Urinary peptidomics and pulse wave velocity: The African-PREDICT study

Increased arterial stiffness is related to early vascular aging and is an independent predictor for cardiovascular disease and mortality. Molecular mechanisms underlying increased arterial stiffness are largely unexplored, especially at the proteome level. We aimed to explore the relationship between pulse wave velocity and urinary proteomics. We included 919 apparently healthy (no chronic illnesses) Black and White men and women (equally distributed) between 20 and 30 years from the African-PREDICT study. Capillary electrophoresis time-of-flight mass spectrometry was used to analyze the urinary proteome. We measured the carotid-femoral pulse wave velocity to estimate arterial stiffness. In the total group, pulse wave velocity correlated positively with collagen-derived peptides including collagen types I, II, III, IV, V, and IX and inversely with collagen type XI (adjusted for mean arterial pressure). Regarding noncollagen-derived peptides, pulse wave velocity positively correlated with polymeric immunoglobulin receptor peptides (n = 2) (all q-value ≤0.05). In multivariable adjusted analyses, pulse wave velocity associated positively and independently with seven urinary peptides (collagen type I, n = 5) (all p-value ≤0.05). We found significant positive and independent associations between pulse wave velocity and the collagen type I-derived peptides, suggesting that dysregulation of collagen type I in the extracellular matrix scaffold could lead to early onset of increased arterial stiffness

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

CITATION: Viraragavan, A. et al. 2020. Model development for predicting in vitro bio-capacity of green rooibos extract based on composition for application as screening tool in quality control. Food & Function, 11:3084-3094. doi:10.1039/C9FO02480HThe original publication is available at http://pubs.rsc.org/en/Journals/Journal/FOMounting 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.Publishers versio

A urinary peptidomics approach for early stages of cardiovascular disease risk: the African-PREDICT study

Cardiovascular disease (CVD) affects individuals across the lifespan, with multiple cardiovascular (CV) risk factors increasingly present in young populations. The underlying mechanisms in early cardiovascular disease development are complex and still poorly understood. We therefore employed urinary proteomics as a novel approach to gain better insight into early CVD-related molecular pathways based on a CVD risk stratification approach. This study included 964 apparently healthy (no self-reported chronic illnesses, free from clinical symptoms of CVD) black and white men and women (aged 20–30 years old) from the African Prospective study on the Early Detection and Identification of Cardiovascular disease and Hypertension (African-PREDICT) study. Cardiovascular risk factors used for stratification included obesity, physical inactivity, tobacco use, high alcohol intake, hyperglycemia, dyslipidemia and hypertension. Participants were divided into low (0 risk factors), medium (1–2 risk factors) and high (≥3 risk factors) CV risk groups. We analyzed urinary peptidomics by capillary electrophoresis time-of-flight mass spectrometry. After adjusting for ethnicity, sex and age, 65 sequenced urinary peptides were differentially expressed between the CV risk groups (all q-values ≤ 0.01). These peptides included a lower abundance of collagen type I- and III-derived peptides in the high compared to the low CV risk group. With regard to noncollagen peptides, we found a lower abundance of alpha-1-antitrypsin fragments in the high compared to the low CV risk group (all q-values ≤ 0.01). Our findings indicate lower abundances of collagen types I and III in the high compared to the low CV risk group, suggesting potential early alterations in the CV extracellular matrix

Identifying a urinary peptidomics profile for hypertension in young adults: the African-PREDICT study

Hypertension is one of the most important and complex risk factors for cardiovascular diseases (CVDs). By using urinary peptidomics analyses, we aimed to identify peptides associated with hypertension, building a framework for future research towards improved prediction and prevention of premature development of CVD. We included 78 hypertensive and 79 normotensive participants from the African-PREDICT study (aged 20–30 years), matched for sex (51% male) and ethnicity (49% black and 51% white). Urinary peptidomics data were acquired using capillary-electrophoresis-time-of-flight-mass-spectrometry. Hypertension-associated peptides were identified and combined into a support vector machine-based multidimensional classifier. When comparing the peptide data between the normotensive and hypertensive groups, 129 peptides were nominally differentially abundant (Wilcoxon p &lt; 0.05). Nonetheless, only three peptides, all derived from collagen alpha-1(III), remained significantly different after rigorous adjustments for multiple comparisons. The 37 most significant peptides (all p ≤ 0.001) served as basis for the development of a classifier, with 20 peptides being combined into a unifying score, resulting in an AUC of 0.85 in the ROC analysis (p &lt; 0.001), with 83% sensitivity at 80% specificity. Our study suggests potential value of urinary peptides in the classification of hypertension, which could enable earlier diagnosis and better understanding of the pathophysiology of hypertension and premature cardiovascular disease development

Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

Biochemical detection (BCD) methods are commonly used to screen plant extracts for specific biological activities in batch assays. Traditionally, bioactives in the most active extracts were identified through time-consuming bio-assay guided fractionation until single active compounds could be isolated. Not only are isolation procedures often tedious, but they could also lead to artifact formation. On-line coupling of BCD assays to high performance liquid chromatography (HPLC) is gaining ground as a high resolution screening technique to overcome problems associated with pre-isolation by measuring the effects of compounds post-column directly after separation. To date, several on-line HPLC-BCD assays, applied to whole plant extracts and mixtures, have been published. In this review the focus will fall on enzyme-based, receptor-based and antioxidant assays

The antioxidant activity of South African wines in different test systems as affected by cultivar and ageing

Thesis (M. Sc.Voedselwet.)--University of Stellenbosch, 2001.ENGLISH ABSTRACT: Phenolic compounds in wine, due to their antioxidant activity, are reportedly responsible for the health-promoting properties of wines. The effect of cultivar and in-bottle ageing on the antioxidant activity of South African wines in different types of antioxidant assays was, therefore, investigated. The antioxidant activity of commercial South African red (Cabernet Sauvignon, Ruby Cabernet, Pinotage, Shiraz, Merlot) and white (Sauvignon blanc, Chenin blanc, Chardonnay, Colombard) cultivar wines was compared using the 2,2’-azino-di-(3-ethylbenzothialozine-sulphonic acid) radical cation (ABTS·+) scavenging, 2,2-diphenyl-1-picrylhydrazyl radical (DPPH·) scavenging and microsomal lipid peroxidation (MLP) assays. The red wines was more effective than the white wines on an “as-is” and an equal total phenol content. The total antioxidant activity (TAAABTS and TAADPPH) of Ruby Cabernet was the lowest of the red wines, but the antioxidant potency (APABTS and APDPPH) of red wine phenolic fractions did not differ (P ³ 0.05). Ruby Cabernet and Pinotage were the least effective inhibitors of MLP, while Merlot was the most effective of the red wines. Pinotage phenolic fractions had lower (P < 0.05) APMLP than that of other red wines. Of the white wines, Chardonnay and Chenin blanc had the highest and lowest effectivity respectively according to all antioxidant parameters. Ascorbic acid present in some wines increased and decreased their TAA and % MLP inhibition respectively. TAA and % MLP inhibition correlated well (r ³ 0.7, P < 0.001) with total phenol content of red and white wines, as well as with flavanol content of red wines and tartaric acid ester content of white wines. The % MLP inhibition also correlated well with flavanol content of white wines. No correlation (P > 0.01) was obtained between TAA or % MLP inhibition and monomeric anthocyanin content of red wines. In the deoxyribose assay, red wines were more pro-oxidant and exhibited lower hydroxyl radical scavenging and metal chelating abilities than white wines. The effect of in-bottle ageing on antioxidant activity of wines was determined using the ABTS·+ and DPPH· scavenging assays. The TAA and total phenol content of experimental red (Pinotage and Cabernet Sauvignon)and white (Chardonnay and Chenin blanc) cultivar wines, decreased (P < 0.05) during 12 months of storage at 0, 15 and 30 ºC. The TAAABTS of Cabernet Sauvignon and Chardonnay, stored at 30 ºC were lower (P < 0.05) than at 0 ºC. The APABTS and APDPPH of most wines also decreased during storage. The monomeric anthocyanin content of red wines decreased (P < 0.05) rapidly at 15 and 30 ºC. The flavanol content of wines (except Chenin blanc) increased during the first 9 months, decreasing again after 12 months, while minor changes in the flavonol and tartaric acid ester content of both red and white wines were observed. The TAAABTS exhibited a good correlation (r ³ 0.7, P < 0.001) with total phenol content of