Raymond L. Baumes

The aroma of grapes. I. Extraction and determination of free and glycosidically bound fractions of some grape aroma components

Abstract A method of extraction and determination of free and glycosidically bound terpenols and of benzyl and 2-phenylethyl alcohols is suggested. The extraction was carried out by adsorbing these compounds on a non-ionic resin, Amberlite XAD-2, and then elution with various selective solvents. Free forms were directly determined by gas chromatography; glycosidically bound forms were first enzymatically hydrolysed to release aglycones. This method was applied to a number of mature grape varieties. The levels of glycosidically bound terpenols and aromatic alcohols were always high, usually much higher than the free forms. The varieties can be classified in two groups: those rich in free and bounds forms which give aromatic wines (Muscat varieties and Gewurztraminer), and those which contain small amounts of these compounds and which give more neutral wines.

Analytical methods for monoterpene glycosides in grape and wine. II. Qualitative and quantitative determination of monoterpene glycosides in grape.

Free and glycosidically bound terpenes of five Vitis vinifera grape cultivars (muscat of Alexandria, muscat of Frontignan, muscat of Hamburg, muscat Ottonel and Gewürztraminer) were investigated. The free and bound fractions were separated by selective retention on Amberlite XAD-2 resin. The glycosidic fractions were analysed by gas chromatography and gas chromatography-mass spectrometry using either enzymic hydrolysis and subsequent analysis of the released aglycones or trimethylsilyl (TMS) and trifluoroacetyl derivatives. The known monoterpenyl, benzyl and 2-phenylethyl beta-D-glucopyranosides, beta-rutinosides, 6-O-alpha-L-arabinofuranosyl-beta-D-glucopyranosides and 6-O-beta-D-apiofuranosyl-beta-D-glucopyranosides were determined. A number of other glycosides were detected and the structures of some of them, mainly apiosylglucosides and glucosides with aglycones in higher oxidation state than linalol, were tentatively identified using the mass spectra of their TMS and TFA derivatives and the results obtained from the analysis of their aglycones.

Monoterpenic and norisoprenoidic glycoconjugates of Vitis vinifera L. cv. Melon B. as precursors of odorants in Muscadet wines.

The volatile monoterpenic and norisoprenoidic compounds released by glycosidase enzyme hydrolysis of C18 reversed-phase isolates from the juice of Vitis vinifera L. cv. Melon B. have been qualitatively and quantitatively determined using GC-MS and GC-FID. The components analyzed were broadly similar to those previously reported for other varieties but the level of bound p-menth-1-en-7,8-diol was higher in this cultivar. Then the monoterpenic and norisoprenoidic volatiles released from the same glycosidic extracts under mild acid conditions, mimicking wine aging conditions, have been analyzed using GC-Olfactometry and GC-MS. The most odorous compounds detected were p-cymene, terpinen-4-ol, cis- and trans-vitispiranes, 1,6,6-trimethyl-1,2-dihydronaphtalene (TDN), beta-damascenone and riesling acetal. To assess their potential levels in corresponding wines after ageing, most of these odorants were generated by harsh acid hydrolysis from the precursors extracts and quantitatively determined using SPME and GC-MS/MS. For the development and application of this analysis, the odorants not commercialy available were synthesized. The total amounts of norisoprenoidic odorants generated by acid hydrolysis of the glycosidic extracts were shown to be proportional to the total amounts of these precursors.

