, and individual class-models lead to unsatisfactory results.

Heat treatment improves the sensory properties of the ultrafiltration by-product of honeybush (Cyclopia genistoides) extract.

BACKGROUND: Ultrafiltration of green honeybush (Cyclopia genistoides) extract results in a by-product (retentate). Application of further separation processes for recovery of polyphenols would entail creation of additional waste. Repurposing the retentate as a food flavour ingredient provides an alternative valorization approach. RESULTS: The retentate, suspended in water (270 g L-1 ), was heat-treated at 80 °C for 2, 4, 8 and 16 h, and at 90 °C for 2, 4, 6 and 8 h to change its sensory profile. The heat-treated retentate, diluted to beverage strength (2.15 g L-1 ), had prominent 'grape/Muscat-like' and 'marmalade/citrus' aroma and flavour notes. Overall, heating for ≤ 4 h increased the intensities of positive flavour and aroma notes, while reducing those of 'green/grass', 'hay' and bitterness, whereafter further heating only had a slight effect on the aroma profile at 80 °C (P < 0.05), but not at 90 °C (P ≥ 0.05). The heat treatments, 80 °C/4 h and 90 °C/4 h, were subsequently applied to different batches of retentate (n = 10) to accommodate the effect of natural product variation. Heating at 90 °C produced higher intensities of positive aroma attributes (P < 0.05), but was more detrimental to the phenolic stability, compared to 80 °C. CONCLUSION: After heat treatment, the phenolic content of C. genistoides retentate, reconstituted to beverage strength, still fell within the range of a typical 'fermented' (oxidized) honeybush leaf tea infusion. The change in phenolic composition will not diminish the benefit of an improved sensory profile for the retentate by-product through heating. © 2021 Society of Chemical Industry.

Shelf-Life Stability of Ready-to-Use Green Rooibos Iced Tea Powder-Assessment of Physical, Chemical, and Sensory Properties.

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.

Identification of a novel di-C-glycosyl dihydrochalcone and the thermal stability of polyphenols in model ready-to-drink beverage solutions with Cyclopia subternata extract as functional ingredient.

Heat processing of ready-to-drink beverages is required to ensure a microbiologically safe product, however, this can result in the loss of bioactive compounds responsible for functionality. The objective of this study was to establish the thermal stability of a novel dihydrochalcone, 3',5'-di-ß-d-glucopyranosyl-3-hydroxyphloretin (2), 3',5'-di-ß-d-glucopyranosylphloretin (3) and other Cyclopia subternata phenolic compounds, in model solutions with or without citric acid and ascorbic acid. The solutions were heated at 93, 121 and 135 °C, relevant to pasteurisation, commercial sterilisation and ultra-high temperature (UHT) pasteurisation, respectively. For most compounds, the acids decreased the second order reaction rate constants, up to 27 times. Compound 2 (46.29 ± 0.53 (g/100 g)-1 h-1), and to a lesser extent compound 3 (5.94 ± 0.01 (g/100 g)-1 h-1) were the most thermo-unstable compounds when treated at 135 °C without added acids. Even though differential effects were observed for compounds at different temperatures and formulations, overall, the phenolic compounds were most stable under UHT pasteurisation conditions.

Honeybush Extracts (Cyclopia spp.) Rescue Mitochondrial Functions and Bioenergetics against Oxidative Injury.

