Development and validation of an UHPLC-HRMS protocol for the analysis of flavan-3-ol metabolites and catabolites in urine, plasma and feces of rats fed a red wine proanthocyanidin extract (original) (raw)
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Foods
Wine phenolic compounds, particularly proanthocyanidins (PAs), play a significant role in wine sensory characteristics, specifically bitterness and astringency. Although not consensual, flavan-3-ols and oligomeric PAs are generally considered the primary contributors to wine bitterness. Patatin, a vegetable protein fining agent, has been explored as an alternative to animal and synthetic fining agents for reducing wine bitterness. However, contradictory results exist regarding its effectiveness in removing flavan-3-ols and oligomeric PAs in red wines. In this work, a UPLC–Q-TOF MS/MS method was optimized and validated for accurately measuring flavan-3-ols, as well as dimeric and trimeric PAs, in red wines. The MS/MS analysis of flavan-3-ols, in addition to the typical fragmentation described in the literature, revealed an intense mass fragment resulting from the loss of C3O2 and C3O2 + H2O from the parent ion. It was observed that flavan-3-ols and PAs undergo oxidation during sample...
British Journal of Nutrition, 2005
Rats were fed ag rape seed extract (GSE) containing ( þ )-catechin,( 2 )-epicatechin and dimers, trimers, tetramers and polymeric procyanidins. Liver, kidney, brain and gastrointestinal (GI) tract together with plasma, urine and faeces were collected over a24h period and their flavan-3-ol content was analysed by HPLC with tandem mass spectrometry and diode array detection. Small amounts of the GSE flavan-3-ols moved out of the stomach and into the duodenum/jejunum, and to ag reater extent the ileum 1h after ingestion, and into the caecum after 2h with relatively small amounts being detected in the colon after 3h.T he GI tract contained the parent GSE flavan-3-ols and procyanidins with only trace amounts of metabolites and there were no indications that proanthocyanidins were depolymerised in the GI tract releasing monomeric flavan-3-ols.Plasma contained exclusively catechin glucuronidesand methylated glucuronide metabolites which were also detected in the liver and kidneys. These metabolites were also present in urine together with sulphated metabolites and low amounts of the procyanidin dimers B 1 ,B 2 ,B 3 and B 4 as well as the trimer C 2 and an unknown GSE trimer. The amounts of ( þ )-catechin and ( 2 )-epicatechin metabolitesexcreted in urine relative to the quantity of the monomers ingested were 27 and 36 %, respectively, after 24 h. This is similar to the levels of urinary excretion reported to occur by other investigators after feeding ( 2 )-epicatechin to rats and provides further, albeit indirect, evidence that the procyanidinoligomers in the GSE were not depolymerised to monomers to any extent after ingestion. No convincing analytical data were obtained for the presence of flavan-3-ol metabolites in the brain. Flavan-3-ols: Procyanidins: Absorption: Metabolism:E xcretion: Rats: HPLC-MS-MS * Corresponding author: Dr P.-L. Teissedre, fax +33 (0) 467548686, emailp teissed@univ-montp1.fr †Authors made an equal contribution.
Identification and Quantitation of Flavanols and Proanthocyanidins in Foods: How Good are the Datas?
Clinical and Developmental Immunology, 2005
Evidence suggesting that dietary polyphenols, flavanols, and proanthocyanidins in particular offer significant cardiovascular health benefits is rapidly increasing. Accordingly, reliable and accurate methods are needed to provide qualitative and quantitative food composition data necessary for high quality epidemiological and clinical research. Measurements for flavonoids and proanthocyanidins have employed a range of analytical techniques, with various colorimetric assays still being popular for estimating total polyphenolic content in foods and other biological samples despite advances made with more sophisticated analyses. More crudely, estimations of polyphenol content as well as antioxidant activity are also reported with values relating to radical scavenging activity. High-performance liquid chromatography (HPLC) is the method of choice for quantitative analysis of individual polyphenols such as flavanols and proanthocyanidins. Qualitative information regarding proanthocyanidi...
