Screening of key odorants and anthocyanin compounds of cv. Okuzgozu (Vitis vinifera L.) red wines with a free run and pressed pomace using GC‐MS‐Olfactometry and LC‐MS‐MS

The principal purpose of the present work is to characterize the aroma, aroma‐active, and anthocyanin profiles of Okuzgozu wines and to observe the effect of the pomace pressing technique on these parameters. A total of 58 and 59 volatile compounds were identified and quantified in free‐run juice wine (FRW) and pressed pomace wine (PW). Alcohols were found as the most dominant group among aroma compounds followed by esters and acids. However, among all these compounds, only 11 and 13 of them could be considered as key odorants in aromatic extracts of FRW and PW, respectively. According to GC‐MS‐O analysis, ethyl octanoate (fruity), phenyl ethyl acetate (fruity), and 2‐phenyl ethanol (flowery) were found as the main contributors to the overall scent of both wines. Beyond the aroma profiles, anthocyanin contents of both types of wines were also investigated, and total 14 and 15 anthocyanins were identified and quantified in FRW and PW. Malvidin‐3‐glycoside and its acetyl and coumaroyl forms were identified as the dominant anthocyanins in both wines. It is worth noting the pressing application (2.0 atm) led to an increase of some unpleasant notes in the aroma providing chemical, pharmacy, and fermented aromas in wine. On the other hand, the wines produced with pressed pomace presented higher amounts of anthocyanins.     

A solid‐phase microextraction gas chromatography/ion trap tandem mass spectrometry method for simultaneous determination of ‘foxy smelling compounds’ and 3‐alkyl‐2‐methoxypyrazines in grape juice

2′‐Aminoacetophenone (o‐AAP) was identified as the main cause of the aging note called ‘hybrid note’ or ‘foxy smell’ which is typical of non‐Vitis vinifera grapes. Together with methyl anthranilate (MA), this compound contributes to the typical foxy taint of wines made with non‐Vitis vinifera grapes. 3‐Alkyl‐2‐methoxypyrazines in grapes, with their herbaceous note and very low sensory thresholds, contribute to the aroma of several wines. In this study, a solid‐phase microextraction gas chromatography/ion trap tandem mass spectrometry (SPME‐GC/IT‐MS/MS) method for simultaneous detection of o‐AAP, MA and ethyl‐, isopropyl‐, sec‐butyl‐ and isobutylmethoxypyrazine in grape juice was developed. The method was applied to the study of several grape juices: the time required for analysis was less than 24 min, and the method was considered to be suitable for analysis of ‘foxy compounds’ and methoxypyrazines in grape juice. The high levels of MA and o‐AAP found in Clinton and Siebel grapes confirmed the ‘hybrid’ character of these varieties. Copyright © 2010 John Wiley & Sons, Ltd.     

A useful approach for the differentiation of wines according to geographical origin based on global volatile patterns

In this study, the feasibility of solid‐phase extraction combined with gas chromatography and mass spectrometry in tandem with partial least squares discriminant analysis was evaluated as a useful strategy to differentiate wines according to geographical origin (Azores, Canary and Madeira Islands) and types (white, red and fortified wine) based on their global volatile patterns. For this purpose, 34 monovarietal wines from these three wine grape‐growing regions were investigated, combining the high throughput extraction efficiency of the solid‐phase extraction procedure with the separation and identification ability. The partial least squares discriminant analysis results suggested that Madeira wines could be clearly discriminated from Azores and Canary wines. Madeira wines are mainly characterized by 2‐ethylhexan‐1‐ol, 3,5,5‐trimethylhexan‐1‐ol, ethyl 2‐methylbutanoate, ethyl dl ‐2‐hydroxycaproate, decanoic acid, 3‐methylbutanoic acid, and (E)‐whiskey lactone, whereas 3‐ethoxypropan‐1‐ol, 1‐octen‐3‐ol, (Z)‐3‐hexenyl butanoate, 4‐(methylthio)‐1‐butanol, ethyl 3‐hydroxybutanoate, isoamyl lactate, 4‐methylphenol, γ‐octalactone and 4‐(methylthio)‐1‐butanol, are mainly associated with Azores and Canary wines. The data obtained in this study revealed that solid‐phase extraction combined with gas chromatography and quadrupole mass spectrometry data and partial least squares discriminant analysis provides a suitable tool to discriminate wines, both in terms of geographical origin as well as wine type and vintage.     