Mitochondrial dysfunction plays a major role not only in the pathogenesis of many oxidative stress or age-related diseases such as neurodegenerative as well as mental disorders but also in normal aging. There is evidence that oxidative stress and mitochondrial dysfunction are the most upstream and common events in the pathomechanisms of neurodegeneration. Cyclopia species are endemic South African plants and some have a long tradition of use as herbal tea, known as honeybush tea. Extracts of the tea are gaining more scientific attention due to their phenolic composition. In the present study, we tested not only the in vitro mitochondria-enhancing properties of honeybush extracts under physiological conditions but also their ameliorative properties under oxidative stress situations. Hot water and ethanolic extracts of C. subternata, C. genistoides, and C. longifolia were investigated. Pretreatment of human neuroblastoma SH-SY5Y cells with honeybush extracts, at a concentration range of 0.1-1 ng/ml, had a beneficial effect on bioenergetics as it increased ATP production, respiration, and mitochondrial membrane potential (MMP) after 24 hours under physiological conditions. The aqueous extracts of C. subternata and C. genistoides, in particular, showed a protective effect by rescuing the bioenergetic and mitochondrial deficits under oxidative stress conditions (400 µM H2O2 for 3 hours). These findings indicate that honeybush extracts could constitute candidates for the prevention of oxidative stress with an impact on aging processes and age-related neurodegenerative disorders potentially leading to the development of a condition-specific nutraceutical.

Model development for predicting in vitro bio-capacity of green rooibos extract based on composition for application as screening tool in quality control.

Mounting evidence of the ability of aspalathin to target underlying metabolic dysfunction relevant to the development or progression of obesity and type 2 diabetes created a market for green rooibos extract as a functional food ingredient. Aspalathin is the obvious choice as a chemical marker for extract standardisation and quality control, however, often the concentration of a single constituent of a complex mixture such as a plant extract is not directly related to its bio-capacity, i.e. the level of in vitro bioactivity effected in a cell system at a fixed concentration. Three solvents (hot water and two EtOH-water mixtures), previously shown to produce bioactive green rooibos extracts, were selected for extraction of different batches of rooibos plant material (n = 10). Bio-capacity of the extracts, tested at 10 µg ml-1, was evaluated in terms of glucose uptake by C2C12 and C3A cells and lipid accumulation in 3T3-L1 cells. The different solvents and inter-batch plant variation delivered extracts ranging in aspalathin content from 54.1 to 213.8 g kg-1. The extracts were further characterised in terms of other major flavonoids (n = 10) and an enolic phenylpyruvic acid glucoside, using HPLC-DAD. The 80% EtOH-water extracts, with the highest mean aspalathin content (170.9 g kg-1), had the highest mean bio-capacity in the respective assays. Despite this, no significant (P≥ 0.05) correlation existed between aspalathin content and bio-capacity, while the orientin, isoorientin and vitexin content correlated moderately (r≥ 0.487; P < 0.05) with increased glucose uptake by C2C12 cells. Various multivariate analysis methods were then applied with Evolution Program-Partial Least Squares (EP-PLS) resulting in models with the best predictive power. These EP-PLS models, based on all quantified compounds, predicted the bio-capacity of the extracts for the respective cell types with RMSECV values ≤ 11.5, confirming that a complement of compounds, and not aspalathin content alone, is needed to predict the in vitro bio-capacity of green rooibos extracts. Additionally, the composition of hot water infusions of different production batches of green rooibos (n = 29) at 'cup-of-tea' equivalence was determined to relate dietary supplementation with the extract to intake in the form of herbal tea.

Impact of Cold versus Hot Brewing on the Phenolic Profile and Antioxidant Capacity of Rooibos (Aspalathus linearis) Herbal Tea.

Consumption of rooibos (Aspalathus linearis) as herbal tea is growing in popularity worldwide and its health-promoting attributes are mainly ascribed to its phenolic composition, which may be affected by the brewing conditions used. An aspect so far overlooked is the impact of cold brewing vs regular brewing and microwave boiling on the poly(phenolic) profile and in vitro antioxidant capacity of infusions prepared from red ('fermented', oxidized) and green ('unfermented', unoxidized) rooibos, the purpose of the present study. By using an untargeted metabolomics-based approach (UHPLC-QTOF mass spectrometry), 187 phenolic compounds were putatively annotated in both rooibos types, with flavonoids, tyrosols, and phenolic acids the most represented type of phenolic classes. Multivariate statistics (OPLS-DA) highlighted the phenolic classes most affected by the brewing conditions. Similar antioxidant capacities (ORAC and ABTS assays) were observed between cold- and regular-brewed green rooibos and boiled-brewed red rooibos. However, boiling green and red rooibos delivered infusions with the highest antioxidant capacities and total polyphenol content. The polyphenol content strongly correlated with the in vitro antioxidant capacities, especially for flavonoids and phenolic acids. These results contribute to a better understanding of the impact of the preparation method on the potential health benefits of rooibos tea.