Prodelphinidins and related flavanols in wine
International Journal of Food Science & Technology, 2000
Liquid chromatography coupled to diode array spectrophotometry and mass spectroscopy detection (LC-DAD-MS) has been applied to the study of the proanthocyanidin composition of red wine. Analytical conditions were optimized using several flavanols previously isolated from grape seeds and pomegranate peel. Higher sensitivity was obtained in positive than in negative ion mode and electrospray ionization (ESI) proved to be more useful than chemical ionization (APCI) for the identification of flavanols with a higher degree of polymerization, since it always provided the molecular ion. Useful information for the identification of compounds was also obtained from their fragmentation patterns (MS-MS spectra). The application of the method of LC-DAD-MS set up permitted the identification of fourteen flavanols in wine: four monomers (C, catechin; EC, epicatechin; GC, gallocatechin; EGC, epigallocatechin), eight dimers (B1, B2, B3, B4, B7, and two trimers . Two other heterogeneous dimers were detected, which were supposed to possess an epi(gallo)catechin unit. No important peaks corresponding to galloyl derivatives were found. As far as we know, prodelphinidins and flavanols containing GC units were identified in wine for the first time.
Journal of Agricultural and Food Chemistry, 2014
This study describes the development and validation of a liquid chromatography−mass spectrometry method for determination of a large number of flavanol colonic derivatives in biological samples. The method was validated with rat plasma after the intake of grape seed flavanols. The minimum plasma volume necessary to maintain good recovery values within the range of 83−110% for all of the standards was determined by micro solid-phase extraction (μ-SPE). In total, 16 commercial standards were used to measure 30 different phenolic compounds present at low concentration levels (micromolar). The chromatographic method enabled reliable quantification of plasma colonic flavanol derivatives with low limits of detection and quantification, achieving values of 0.03 nM and 0.10 nM, respectively. The developed method can be readily applied to determine all of the flavanol metabolites that are most likely responsible for the majority of biological effects of poorly absorbed flavanols.
Fractionation of Grape Seed Proanthocyanidins for Bioactivity Assessment
Grape seed extract (GSE) is rich in flavonoids, particularly proanthocyanidins (PAs). PAs and their monomers are strong antioxidants and have been ascribed a number of potential activities beneficial to health, including protection against cancers, cardiovascular disease, and Alzheimer’s disease. However, little is known about the chemical nature and the concentration of bioactive polyphenols to which the body is exposed after ingestion of GSE. This chapter summarizes recent advances in fractionation techniques for PAs, including work from our laboratory on fractionation of GSE in quantities sufficient for analysis of its protective effects in whole animal models of Alzheimer’s disease. Several techniques, ranging from fractionation using liquid–liquid extraction to separation on Toyopearl resin and normal-phase HPLC, have been optimized to separate and purify monomeric, oligomeric, and polymeric PAs from GSE. The extraction and separation efficiencies of the various techniques are discussed.
Journal of Mass Spectrometry, 2003
The proanthocyanidins in three foods (pinto beans, plums and cinnamon) were studied with electrospray ionization (ESI) mass spectrometry (MS) in the negative mode following separation by normal-phase high-performance liquid chromatography. The MS/MS analysis demonstrated that the major ions derived from heterocyclic ring fission and retro-Diels-Alder reaction of flavan-3-ol provided information about the hydroxylation pattern and type of interflavan bond. The connection sequence of the oligomers was identified through diagnostic ions derived from quinone methide (QM) cleavage of the interflavan bond. Novel heterogeneous B-type proanthocyanidins containing (epi)afzelechin as subunits were identified in pinto beans. Proanthocyanidins with interestingly different A-type linkages were identified in plums and cinnamon. In efforts aimed at extending the identification capacity of ESI-MS to polymers, we found that the polymeric procyanidins fragmented readily instead of forming multiply charged ions in the negative ESI mode. Fragmentation patterns were proposed based on our data obtained by ESI-MS/MS and ESI time-of-flight MS.