Phenolic Composition and Total Antioxidant Capacity by Electrochemical,Spectrophotometric and HPLC‐EC Evaluation in Portuguese Red and White Wines

Phenolic composition and in vitro total antioxidant capacity have been evaluated, in red and white Portuguese wines, by electrochemical methods, spectrophotometric methods, and reverse‐phase high performance liquid chromatography with electrochemical detection (HPLC‐EC). The different phenolic compounds present in seventeen red and white wines, with different grape varieties, and from different geographical locations, were successfully identified and detected. The wines in vitro total antioxidant capacity, using the method of capture of diphenilpicrilhydrazil (DPPH^.) free radical “efficient concentration” (EC₅₀), and the electrochemical quantitative index (EI), was determined. The wine with the highest total antioxidant capacity exhibited the lowest antioxidant power, and the EI and EC₅₀ assays had a very good correlation.     

Solid phase extraction, multidimensional gas chromatography mass spectrometry determination of four novel aroma powerful ethyl esters. Assessment of their occurrence and importance in wine and other alcoholic beverages

A method for the quantitative determination of four powerful aromatic ethyl esters recently identified in some wines has been developed, validated and applied to the determination of these compounds in different samples of wine, whisky and brandy. Ethyl 2-, 3-, and 4-methylpentanoate and ethyl cyclohexanoate are extracted from 100ml of sample by solid phase extraction (SPE) on a 200mg LiChrolut EN bed. Major compounds are eliminated by rinsing with a water-methanol (50:50) solution containing 1% sodium bicarbonate, and analytes are eluted with 1.5ml of dichloromethane. Fifty microlitres of this extract are then injected in a multidimensional gas chromatography-mass spectromety (GC-GC-MS) system. Recoveries in the SPE are quantitative. Method repeatability is satisfactory (5-12% for a 5-10ngl(-1) level, and less than 7% for 25-50ngl(-1) level), the method linearity holds along the whole range of occurrence of analytes (2-2700ngl(-1)), and the signal is independent on the matrix. Method detection limits are below 1ngl(-1) in all cases. Results suggest that these compounds are formed by the slow esterification with ethanol of the corresponding acids formed by different microorganisms. The levels of these compounds are above the corresponding thresholds in most samples of aged wines or distillates, but are particularly high in some sweet wines, whiskeys and brandies where they may constitute the most important contributors to the sweet-fruity notes reaching concentrations up to 85-350 times higher than the corresponding odor thresholds.     

Volatile flavour constituent patterns of Terras Madeirenses red wines extracted by dynamic headspace solid-phase microextraction

A suitable analytical procedure based on static headspace solid-phase microextraction (SPME) followed by thermal desorption gas chromatography-ion trap mass spectrometry detection (GC-(ITD)MS), was developed and applied for the qualitative and semi-quantitative analysis of volatile components of Portuguese Terras Madeirenses red wines. The headspace SPME method was optimised in terms of fibre coating, extraction time, and extraction temperature. The performance of three commercially available SPME fibres, viz. 100 mum polydimethylsiloxane; 85 mum polyacrylate, PA; and 50/30 mum divinylbenzene/carboxen on polydimethylsiloxane, was evaluated and compared. The highest amounts extracted, in terms of the maximum signal recorded for the total volatile composition, were obtained with a PA coating fibre at 30 degrees C during an extraction time of 60 min with a constant stirring at 750 rpm, after saturation of the sample with NaCl (30%, w/v). More than sixty volatile compounds, belonging to different biosynthetic pathways, have been identified, including fatty acid ethyl esters, higher alcohols, fatty acids, higher alcohol acetates, isoamyl esters, carbonyl compounds, and monoterpenols/C(13)-norisoprenoids.     

Determination of Volatile Compounds in Sultaniye Wine by Solid-Phase Microextraction Techniques

Volatile compounds of wines made from Sultaniye grape varieties grown in Denizli-Turkey were investigated. Volatile compounds in wines were extracted by headspace and immersion solid-phase microextraction (SPME) methods and identified by GC-MS. According to the results of analyses, esters were mainly responsible for the flavor of Sultaniye wine, and among these, ethyl decanoate, ethyl octanoate, ethyl hexanoate, and 3-methylbutyl acetate were found to be dominant. __________ Published in Khimiya Prirodnykh Soedinenii, No. 4, pp. 309–310, July–August, 2005.     