red and white wines, as well as with flavonol and tartaric acid ester content of red and white wines and flavanol content of white wines. The monomeric anthocyanin content of red wines correlated (r = 0.50, P < 0.001) weakly with TAAABTS. The decrease in the TAAABTS of wines could thus be mainly attributed to a decrease in their total phenol content.AFRIKAANSE OPSOMMING: Die antioksidant aktiwiteit van fenoliese komponente in wyn is waarskynlik verantwoordelik vir die gesondheidsvoordele daarvan. Die studie het dus gepoog om effek van kultivar en veroudering na bottelering op die antioksidant aktiwiteit van Suid-Afrikaanse wyne te ondersoek. Die antioksidant aktiwiteit van kommersiële Suid-Afrikaanse rooi (Cabernet Sauvignon, Ruby Cabernet, Pinotage, Shiraz, Merlot) en wit (Sauvignon blanc, Chenin blanc, Chardonnay, Colombard) kultivarwyne is vergelyk deur middel van die 2,2’-azino-di-(3-etielbensotialosien-sulfoon suur)-radikaal katioon (ABTS·+) vernietigingstoets, 2,2-difeniel-1-pikrielhidrasielradikaal (DPPH·) vernietigingstoets en mikrosomale lipiedperoksidasietoets (MLP). Die antioksidant aktiwiteit en die antioksidant kragtigheid (AK) van die rooiwyne was beter as dié van witwyne in al drie antioksidant toetse. Die totale antioksidant aktiwiteit (TAAABTS en TAADPPH) van Ruby Cabernet was die laagste van die rooiwyne, terwyl die AKABTS en AKDPPH van rooiwyn fenoliese fraksies nie van mekaar verskil (P ³ 0.05) het nie. Van die rooiwyne, het Ruby Cabernet en Pinotage die laagste en Merlot die hoogste effektiwiteit in die MLP toets getoon. Die AKMLP van Pinotage se fenoliese fraksies was die laagste van die rooiwyne. Die witwyne, Chardonnay en Chenin blanc, het onderskeidelik die beste en swakste antioksidant aktiwiteit en AK van die witwyne getoon in al drie antioksidant toetse. Askorbiensuur wat in sommige witwyne voorgekom het, het die TAA van hierdie wyne verhoog, maar hul % MLP inhibisie verlaag. Die TAA en % MLP inhibisie het goed gekorreleer (r ³ 0.7, P < 0.001) met die totale fenolinhoud van rooi- en witwyne, asook die flavanolinhoud van rooiwyne en die wynsteensuur-esterinhoud van witwyne. Die % MLP inhibisie het ook goed gekorreleer met die flavanolinhoud van witwyne. Geen korrelasie (P > 0.1) is waargeneem tussen antioksidant aktiwiteit van rooiwyne en hul monomeriese antosianien-inhoud. Rooiwyn was meer pro-oksidatief in die deoksieribose toets as witwyne, maar was die swakste hidroksieradikaalvernietigers en metaalcheleerders.Die effek van veroudering na bottelering op die antioksidant aktiwiteit van wyne soos bepaal met die ABTS·+ en DPPH· vernietigingstoetse, is ondersoek. Die TAA en die totale fenolinhoud van eksperimentele rooi- (Pinotage en Cabernet Sauvignon) en witwyne (Chardonnay en Chenin blanc) het afgeneem (P < 0.05) tydens opberging na bottelering by 0, 15 en 30 ºC oor 12 maande. Opberging by 30 ºC het ‘n groter vermindering (P < 0.05) in die TAAABTS waarde vir Cabernet Sauvignon en Chardonnay veroorsaak as by 0 ºC. Die meeste wyne se APABTS en APDPPH waardes het ook verminder (P < 0.05) na 12 maande. Drastiese vermindering (P < 0.05) in die monomeriese antosianieninhoud van rooiwyne is opgemerk tydens opberging by 15 en 30 ºC. Tydens die eerste 9 maande se opberging het die flavanolinhoud van wyne toegeneem (P < 0.05) en daarna afgeneem (P < 0.05) tot by 12 maande, terwyl flavonol- en wynsteensuuresterinhoud van beide rooi- en witwyne min verandering ondergaan het. Die totale fenolinhoud van rooi- en witwyne, asook die flavonol en wynsteensuur-esterinhoud van rooi-en witwyne en die flavanolinhoud van witwyne, het goed gekorreleer (r ³ 0.7, P < 0.001) met die TAAABTS. In teenstelling met die resultate vir kommersiële kultivarwyne, was die TAAABTS van rooiwyne swak gekorreleer (r = 0.5, P < 0.001) met hul monomeriese antosianieninhoud. Die afname in TAAABTS van wyne tydens veroudering kon dus meestal toegeskryf word aan die afname in hul totale fenolinhoud.Maste

Food Ingredient Extracts of Cyclopia subternata (Honeybush): Variation in Phenolic Composition and Antioxidant Capacity