Journal of Chromatography A, 1994
A method for characterizing flavan-3-01s and procyanidins by HPLC separation in connection with on-line UV detection and chemical reaction detection (CRD) is described. The post-column derivatization of the polyphenols with 4-dimethylaminocinnamaldehyde in the presence of sulphuric acid improves the sensitivity as compared to UV detection. The extent of improvement strongly depends on the structure of the procyanidin, hence resulting in characteristic absorbance ratios (CRDNJV) for the individual compounds under given reaction conditions. The ratios are further influenced by the solvent composition, by reaction time and by temperature.
Improving the Phloroglucinolysis Protocol and Characterization of Sagrantino Wines Proanthocyanidins
Molecules
Proanthocyanidins are key metabolites that explain wine sensorial character (bitterness and astringency) and red wine color changes during aging. Therefore, a fast and accurate method to evaluate the degree of polymerization and the structural composition of the polymeric proanthocyanidins is a crucial analytical tool. Phloroglucinolysis is the most used method for this analysis but, unfortunately, the phloroglucinol adducts of the monomeric flavan-3-ols are not commercially available, making the results less accurate. The aim of this work was the isolation by semi-preparative high performance liquid chromatography (HPLC) of these non-commercial compounds and their use for the development of an accurate UHPLC-MS/MS protocol. The purity of each adduct was established via quantitative 1H-nuclear magnetic resonance (NMR) measurements with 3-trimethylsilyl-propionic-d4 acid sodium salt as the calibration standard. The developed method was applied to evaluate the proanthocyanidins profil...
2007
Representing the most common flavonoid consumed in the American diet, the flavanols and their polymeric condensation products, the proanthocyanidins, are regarded as functional ingredients in various beverages, whole and processed foods, herbal remedies and supplements. Their presence in food affects food quality parameters including astringency, bitterness, sourness, sweetness, salivary viscosity, aroma, and color formation. The ability of flavonoids to aid food functionality has also been established in terms of microbial stability, foamability, oxidative stability, and heat stability. polymerize to form the flavonoid subclass proanthocyanidins (Figure 1.3)(Marais, Deavours et al. 2006). The proanthocyanidins. The use of various terms to describe polymeric flavan-3-ols has made understanding proanthocyanidin nomenclature a bit of a daunting task. Of the several terms used in literature (tannins, condensed tannins, vegetable tannins, flavans, flavolans, polyflavans, catechins, macromolecular phenolic substances, leucoanthocyanidins, condensed proanthocyanidins, polymeric proanthocyanidins, oligomeric proanthocyanidins, procyanidins, procyanidolic oligomers, plant polyphenols, and pycnogenols), the terms 'proanthocyanidins' and 'condensed tannins' are used most frequently (Chung, Wong et al. 1998; Haslam 1998; Khanbabaee and VanRee 2001; Ferreira, Marais et al. 2003). The term 'tannin' is also commonly employed, however its use should be carefully considered as it can refer to other classes of plant polyphenols such as the hydrolysable and complex tannins (Khanbabaee and VanRee 2001; Ferreira, Marais et al. 2003). The IUPAC system also works for naming proanthocyanidins. Although its use becomes troublesome in naming larger molecules, the system has more recently been adapted and applied to define absolute stereochemistry of flavonoids at all chiral centers. As most flavan-3-ols are 2R isomers, the prefix 'ent' is used in describing 2S isomers. Location and stereochemistry about the interflavanoid bond are denoted by the symbols α and β. Terms ending in-in possess a glycone, while those ending inidin lack a glycone (Ribereau-Gayon 1972). Proanthocyanidins can be further classified into two groups based on subunit A ring hydroxylation: the phloroglucinol type (meta-trihydroxylated) or resorcinol type