Determination of 3-methyl-2,4-nonanedione in red wines using methanol chemical ionization ion trap mass spectrometry

A compound associated with oxidized flavor in red wines was recently-identified as 3-methyl-2,4-nonanedione (MND). In order to quantify it, positive chemical ionization (PCI) in an ion trap was studied using conventional liquid reagents such as methanol, acetonitrile, and acetone, as well as non-conventional liquid reagents such as ethyl acetate, diethyl ether, pentane, isohexane, and heptane. Under laboratory conditions, very different response factors were obtained with MND depending on the gas. We also compared the detection limit of conventional CI with hybrid chemical ionization (HCI). Finally, this compound was quantified in red wines by liquid/liquid extraction without any derivatization steps, followed by GC/MS-CI analysis, using methanol as the reagent gas. Coelutions of compounds with the same m/z were checked using methanol-d(4). The method we developed was linear in the 10-300 ng/L range of MND concentrations, with satisfactory repeatability. The detection limit was 4.3 ng/L, over 3 times lower than the olfactory perception threshold of this compound (16 ng/L). The suitability of this method for assaying this diketone in red wine was demonstrated by the analyzing many wines from different vintages.     

Determination of flurazepam and its hydroxyethyl and dealkyl metabolites by some packed and capillary ECD-GLC methods: Application to pharmacokinetic studies

A sensitive gas-liquid chromatographic assay was developed for plasma determinations of flurazepam (F), N1-desalkylflurazepam (DF), and N1-hydroxyethylflurazepam (HF). The internal standards used were delorazepam for F and DF and 1 ã-hydroxymidazolam for HF. Following extraction and derivatization for HF, these compounds were analyzed by ECD-GLC, using either glass packed columns (poly A 103 for F and DF, OV 225 for HF) or CP-Sil 5 capillary columns for DF and HF. The detection limit of the packed-column assay was approximately 3ng/ml for HF and DF, and 5ng/ml for F; with capillary GLC analysis, the sensitivity limit for HF and DF was lower than 1 ng/ml. These methods were used for pharmacokinetic investigations after i. v. administration of these compunds.     

Development of a dynamic headspace solid-phase microextraction procedure coupled to GC-qMSD for evaluation the chemical profile in alcoholic beverages

In the present study, a simple and sensitive methodology based on dynamic headspace solid-phase microextraction (HS-SPME) followed by thermal desorption gas chromatography with quadrupole mass detection (GC-qMSD), was developed and optimized for the determination of volatile (VOCs) and semi-volatile (SVOCs) compounds from different alcoholic beverages: wine, beer and whisky. Key experimental factors influencing the equilibrium of the VOCs and SVOCs between the sample and the SPME fibre, as the type of fibre coating, extraction time and temperature, sample stirring and ionic strength, were optimized. The performance of five commercially available SPME fibres was evaluated and compared, namely polydimethylsiloxane (PDMS, 100 μm); polyacrylate (PA, 85 μm); polydimethylsiloxane/divinylbenzene (PDMS/DVB, 65 μm); carboxen™/polydimethylsiloxane (CAR/PDMS, 75 μm) and the divinylbenzene/carboxen on polydimethylsiloxane (DVB/CAR/PDMS, 50/30 μm) (StableFlex).An objective comparison among different alcoholic beverages has been established in terms of qualitative and semi-quantitative differences on volatile and semi-volatile compounds. These compounds belong to several chemical families, including higher alcohols, ethyl esters, fatty acids, higher alcohol acetates, isoamyl esters, carbonyl compounds, furanic compounds, terpenoids, C13-norisoprenoids and volatile phenols. The optimized extraction conditions and GC-qMSD, lead to the successful identification of 44 compounds in white wines, 64 in beers and 104 in whiskys. Some of these compounds were found in all of the examined beverage samples.The main components of the HS-SPME found in white wines were ethyl octanoate (46.9%), ethyl decanoate (30.3%), ethyl 9-decenoate (10.7%), ethyl hexanoate (3.1%), and isoamyl octanoate (2.7%). As for beers, the major compounds were isoamyl alcohol (11.5%), ethyl octanoate (9.1%), isoamyl acetate (8.2%), 2-ethyl-1-hexanol (5.9%), and octanoic acid (5.5%). Ethyl decanoate (58.0%), ethyl octanoate (15.1%), ethyl dodecanoate (13.9%) followed by 3-methyl-1-butanol (1.8%) and isoamyl acetate (1.4%) were found to be the major VOCs in whisky samples.     

