Quantitative plasma profiling by 1H NMR-based metabolomics: impact of sample treatment

Introduction: There is evidence that sample treatment of blood-based biosamples may affect integral signals in nuclear magnetic resonance-based metabolomics. The presence of macromolecules in plasma/serum samples makes investigating low-molecular-weight metabolites challenging. It is particularly relevant in the targeted approach, in which absolute concentrations of selected metabolites are often quantified based on the area of integral signals. Since there are a few treatments of plasma/serum samples for quantitative analysis without a universally accepted method, this topic remains of interest for future research.Methods: In this work, targeted metabolomic profiling of 43 metabolites was performed on pooled plasma to compare four methodologies consisting of Carr-Purcell-Meiboom-Gill (CPMG) editing, ultrafiltration, protein precipitation with methanol, and glycerophospholipid solid-phase extraction (g-SPE) for phospholipid removal; prior to NMR metabolomics analysis. The effect of the sample treatments on the metabolite concentrations was evaluated using a permutation test of multiclass and pairwise Fisher scores.Results: Results showed that methanol precipitation and ultrafiltration had a higher number of metabolites with coefficient of variation (CV) values above 20%. G-SPE and CPMG editing demonstrated better precision for most of the metabolites analyzed. However, differential quantification performance between procedures were metabolite-dependent. For example, pairwise comparisons showed that methanol precipitation and CPMG editing were suitable for quantifying citrate, while g-SPE showed better results for 2-hydroxybutyrate and tryptophan.Discussion: There are alterations in the absolute concentration of various metabolites that are dependent on the procedure. Considering these alterations is essential before proceeding with the quantification of treatment-sensitive metabolites in biological samples for improving biomarker discovery and biological interpretations. The study demonstrated that g-SPE and CPMG editing are effective methods for removing proteins and phospholipids from plasma samples for quantitative NMR analysis of metabolites. However, careful consideration should be given to the specific metabolites of interest and their susceptibility to the sample treatment procedures. These findings contribute to the development of optimized sample preparation protocols for metabolomics studies using NMR spectroscopy

Descriptive analysis of dietary (poly)phenol intake in the subcohort MAX from DCH-NG: "Diet, Cancer and Health-Next Generations cohort"

Purpose (Poly)phenols are bioactive compounds widely distributed in plant-based foods. Currently, limited data exist on the intake distribution of (poly)phenols across meals. This study aimed to estimate dietary intakes of all individual (poly)phenols and total intake per class and subclass by meal event, and to identify their main food sources in the subcohort MAX from the Diet, Cancer and Health-Next Generations cohort (DCH-NG). Methods Dietary data were collected using three web-based 24-h dietary recalls over 1 year. In total, 676 participants completed at least one recall. The dietary data were linked to Phenol-Explorer database using standardized procedures and an in-house software. We categorized foods/drinks into five options of meal events selected by the participant: \u27Breakfast\u27, \u27Lunch\u27, \u27Evening\u27, \u27Snack\u27, and \u27Drink\u27. Results Adjusted total (poly)phenols mean intake by meal was the highest in the drink event (563 mg/day in men and 423 mg/day in women) and the lowest in the evening event (146 mg/day in men and 137 mg/day in women). The main overall (poly)phenol class contributor was phenolic acids (55.7-79.0%), except for evening and snack events where it was flavonoids (45.5-60%). The most consumed (poly)phenol subclasses were hydroxycinnamic acids and proanthocyanidins. Nonalcoholic beverages (coffee accounted for 66.4%), cocoa products, and cereals were the main food sources of total (poly)phenols. Conclusion This study provides data on the variability in the intake of classes and subclasses of (poly)phenols and their main food sources by meal event according to lifestyle data, age, and gender in a Danish population

Discrimination of beer flavours by analysis of volatiles using the mass spectrometer as an electronic nose

Entire mass spectra of beer headspace components were used as fingerprints for beer brand classification and differentiation of beer samples stored under various conditions. Chemometric analysis of the mass spectra allowed for the discrimination of beer brands and for the detection of beer aging and photodegradation. The numeric methods used include unsupervised PCA modelling and discrimination using kappa NN, LDA, and D-PLS methods