Potential of benzophenones and flavanones to modulate the bitter intensity of Cyclopia genistoides herbal tea.

Variation in the bitter taste of Cyclopia genistoides (honeybush) herbal tea and reported modulation between its major xanthones, mangiferin and isomangiferin, prompted further investigation into the potential modulatory effects of honeybush phenolics. Combinations of crude benzophenone (BF)-, xanthone (XF)-, and flavanone (FF)-rich fractions and their major individual phenolic compounds were analysed by descriptive sensory analysis. The fractions were prepared from a bitter, hot water extract of green C. genistoides. Fraction BF, which is below the bitter threshold (intensity 10 on 100-point scale), enhanced the bitter intensity of XF and FF slightly (p < 0.05), although none of the major individual benzophenones retained this bitter enhancing effect. On the contrary, 3-ß-d-glucopyranosyl-4-ß-d-glucopyranosyloxyiriflophenone, the major benzophenone in BF, significantly (p < 0.05) decreased the bitter taste of XF, at a low concentration, whereas FF suppressed the bitter intensity of XF and mangiferin, the major xanthone present in XF. Hesperidin, however, had no effect on the bitter intensity of XF. In contrast, (2S)-5-[α-L-rhamnopyranosyl-(1→2)-ß-d-glucopyranosyloxy]-naringenin, the major compound of FF, significantly (p < 0.05) enhanced the bitter taste of XF when added at concentrations comparable to that of 'fermented' honeybush tea infusions. The concentration-dependence of these bitter taste interactions may be responsible for the variable bitter intensity of C. genistoides herbal tea.

Membrane selection and optimisation of tangential flow ultrafiltration of Cyclopia genistoides extract for benzophenone and xanthone enrichment.

Ultrafiltration of Cyclopia genistoides extract was optimised to increase its benzophenone and xanthone content as quantified using HPLC-DAD. Regenerated cellulose (RC) and polyethersulphone membranes with molecular weight cut-offs of 10 and 30 kDa were evaluated in terms of compound enrichment, permeate flux and permeate yield, using dead-end ultrafiltration. Compound enrichment was subsequently optimised using the 10 kDa RC membrane and tangential flow ultrafiltration (TFU). The effect of extract composition on compound enrichment, due to natural variation in the source material, was assessed using extracts from different batches of plant material (n = 11). Transmembrane pressure and feed flow rate affected (p < 0.05) process efficiency (mean permeate flux, compound enrichment and membrane fouling). TFU achieved ≥20% enrichment of the target compounds, proving its suitability for preparation of a nutraceutical extract of C. genistoides.

Rooibos agro-processing waste as herbal tea products: optimisation of soluble solids extraction from dust and application to improve sensory profile, colour and flavonoid content of stem infusions.

BACKGROUND: Rooibos represents 10% of the global herbal tea market. Shrinking production areas as a result of climate change necessitate the maximum conversion of plant biomass to product. The present study aimed to determine the potential of rooibos tea processing waste (i.e. fine dust and coarse stems) as potential flavour and herbal tea ingredients, respectively. RESULTS: Hot water extraction of soluble solids (SS) from rooibos dust was optimised and extracts from different production batches (n = 20) were prepared. Their sensory profiles were similar, although less intense than that of infusions of commercial rooibos (n = 20) when diluted to the same SS content. The turbidity and flavonoid content of the diluted extracts was mostly lower (P < 0.05) than that of commercial rooibos. An atypical and negative aroma attribute, 'planky/pencil shavings', was predominant in the stem infusions (n = 20), which contained less SS (P < 0.05) than commercial rooibos. Blends of stem infusion and extract could not effectively mask this negative aroma note (P > 0.05). CONCLUSION: Rooibos dust could be used to produce a rooibos flavour extract, whereas the prominent atypical, negative 'planky/pencil shavings' aroma note of the stems would limit their inclusion in commercial rooibos blends. © 2019 Society of Chemical Industry.