Characterization of Aromatic Liquor by Gas Chromatography and Principal Component Analysis

The differentiation of aromas of Chinese liquor is important for their unique flavors. In this work, aromas of Chinese liquor were characterized by gas chromatography and chemometrics. Ten representative aroma compounds, including three alcohols, four esters, two organic acids, and acetal in 16 Chinese liquor were determined by gas chromatography with flame ionization detection. The relationship between these compounds and six classic aromas was investigated using principal component analysis and k-means clustering. The cumulative contribution of the first three principal components reached up to 84.607%, which effectively differentiated the liquors. The variables with the highest loading absolute value were acetal and ethyl acetate for principal component 1, ethyl butanoate and ethyl hexanoate for principal component 2, and the hexanoic acid and ethyl butanoate for principal component 3. The aromas of the liquors were characterized by k-means clustering with the first three principal component scores, indicating that the acetal, ethyl acetate, ethyl butanoate, ethyl hexanoate, and hexanoic acid are important for the aroma of Chinese liquors. This work demonstrated that the gas chromatography with chemometrics is effective for the characterization of aromatic liquor.     

Determination of major compounds in sweet wines by headspace solid-phase microextraction and gas chromatography

Headspace solid-phase microextraction (HS-SPME) was studied by high resolution gas chromatographic analysis of major compounds (ethyl acetate, methanol, 1-butanol, 2-butanol, 1-propanol, isobutanol, 2-methyl-1-butanol and 3-methyl-1-butanol) in sweet wines. Five different SPME fibres were tested and the influence of different factors such as temperature and time of desorption, extraction time, stirring, sample and vial volume, sugar and ethanol content were studied and optimized using model solutions. The SPME method was validated with the direct injection method. The proposed HS-SPME-GC method is an appropriate technique for the quantitative analysis of the mentioned analytes in real sweet wines.     

Classification of wines from five Spanish origin denominations by aromatic compound analysis

Wine is a complex matrix in which aroma compounds play an important role in the characterization of the flavor pattern of a given wine. Twelve volatile compounds were determined in 244 samples of Spanish red wines from different denominations of origin: Rioja, Navarra, Valdepe?as, La Mancha, and Cari?ena. The samples were analyzed by GC using headspace solid-phase microextraction. The concentration (mg/mL) intervals obtained were 3-methyl-butyl acetate (3.9 to 116), 3-methyl-1-butanol (93 to 724), ethyl hexanoate (0.8 to 39), 1-hexanol (0.3 to 6.7), ethyl octanoate (1.4 to 41), diethyl succinate (0.2 to 13), 2-phenyl ethyl acetate (0 to 5.3), hexanoic acid (0 to 8.3), geraniol (0 to 3.0), 2-phenylethanol (1.5 to 56), octanoic acid (0 to 20), and decanoic acid (0 to 3.3). Wines were classified by multivariate statistical methods: principal component analysis, and lineal discriminant analysis. A correct differentiation among wines according to their origin was obtained by lineal discriminant analysis.     

Determination of phenolic compounds in wines by novel matrix solid-phase dispersion extraction and gas chromatography/mass spectrometry

A novel matrix solid-phase dispersion (MSPD) extraction method was developed to extract simultaneously 23 phenolic compounds from wine samples prior to determination by gas chromatography with mass spectrometric detection in the selected ion monitoring mode. Different parameters of the MSPD technique such as dispersant solid-phase, eluting solvent, and sample ionic strength and pH were optimized. The optimized MSPD procedure requires a small volume of wine (1 mL), commercial silica gel (1.5 g) as dispersant solid-phase and a small volume of ethyl acetate (5 mL) as eluting solvent. Under these conditions, the extraction of the studied compounds was almost complete (mean values of recoveries between 87 and 109%) in a short time (15 min). Moreover, satisfactory standard deviations of repeatability (RSD<9% in most cases), linear regression coefficients (r(2)>0.993) and detection limits (<8 microg/L) confirm the usefulness of the methodology for routine monitoring of the concentration of individual phenolic antioxidants in wines. Application was illustrated by analysis of different wine samples.     

Identification of Free and Glycosidically Bound Volatiles and Glycosides by Capillary GC and Capillary GC‐MS in “Lulo del Chocó” (Solanum topiro)

The volatile constituents of lulo del Chocó (Solanum topiro) fruit pulp obtained by liquid‐liquid extraction were analyzed by capillary GC and capillary GC‐MS. In total, 30 components were identified with methyl salicylate, hexadecanoic acid, hexanal, guaiacol, ethyl butanoate, and ethyl acetate being the major components. Chirospecific MDGC analysis revealed the predominance of (R)‐ethyl‐3‐hydroxybutanoate (ee 40%) and the presence of racemic mixtures both of δ‐octalactone and of δ‐decalactone. For γ‐hexalactone, γ‐octalactone, and γ‐decalactone enantiomeric distributions of 22.4 : 77.6, 22.9 : 77.1, and 20.0 : 80.0, (R) : (S), respectively, were determined. Glycosidically bound aroma compounds were identified by capillary GC and capillary GC‐MS after isolation of the glycosidic fraction obtained by Amberlite XAD‐2 adsorption and methanol elution followed by hydrolysis with a commercial pectinase enzyme. In total 13 bound aroma compounds (aglycones) were identified. These aglycones mainly consisted of compounds exhibiting aromatic structures. Additionally, with the aid of capillary GC and capillary GC‐MS (EI and NCI) of trifluoroacetylated derivatives we identified eight glucosides: the novel 3,6‐epoxy‐7‐megastigmen‐5,9‐diol β‐D‐glucopyranoside and the hexyl, benzyl, linalyl oxide (furanic), 2‐phenylethyl, vomifolyl (isomer 1), (6S,9R)‐vomifolyl, and scopoletin β‐D‐glucopyranosides.     

