Jacques Rigaud

Oligomeric and polymeric procyanidins from grape seeds

Abstract Five tannin fractions were isolated from grape ( Vitis vinifera ) seeds. HPLC analysis of the degradation products released by thioacidolysis showed that all fractions consisted of catechin, epicatechin and epicatechin gallate units. Epicatechin was the major component in the extended chain. Catechin was more abundant in terminal units than in extension units. The proportion of galloyllated units varied from 13% to 29% as the M r , increased. Mean degrees of polymerization ranging from 2.3 to 15.1 (by thiolysis) and from 2.4 to 16.7 (by GPC) were determined.

A new class of wine pigments generated by reaction between pyruvic acid and grape anthocyanins

A new class of stable red pigments detected in grape pomace was analysed by electrospray ionisation mass spectrometry. They were shown to be pyruvic acid derivatives of genuine grape anthocyanins by synthesis experiments. The major product was identified by NMR (1H, NOE, HSQC, HMBC) experiments as the malvidin-3-monoglucoside pyruvic acid adduct. Its formation results from cyclisation between C-4 and the hydroxyl group at C-5 of the original flavylium moiety with the double bond of the enolic form of pyruvic acid, followed by dehydration and rearomatisation steps. This type of reaction leads to increased colour stability. Various yeast metabolites other than pyruvic acid were shown to react with grape anthocyanins following this mechanism, suggesting that it may be an important route of conversion into stable pigments during the maturation and ageing of wine.

Procyanidin dimers and trimers from grape seeds

Abstract Fractionation of a grape (Vitis vinifera) seed extract yielded various natural procyanidin dimers and trimers isolated and identified for the first time in grapes: procyanidin dimers B5 [epicatechin-(4β → 6)-epicatechin], B6 [catechin-(4α → 6)-catechin] and B8 [catechin-(4α → 6)-epicatechin]; four procyanidin trimers [epicatechin-(4β → 6)-epicatechin-(4β → 8)-epicatechin], [epicatechin-(4β → 8)-epicatechin-(4β → 6)-catechin], [epicatechin-(4β → 8)-epicatechin-(4β → 6)-epicatechin], [epicatechin-(4β → 6)-epicatechin-(4β → 8)-catechin] along with five galloyl procyanidins: B2 3-O-gallate, B1 3-O-gallate, B4 3′-O-gallate, B2 3,3′-di-O-gallate and [epicatechin-(4β → 8)-epicatechin 3-O-gallate-(4β → 8)-catechin]. The presence of (+)-catechin, (−)-epicatechin, (−)-epicatechin 3-O-gallate, procyanidins dimers B1, B2, B3, B4, B7, B2 3′-O-gallate and procyanidin trimers C1 and [epicatechin-(4β → 8)-epicatechin-(4β → 8)-catechin] was also confirmed. The structures of all these compounds were elucidated by enzymatic hydrolysis, complete acid hydrolysis, partial acid-catalysed degradation with phloroglucinol and phenylmethanethiol, FABMS and 1H NMR. Separation of procyanidins and of their phloroglucinol or phenylmethanethiol adducts was achieved by TLC on silica plates and HPLC.

Normal-phase high-performance liquid chromatographic separation of procyanidins from cacao beans and grape seeds

Abstract An HPLC method using a normal-phase silica column and a gradient of dichloromethane—methanol—formic acid—water mixtures as the eluent was developed to separate procyanidins on a molecular mass basis, without derivatization. It was successfully applied to the analysis of procyanidin extracts from cacao beans and grape seeds. The monomers and major dimers were resolved as discrete peaks. Oligomeric and polymeric components were eluted in order of increasing degree of polymerization, as confirmed by determining the average molecular mass of successive fractions collected from the normal-phase column using gel permeation chromatography, after acetylation.

Micro method for the identification of proanthocyanidin using thiolysis monitored by high-performance liquid chromatography

Abstract The classical method of procyanidin thiolysis was modified so as to render it suitable for the identification of small amount (

[15] Size separation of condensed tannins by normal-phase high-performance liquid chromatography

Publisher Summary This chapter discusses the size separation of condensed tannins by normal-phase high performance liquid chromatography (HPLC). Condensed tannins are also called “proanthocyanidins” because they release anthocyanins when heated in acidic conditions. They are ubiquitous plant components, consisting of chains of flavan-3-ol units. Tannin properties—including radical scavenging effects protein binding ability—depend largely on their structure and particularly on the number of constitutive units. Several methods have been developed to determine oligomeric and polymeric proanthocyanidins. Separation of lower molecular weight proanthocyanidins is usually achieved by reversed-phase HPLC, but the elution order is unrelated to the degree of polymerization. Various other techniques have been used to separate proanthocyanidins following this criterion—for instance, liquid chromatography on sephadex LH-204 or fractogel. HPLC method presented in this chapter allows separation of proanthocyanidins on a molecular weight basis, without derivatization. Although this method is suitable in most cases, it may require modifications, depending on the nature of tannins to be extracted and on the presence of other plant constituents. In particular, the extraction yield may be improved by sonicating and the samples may be protected against oxidation at this stage—for instance, by sulfiting or working under inert gas and/or at low temperature.

Oxidation kinetics of trans-caffeoyltartrate and its glutathione derivatives in grape musts

Abstract The oxidation of trans -caffeoyl tartrate and the formation of cafteoyl tartrate o -quinone, 2- S -glutathionyl caffeoyl tartrate (grape reaction product, GRP), GRP o -quinone, and 2,5-di- S -glutathionyl cafteoyl tartrate (GRP2) in model solutions containing caffeoyl tartrate, glutathione, and crude grape enzymatic extract were monitored by HPLC and shown to depend on the caffeoyl tartrate to glutathione molar ratio. Similar kinetics were obtained in oxidizing grape musts. However, it seems that other grape components can trap the o -quinones in the same way as glutathione. An initial lag phase, due to the presence of powerful reductants such as ascorbic acid, was observed.

Structure of Procyanidin Oligomers Isolated From Grape Seeds in Relation to Some of their Chemical Properties

A micro-method using thiolysis followed by HPLC monitoring of the degradation products was developed to determine the structure of procyanidin oligomers available in small quantities (≤ 0.1 mg). It was successfully applied to a series of procyanidins isolated from grape seeds, allowing the identification of procyanidins dimers B5 to B8 linked by C4 → C6 bonds, six procyanidin trimers with C4 → C8 and C4 → C6 linkages, and six galloylated dimers and trimers, along with the four C4 → C8 linked dimers, Bl to B4. The influence of procyanidin structural variations on their interactions with proteins, free radical scavenging effect, and oxidation was studied using these compounds. Protein-procyanidin interactions increased with the degree of polymerization, the number of galloyl substituents, and the extent of C4→ C6 linkages. Galloylation increased scavenger capacity of procyanidin dimers for Superoxide anion (O2 -•) and hydroxyl (su•OH) radicals. The type of C → C linkage and the esterification position were also important. None of the molecules tested was oxidizable by grape polyphenoloxidase, but they were all oxi-dized by the enzymically generated caffeoyl tartaric acid o-quinones. In addition, galloylated procyanidins seemed to undergo condensation reactions faster than the corresponding non-galloylated ones, even though their rate of oxidation was similar.