Clinical phenotype clustering in cardiovascular risk patients for the identification of responsive metabotypes after red wine polyphenol intake

© 2015 Elsevier Inc. This study aims to evaluate the robustness of clinical and metabolic phenotyping through, for the first time, the identification of differential responsiveness to dietary strategies in the improvement of cardiometabolic risk conditions. Clinical phenotyping of 57 volunteers with cardiovascular risk factors was achieved using k-means cluster analysis based on 69 biochemical and anthropometric parameters. Cluster validation based on Dunn and Figure of Merit analysis for internal coherence and external homogeneity were employed. k-Means produced four clusters with particular clinical profiles. Differences on urine metabolomic profiles among clinical phenotypes were explored and validated by multivariate orthogonal signal correction partial least-squares discriminant analysis (OSC-PLS-DA) models. OSC-PLS-DA of 1H-NMR data revealed that model comparingPostprint (published version

Comparison of flavonoid intake assessment methods using USDA and phenol explorer databases: Subcohort diet, cancer and health-next generations—MAX study

Flavonoids are bioactive plant compounds that are widely present in the human diet. Estimating flavonoid intake with a high degree of certainty is challenging due to the inherent limitations of dietary questionnaires and food composition databases. This study aimed to evaluate the degree of reliability among flavonoid intakes estimated using four different approaches based on the two most comprehensive flavonoid databases, namely, United States Department of Agriculture (USDA) and Phenol Explorer (PE). In 678 individuals from the MAX study, a subcohort of the Diet, Cancer and Health-Next Generations cohort, dietary data were collected using three 24-h diet recalls over 1 year. Estimates of flavonoid intake were compared using flavonoid food content from PE as (1) aglycones (chromatography with hydrolysis), (2) aglycones transformed (converted from glycosides by chromatography without hydrolysis), (3) as they are in nature (glycosides, aglycones, and esters), and 4) using flavonoid content from USDA as aglycones (converted). Spearman\u27s intra-class correlation (ICC) coefficient and weighted kappa (K) coefficient were calculated for the reliability analysis. When comparing PE total aglycones to USDA total aglycones, there was a moderate reliability when a continuous variable was used [ICC: 0.73, 95% confidence interval (CI): 0.70–0.76] and an excellent reliability when flavonoid intake was modeled as a categorical variable (K: 0.89, 95% CI: 0.88–0.90). The degree of reliability among all methods of estimated flavonoid intakes was very similar, especially between database pairs, for the flavanol subclass, while larger differences were observed for flavone, flavonol, and isoflavone subclasses. Our findings indicate that caution should be taken when comparing the results of the associations between flavonoid intakes and health outcomes from studies, when flavonoid intakes were estimated using different methods, particularly for some subclasses

HPLC–Tandem Mass Spectrometric Method to Characterize Resveratrol Metabolism in Humans

AbstractBackground: Nutritional biomarkers are alternatives to traditional dietary assessment tools. We sought to develop a method for nutritional analysis of resveratrol, a phenolic compound with purported health-promoting properties, and to determine all resveratrol metabolites.Methods: We obtained LDL and urine samples from 11 healthy male volunteers who had consumed 250 mL of Merlot red wine. We measured resveratrol and its metabolites with 96-well solid-phase extraction plates coupled with HPLC-tandem mass spectrometry. Hexestrol was used as the internal standard. Gradient chromatography in multiple reaction monitoring mode was performed on a Luna C18 column, maintained at 40 °C; m/z transitions were as follows: resveratrol, 227/185; resveratrol glucosides, 389/227; resveratrol glucuronides, 403/227; resveratrol sulfates, 307/227; taxifolin, 303/285; and hexestrol, 269/134.Results: Standard calibration curves were linear at 4.4–3289.5 nmol/L. Residual analyses were 100% (3.2) for trans-resveratrol and 100% (11.1) for trans-piceid. In both matrices, imprecision (CV) was <10.8% at all concentrations. Detection limits for resveratrol were 0.2 nmol/L (LDL), 0.3 nmol/L (synthetic urine), and 4.0 nmol/L (blank urine). Resveratrol and metabolites were checked for stability, and no degradation was observed.Conclusions: The HPLC–tandem mass spectrometry method enabled us to identify resveratrol sulfates in human LDL and to characterize the complete profile of resveratrol metabolism in human LDL and urine. This method provides an accurate index of exposure to resveratrol and its metabolites, which can be used as nutritional biomarkers for evaluating the biological effects of moderate wine intake on human health