Quantitative determination of free and hydrolytically liberated β-damascenone in red grapes and wines using a stable isotope dilution assay

Quantitative analysis of free and hydrolytically liberated beta-damascenone in grapes and wines was developed, using a stable isotope dilution assay. Free beta-damascenone was isolated from grapes and wines by diethyl ether-hexane (1:1, v/v) extraction and the precursor(s) (glycosidic, polyols) of beta-damascenone using Sep-Pak Plus C18 RP cartridges. Hydrolytically liberated beta-damascenone was generated by acid hydrolysis from the precursor(s) extract. Red wines from Bordeaux (Merlot, Cabernet Sauvignon and Cabernet Franc, 1995 and 1996 vintage), Burgundy (Pinot Noir, 1995 and 1996 vintage) regions and Grenache wines from Chateauneuf du Pape and Côtes du Rhône (1995 vintage) were analysed to quantify free beta-damascenone. The wines made from Grenache and Cabernet Sauvignon (1996 vintage) grapes presented the highest mean amounts of free beta-damascenone, 5.4 and 5.5 micrograms l-1, respectively. Merlot, Cabernet Sauvignon and Cabernet Franc grapes of Bordeaux (1996 vintage) and their corresponding wines were analysed for quantification of free and hydrolytically liberated beta-damascenone. The levels of hydrolytically liberated beta-damascenone in grapes could predict closely the levels of free beta-damascenone in the corresponding wines after one year of ageing, i.e., almost half the levels found for the grape samples. The influence of enzyme and heat treatment of Merlot wine samples on their beta-damascenone levels was studied. Heat treatment doubled the levels of this compound, but enzyme treatment generated, in the corresponding wines, half the levels of beta-damascenone found in the non-enzyme treated wines.

Analytical methods for monoterpene glycosides in grape and wine: I. XAD-2 extraction and gas chromatographic—mass spectrometric determination of synthetic glycosides☆

Synthetic monoterpene and aromatic beta-D-glucopyranosides, beta-rutinosides and 6-O-(alpha-L-arabinofuranosyl)-beta-D-glucopyranosides and their corresponding alcohols, diluted in a synthetic solution imitating wine, were isolated and separated by selective retention on Amberlite XAD-2. The corresponding recoveries and the conditions for the direct determination of these glycosides by gas chromatography and gas chromatography-mass spectrometry after derivatization were determined. Trifluoroacetylation gave the best results but trimethylsilylation provided complementary results. The separation of some diastereoisomeric monoterpene glycosides was also examined.

Quantitative determination of β-ionone in red wines and grapes of Bordeaux using a stable isotope dilution assay

Quantitative analysis of beta-ionone has been developed, using a stable isotope dilution assay. This was applied to red wines from different cultivars and regions. The Burgundy Pinot noir wines exhibited the highest levels of beta-ionone. The variation in the levels of beta-ionone in grape samples and in their corresponding wines, of Merlot, Cabernet Sauvignon and Cabernet franc from Bordeaux regions was monitored at four different stages towards the end of maturation: the levels of beta-ionone were almost similar, exhibiting only a slight decrease during maturation. beta-Ionone occurred in all the grapes and wines samples analysed at levels higher than, or close to its odour threshold which was determined in a model wine solution and was found to be of 90 ng l-1. In a previously supplemented model wine solution with beta-ionone (250 ng l-1), the odour threshold was found to be of 980 ng l-1 (total levels).

Enzymatic synthesis of monoterpenyl β-D-glucosides by various β-glucosidases

Abstract β-glucosidases from Aspergillus niger, Trichoderma reesei, Candida molischiana , and almond have been shown to catalyze synthesis of β-glucosides of primary monoterpene alcohols, such as geraniol, nerol, and citronellol, using cellobiose as carbohydrate donor. Enzymes had the strongest glucosyl transferase activity for geraniol. Glucosylation of (±)-citronellol was nonselective. Among organic solvents tested, 30% acetone in enzyme assay gave the highest yield in glucoside synthesis. Monoterpene alcohols, such as linalool, α-terpineol, and menthol, were not used as acceptors in transglycosylation reactions.

Quantitative determination of 2-methoxy-3-isobutylpyrazine in red wines and grapes of Bordeaux using a stable isotope dilution assay.