Bitter profiling of phenolic fractions of green Cyclopia genistoides herbal tea.

The bitter taste of Cyclopia genistoides infusions is unacceptable to consumers, who are used to the slightly sweet taste of the herbal teas produced from other Cyclopia species. Bitter taste intensities of crude phenolic fractions of a bitter hot water extract of C. genistoides were determined by a trained panel to identify the fraction contributing most to the bitter taste. Fractions, enriched in benzophenones (B), xanthones (X) and flavanones (F), and each tested at their infusion equivalent concentration (IEC) scored 5, 31 and 13 (on a 100-point scale), respectively. Fraction B, containing mostly iriflophenone glucosides, was perceived as not bitter. The major xanthone in fraction X, mangiferin, was significantly (p < 0.05) more bitter than its regio-isomer, isomangiferin, at equal concentration. A mixture of isomangiferin and mangiferin at their IECs was significantly (p < 0.05) less bitter than the mangiferin solution alone, indicating bitter suppression by isomangiferin.

Impact of steam treatment on shelf-life stability of a xanthone-rich green herbal tea (Cyclopia maculata Andrews Kies) - identifying quality changes during storage.

BACKGROUND: Steam treatment of shredded, fresh C. maculata (honeybush) plant material improves the aroma of this green herbal tea with a slight impact on color and phenolic content, but the effect on storage stability is not known. RESULTS: Steam-treated (60 s before drying) and untreated (control) dried plant material was stored under normal storage conditions in semi-permeable sachets at 25 °C and 60% relative humidity. Reference samples of treated (steamed) and untreated (control) material were stored at 0 °C in impermeable pouches for maximum retention of quality. The stability of the herbal tea was assessed in terms of sensory profile, phenolic composition and color over a storage period of 6 months. Normal storage conditions resulted in a significant (P < 0.05) decrease in green color, especially in steamed samples. Intensities of fruity and sweet-associated aroma attributes increased progressively during storage, while the opposite was observed for vegetal and cereal-like attributes. These changes in the aroma profile were more pronounced in untreated (control) samples. Individual phenolic content remained stable during storage. CONCLUSIONS: Storage of 3 to 6 months may result in a more appealing aroma profile and enhanced product quality, despite loss of green color. © 2018 Society of Chemical Industry.

Bitter Taste Impact and Thermal Conversion of a Naringenin Glycoside from Cyclopia genistoides.

A naringenin derivative, isolated from Cyclopia genistoides, a bitter tasting herbal tea, especially when in green (unoxidized) form, was identified as (2 S)-5-[α-l-rhamnopyranosyl-(1→2)-ß-d-glucopyranosyloxy]naringenin (1). The compound partially epimerizes to (2 R)-5-[α-l-rhamnopyranosyl-(1→2)-ß-d-glucopyranosyloxy]naringenin (2) when heated at different temperatures (80, 90, 100, 110, and 120 °C) for a prolonged period in a phosphate buffer at pH 5. The fractional conversion model predicted the decrease in the concentration of compound 1 the best. The activation energy of the conversion reaction was calculated as 99.16 kJ mol-1. Prolonged heating resulted not only in formation of compound 2 but eventually a decrease in its concentration and the formation of another conversion product, ( E)-2'-[α-l-rhamnopyranosyl-(1→2)-ß-d-glucopyranosyloxy]-4',6',4-trihydroxychalcone (3). In contrast, naringin, glycosylated at C-7, remained stable when heated under the same conditions (100 °C for 6 h at pH 5). The bitter intensity of compound 1 was substantially less than that of naringin, both tested at 0.04 mM, a concentration typical of compound 1 in an herbal tea infusion of green C. genistoides. This comparison indicates that the position of the sugar moiety plays an important role in determining both bitter intensity and heat stability of naringenin glycosides.