Dietary polyphenols, metabolic syndrome and cardiometabolic risk factors: An observational study based on the DCH-NG subcohort

Background and aims: Polyphenol-rich foods have beneficial properties that may lower cardiometabolic risk. We aimed to prospectively investigate the relationship between intakes of dietary polyphenols, and metabolic syndrome (MetS) and its components, in 676 Danish residents from the MAX study, a subcohort of the Danish Diet, Cancer and Health–Next Generations (DCH-NG) cohort. Methods and results: Dietary data were collected using web-based 24-h dietary recalls over one year (at baseline, and at 6 and 12 months). The Phenol-Explorer database was used to estimate dietary polyphenol intake. Clinical variables were also collected at the same time point. Generalized linear mixed models were used to investigate relationships between polyphenol intake and MetS. Participants had a mean age of 43.9y, a mean total polyphenol intake of 1368 mg/day, and 75 (11.6%) had MetS at baseline. Compared to individuals with MetS in Q1 and after adjusting for age, sex, lifestyle and dietary confounders, those in Q4 – for total polyphenols, flavonoids and phenolic acids–had a 50% [OR (95% CI): 0.50 (0.27, 0.91)], 51% [0.49 (0.26, 0.91)] and 45% [0.55 (0.30, 1.00)] lower odds of MetS, respectively. Higher total polyphenols, flavonoids and phenolic acids intakes as continuous variable were associated with lower risk for elevated systolic blood pressure (SBP) and low high-density lipoprotein cholesterol (HDL-c) (p < 0.05). Conclusions: Total polyphenol, flavonoid and phenolic acid intakes were associated with lower odds of MetS. These intakes were also consistently and significantly associated with a lower risk for higher SBP and lower HDL-c concentrations

Iberian cured-ham consumption improves endothelial function in healthy subjects

Objectives: Previous studies have shown that dietary components such as oleic acid or polyphenols exert beneficial effects on endothelium. We aimed to assess the impact of regular consumption of Iberian cured-ham (ICH) on endothelial function. Design: An open-label, randomized controlled parallel study. Setting: Volunteers recruited through advertisements at a hospital in Madrid, Spain. Participants: 102 Caucasian adults (76.8% females) aged 25-55 years, and free from cardiometabolic disease. Intervention: Participants were randomized to an ICH-enriched ad libitum diet or an ad libitum diet without ICH for 6 weeks. Subjects in ICH group were randomly provided with either acorn- or mixed-fed ICH, and followed up for an additional 6-week period under their usual diet. Measurements: Clinical parameters, biomarkers of endothelial function and oxidative stress, microvascular vasodilatory response to hyperemia and arterial stiffness were measured before and after the intervention. Results: After 6 weeks, a larger decrease in PAI-1 was observed in subjects consuming ICH compared to the Control group (-6.2±17.7 vs. 0.3±1.4 ng/ml; p=0.020). Similarly, microvascular vasodilatory response to hyperemia showed a significant increase (112.4±391.7 vs. -56.0±327.9%; p=0.007). However, neither oxidative stress, hemodynamic nor clinical parameters differed significantly over the study. Additionally, after stopping ICH consumption, improvements in PAI-1 remained for 6 additional weeks with respect to baseline (p=0.006). Conclusion: The present study demonstrates, for the first time, that regular consumption of ICH improves endothelial function in healthy adults. Strategies aimed to preserve or improve the endothelial function may have implications in vascular aging beyond the prevention of the atherothrombotic disease