Quantitative analysis of 2-methoxy-3-isobutylpyrazine (MIBP) in grapes and wines was developed, using a stable isotope dilution assay. This was applied to red grapes and wines from the Bordeaux region. The grapes and the wines of the 1995 and 1996 vintages came from the three most frequently used varieties of the region, Merlot, Cabernet Franc and Cabernet Sauvignon. The wines made from Cabernet Sauvignon grapes exhibited levels of MIBP (mean concentration, 12 ng l-1 for 1996 vintage and 13 ng l-1 for 1995 vintage) close to or higher than its odour threshold in wines (10 ng l-1) and slightly higher than the amounts found in the Merlot wines (mean concentration, 8 ng l-1 for 1996 vintage and 4 ng l-1 for 1995 vintage), especially those of the 1996 vintage. The variation in the levels of MIBP in grape samples and in their corresponding wines was monitored at four different stages towards the end of maturation. MIBP was present in all grapes and wines analysed, even in surmaturation. A linear trend was observed between grapes and wines of the three cultivars during maturation.

Volatile and Glycosidically Bound Composition of Loureiro and Alvarinho Wines

Composition of Loureiro and Alvarinho wines from the Vinhos Verdes region, regarding free volatile compounds as well as glycosidically bound aroma precursors, was exhaustively determined by gas chromatography—mass spectrometry after adsorption on XAD-2 resin. On the whole, were identified and quantified 120 volatile compounds in the free fraction and 77 glycosidically bound compounds, belonging to C6-compounds, alcohols, fatty acids ethyl esters, esters of organic acids, acetates, monoterpenic alcohols, monoterpenic oxides and diols, C13-norisoprenoids, volatile phenols, volatile fatty acids, and carbonyl compounds. Globally, the wines of the two cultivars present similar composition on volatiles. However, regarding varietal compounds, Loureiro wines were richer than Alvarinho ones with respect to C6-compounds and monoterpenic compounds, occurring the opposite for volatile phenols. It was also demonstrated that wines of both varieties might benefit the aroma reserve, present as glycoconjugates, as it is susceptible of being technologically explored. Linalool, Ho-trienol, (α-terpineol, contributing with fruity and floral notes, and (β-damascenone mostly for Alvarinho, confering tropical fruit notes, are the varietal compounds which may particularly influence the aroma of these wines. Respecting fermentative compounds, Alvarinho is also particularly rich in fatty acids ethyl esters related to lipid metabolism and acetates of fusel alcohols, which can provide it a fruity character; Loureiro contains higher levels of esters of organic acids and 2-phenylethanol, conferring fruity and floral notes. Sensory analysis agreed with chemical analyses showing a pronounced tree and tropical fruit character for Alvarinho wines while Loureiro wines present more intense citrus fruit notes.

Identification of S-methylmethionine in Petit Manseng grapes as dimethyl sulphide precursor in wine.

A procedure for the extraction of free amino acids was applied to isolate S-methylmethionine (SMM) from late harvest Petit Manseng grapes. Grapes were destemmed and crushed, and the obtained clarified must was percolated through cation-exchange resins (Dowex 50 WX4-100). The retained compounds were eluted with ammonia solution and the extract was finally concentrated. Taking into account the potential DMS (PDMS using heat-alkaline treatment assay) of the initial grape juice used (51.5nmolmL(-1)) and the concentration factor of the extract (17.9-fold), the PDMS of the final extract (678nmolmL(-1)) gave an overall recovery of 73.5% for juice SMM. This compound was identified and quantified (484.5nmolmL(-1) relatively to [(2)H(3)]-SMM used as internal standard) by its selective detection in this extract without derivatization by MALDI-TOF-MS using instrumentation and procedures previously reported to analyze SMM in complex natural extracts. SMM and 22 other amino acids in the initial must and in the final SMM extract were also determined using a Biochrom 30 amino acid analyser with post-column ninhydrin derivatization. SMM peak identification and quantification (401.2nmolmL(-1) relatively to norleucine used as internal standard) were carried out by comparison with commercial SMM.