Chemoprevention of LA7-Induced Mammary Tumor Growth by SM6Met, a Well-Characterized Cyclopia Extract.

Breast cancer (BC) is the leading cause of cancer-related deaths in women. Chemoprevention of BC by using plant extracts is gaining attention. SM6Met, a well-characterized extract of Cyclopia subternata with reported selective estrogen receptor subtype activity, has shown tumor suppressive effects in a chemically induced BC model in rats, which is known to be estrogen responsive. However, there is no information on the estrogen sensitivity of the relatively new orthotopic model of LA7 cell-induced mammary tumors. In the present study, the potential chemopreventative and side-effect profile of SM6Met on LA7 cell-induced tumor growth was evaluated, as was the effects of 17ß-estradiol and standard-of-care (SOC) endocrine therapies, such as tamoxifen (TAM), letrozole (LET), and fulvestrant (FUL). Tumor growth was observed in the tumor-vehicle control group until day 10 post tumor induction, which declined afterward on days 12-14. SM6Met suppressed tumor growth to the same extent as TAM, while LET, but not FUL, also showed substantial anti-tumor effects. Short-term 17ß-estradiol treatment reduced tumor volume on days prior to day 10, whereas tumor promoting effects were observed during long-term treatment, which was especially evident at later time points. Marked elevation in serum markers of liver injury, which was further supported by histological evaluation, was observed in the vehicle-treated tumor control, TAM, LET, and long-term 17ß-estradiol treatment groups. Alterations in the lipid profiles were also observed in the 17ß-estradiol treatment groups. In contrast, SM6Met did not augment the increase in serum levels of liver injury biomarkers caused by tumor induction and no effect was observed on lipid profiles. In summary, the results from the current study demonstrate the chemopreventative effect of SM6Met on mammary tumor growth, which was comparable to that of TAM, without eliciting the negative side-effects observed with this SOC endocrine therapy. Furthermore, the results of this study also showed some responsiveness of LA7-induced tumors to estrogen and SOC endocrine therapies. Thus, this model may be useful in evaluating potential endocrine therapies for hormone responsive BC.

Phenolic and physicochemical stability of a functional beverage powder mixture during storage: effect of the microencapsulant inulin and food ingredients.

BACKGROUND: The need for a convenience herbal iced tea product with reduced kilojoules merited investigation of the shelf-life of powder mixtures containing a green Cyclopia subternata Vogel (honeybush) extract with proven blood glucose-lowering activity and alternative sweetener mixture. RESULTS: Prior to long-term storage testing, the wettability of powder mixtures containing food ingredients and the compatibility of their components were confirmed using the static sessile drop method and isothermal microcalorimetry, respectively. The powders packed in semi-sealed containers remained stable during storage at 25 °C/60% relative humidity (RH) for 6 months, except for small losses of specific phenolic compounds, namely mangiferin, isomangiferin, 3-ß-d-glucopyranosyliriflophenone, vicenin-2 and 3',5'-di-ß-d-glucopyranosylphloretin, especially when both citric acid and ascorbic acid were present. These acids drastically increased the degradation of phenolic compounds under accelerated storage conditions (40 °C/75% RH). Accelerated storage also caused changes in the appearance of powders and the colour of the reconstituted beverage solutions. Increased moisture content and aw of the powders, as well as moisture released due to dehydration of citric acid monohydrate, contributed to these changes. CONCLUSION: A low-kilojoule honeybush iced tea powder mixture will retain its functional phenolic compounds and physicochemical properties during shelf-life storage at 25 °C for 6 months. © 2017 Society of Chemical Industry.

Polyphenol-Enriched Fractions of Cyclopia intermedia Selectively Affect Lipogenesis and Lipolysis in 3T3-L1 Adipocytes.