Cyclopia subternata plants are traditionally used for the production of the South African herbal tea, honeybush, and recently as aqueous extracts for the food industry. A C. subternata aqueous extract and mangiferin (a major constituent) are known to have anti-diabetic properties. Variation in phenolic composition and antioxidant capacity is expected due to cultivation largely from seedlings, having implications for extract standardization and quality control. Aqueous extracts from 64 seedlings of the same age, cultivated under the same environmental conditions, were analyzed for individual compound content, total polyphenol (TP) content and total antioxidant capacity (TAC) in a number of assays. An HPLC method was developed and validated to allow quantification of xanthones (mangiferin, isomangiferin), flavanones (hesperidin, eriocitrin), a flavone (scolymoside), a benzophenone (iriflophenone-3-C-β-glucoside) and dihydrochalcones (phloretin-3\u27,5\u27-di-C-β-glucoside, 3-hydroxyphloretin-3\u27,5\u27-di-C-hexoside). Additional compounds were tentatively identified using mass spectrometric detection, with the presence of the 3-hydroxyphloretin-glycoside, an iriflophenone-di-O,C-hexoside, an eriodictyol-di-C-hexoside and vicenin-2 being demonstrated for the first time. Variability of the individual phenolic compound contents was generally higher than that of the TP content and TAC values. Among the phenolic compounds, scolymoside, hesperidin and iriflophenone-3-C-β-glucoside contents were the most variable. A combination of the measured parameters could be useful in product standardization by providing a basis for specifying minimum levels

Antioxidant capacity of Pinotage wine as affected by viticultural and enological practices

Thesis (PhD (Food Science))--University of Stellenbosch, 2006.The aim of the study was to provide the South African wine industry with guidelines for the production of Pinotage wines with optimal total antioxidant capacity (TAC), while retaining sensory quality. The contribution of individual phenolic compounds to the wine TAC is important in this regard. The wine TAC was measured with the 2,2 -azino-di(3-ethylbenzo-thiazoline-sulphonic acid radical cation) (ABTS +) scavenging assay. The contributions of individual phenolic compounds to the wine TAC were calculated from their content in the wines and the Trolox equivalent antioxidant capacity (TEAC) of pure phenolic standards. The effects of climate region, vine structure, enological techniques (pre-fermentation maceration, juice/skin mixing, addition of commercial tannins, extended maceration) and maturation (oak barrels, alternative oak products, oxygenation) on the phenolic composition, TAC and sensory quality of Pinotage wines were also investigated. The TEAC values of quercetin-3-galactoside, isorhamnetin and peonidin-3-glucoside were reported for the first time. TEAC values observed for most compounds were much lower than those reported previously, although TEAC values for gallic acid, caftaric acid, caffeic acid and kaempferol were consistent with some previous reports. Caftaric acid and malvidin-3-glucoside were the largest contributors to the wine TAC. The contents of monomeric phenolic compounds and procyanidin B1, however, only explained a small amount (between 11 and 24%) of the wine TAC, with the remaining TAC attributed to oligomeric and polymeric phenolic compounds and other unknown compounds. Some synergy between different monomeric phenolic compounds was also demonstrated. All the viticultural and enological factors investigated affected the phenolic composition of Pinotage wines, while the wine TAC was only affected by some treatments. Changes in wine TAC could not always be explained by changes in phenolic composition as the contribution of oligomeric, polymeric and unknown compounds could not be assessed, but could play a large role. Differences in wine colour were also difficult to explain due to the large number of factors involved and the dark wine colour, which made objective measurements difficult. The concentration of vitisin A, an orange-red pyranoanthocyanin, was increased consistently as a result of prefermentation maceration treatments and affected the wine colour of oxygenated wines. Increased wine TAC was observed when cultivating Pinotage grapes on bush vines and in cooler climatic regions, compared to cultivation on trellised vines in warmer climatic regions. All the climatic regions and vine structure treatments, however, resulted in wines with good sensory quality. In terms of enological techniques, pumping-over, as opposed to punching-down and rotor treatments, is not recommended as a juice/skin mixing technique, due to reduced wine TAC, colour and sensory quality. Pre-fermentation maceration, addition of commercial tannin preparations, and oak maturation using traditional and alternative treatments, resulted in improved sensory quality, but with no change in wine TAC. However, optimisation of the tannin addition protocol may result in increased wine TAC if additions are made after fermentation or higher dosages are used. Oxygenation of Pinotage wine needs further investigation to optimise the protocol, as improvements to the wine colour and fullness were observed for some treatments, but loss of sensory quality and TAC were observed in most cases