Cyclopia species are increasingly investigated as sources of phenolic compounds with potential as therapeutic agents. Recently, we demonstrated that a crude polyphenol-enriched organic fraction (CPEF) of Cyclopia intermedia, currently forming the bulk of commercial production, decreased lipid content in 3T3-L1 adipocytes and inhibited body weight gain in obese db/db mice. The aim of the present study was to determine whether a more effective product and/or one with higher specificity could be obtained by fractionation of the CPEF by purposely increasing xanthone and benzophenone levels. Fractionation of the CPEF using high performance counter-current chromatography (HPCCC) resulted in four fractions (F1-F4), predominantly containing iriflophenone-3-C-ß-D-glucoside-4-O-ß-D-glucoside (benzophenone: F1), hesperidin (flavanone: F2), mangiferin (xanthone: F3), and neoponcirin (flavone: F4), as quantified by high-performance liquid chromatography with diode array detection (HPLC-DAD), and confirmed by LC-DAD with mass spectrometric (MS) and tandem MS (MSE) detection. All fractions inhibited lipid accumulation in 3T3-L1 pre-adipocytes and decreased lipid content in mature 3T3-L1 adipocytes, although their effects were concentration-dependent. F1-F3 stimulated lipolysis in mature adipocytes. Treatment of mature adipocytes with F1 and F2 increased the messenger RNA expression of hormone sensitive lipase, while treatment with F1 and F4 increased uncoupling protein 3 expression. In conclusion, HPCCC resulted in fractions with different phenolic compounds and varying anti-obesity effects. The activities of fractions were lower than the CPEF; thus, fractionation did not enhance activity within a single fraction worthwhile for exploitation as a nutraceutical product, which illustrates the importance of considering synergistic effects in plant extracts.

Thermal stability of the functional ingredients, glucosylated benzophenones and xanthones of honeybush (Cyclopia genistoides), in an aqueous model solution.

Thermal stability of the benzophenones, 3-ß-d-glucopyranosyl-4-ß-d-glucopyranosyloxyiriflophenone (1), 3-ß-d-glucopyranosylmaclurin (2) and 3-ß-d-glucopyranosyliriflophenone (3), and the xanthones, mangiferin (4) and isomangiferin (5), was assessed separately in an aqueous model solution (pH 5) to delineate their major degradation products and mechanism(s). Degradation followed first-order reaction kinetics and the temperature-dependence of the respective reaction rate constants complied with the Arrhenius equation. The stability of the compounds increased in the order 2>4>3>5>1. 4-O-Glucosylation significantly stabilised 1 against degradation compared to 3, enediol B-ring functionality of 2 decreased stability compared to 3 and position of glucosylation affected the stability of the xanthones with 5 being more stable than 4. The xanthone nucleus (C-ring) conferred higher stability to 4 and 5 compared to their benzophenone analogue 2. Cyclisation of 2 to 4 and 5 would underestimate their degradation in mixtures. Other reactions were isomerisation, dimerisation, acetylation and hydrolysis.

Minimising variation in aspalathin content of aqueous green rooibos extract: optimising extraction and identifying critical material attributes.

BACKGROUND: High levels of aspalathin, an antidiabetic dihydrochalcone, in green rooibos underpins interest in the production of a standardised extract. Elements of a quality-by-design approach were applied to optimise extraction conditions, aiming at the delivery of a dry matter yield (DMY) ≥ 160 g kg-1 and an extract with an aspalathin content (AC) ≥ 80 g kg-1 . RESULTS: Hot water extraction parameters, namely extraction time, extraction temperature and water-to-plant material ratio, were optimised for DMY and aspalathin extraction efficiency (AEE) using Design of Experiments. Good polynomial prediction models were obtained and multiresponse desirability plots indicated 37 min, 93 °C and 23:1 as optimal conditions. Even when using 30 min and 10:1 instead for practical reasons, the target DMY and AC values could be achieved with the caveat that plant material with an AC ≥ 30 g kg-1 is used. Particle size distribution and stem content were identified as contributing to variation in the AC of raw material. CONCLUSION: By setting raw material specifications in terms of AC, as well as applying practical optimum extraction conditions, 160 g kg-1 extract with an AC ≥ 80 g kg-1 could be consistently achieved from green rooibos plant material. © 2017 Society of Chemical Industry.