Use of vancomycin chiral stationary phase for the enantiomeric resolution of basic and acidic compounds by nano-liquid chromatography

In this paper we studied the potentiality of nano-liquid chromatography (nano-LC) for the enantiomeric resolution of both basic and acidic compounds of pharmaceutical interest using a vancomycin modified silica stationary phase. Experiments were carried out in a fused silica capillary of 75 microm I.D. packed with chiral modified silica particles of 5 microm diameter, the detection, was done on-line at 195 nm. Enantiomeric resolution of alprenolol, atenolol, metoprolol, oxprenolol, pindolol, propranolol (basic compounds) and some acidic analytes, namely 2-[(5'-benzoyl-2'-hydroxy)phenyl]propionic acid (DF1738Y), 2-[(4'-benzoyloxy-2'-hydroxy)phenyl]propionic acid (DF1770Y), ketoprofen, indoprofen and suprofen was studied by nano-LC utilizing mobile phases containing methanol-acetonitrile-ammonium formate or acetate. The effect of mobile phase composition (buffer type and concentration, organic modifier type and concentration) on chiral resolution (Rs), retention factor (k) and retention time (tR) was also investigated. Good enantiomeric resolution was achieved for basic compounds utilizing the mobile phase containing 90% (MeCN-MeOH)/5% water/5% of 100 mM ammonium acetate pH 4.5. Acidic compounds such as DF1738Y and DF1770Y were better resolved at lower pH 3.5 while ketoprofen, indoprofen and suprofen exhibited the highest resolution at pH 4.5; in this case the mobile phase contained MeOH or MeCN (90%), 5% buffer and 5% of water. The nano-LC method was validated using R-(+)-propranolol as an internal standard finding good repeatability, detection limit, correlation coefficient and recovery and applied to the assay of a pharmaceutical formulation containing a racemic mixture of metoprolol.     

Aroma composition of red wines by different extraction methods and Gas Chromatography-SIM/MASS spectrometry analysis

One hundred and one volatile compounds, reported in literature as powerful odorants of wine, were quantified by Gas Chromatography-Selective Ion Monitoring/Mass Spectrometry (GC-SIM/MS) in Primitivo, Aglianico, Merlot and Cabernet Sauvignon red wines. Wine samples were extracted by 3 different extraction methods: 1) separation of the alcoholic fraction from the aqueous phase by salting-out and subsequent extraction by liquid-liquid micro-extraction with 1,1,2-trichlorotrifluoroethane (Freon 113); 2) extraction by liquid-liquid micro-extraction with dichloromethane; 3) solid phase extraction (SPE cartridge: 800 mg of LiChrolut EN resin) with pentane-dichloromethane (20:1) and dichloromethane. The selection of the ion fragments used for quantification was directly performed on a red wine sample. For each compound the area of the corresponding peak was normalized respect to the peak of the internal standard and then interpolated in a calibration curve obtained analysing a model wine solution (water, ethanol, tartaric acid and known amounts of analytes and of internal standard). The methods showed a good linearity: r2>0.990, except for farnesol (isomer a and c), octanal, decanal, furaneol and phenylacetic acid with 0.966 < or = r2 < or = 0.990. The 7 most powerful odorants were: beta-damascenone, acetaldehyde, maltol, ethyl 2-methylbutanoate, ethyl 3-methylbutanoate, 3-methylbutanoic acid and acetal; 7 other slightly less important were: ethyl hexanoate, ethyl acetate, 1-octen-3-ol, butanoic acid, rose oxide, furaneol and sotolon. The Aglianico wines were characterised by the major fermentation compounds (esters, fatty acids and 2-phenylethanol), beta-damascenone, beta-ionone and linalool. The Primitivo wines were characterized by furaneol, methoxypyrazine, gamma-nonalactone and acetaldehyde, while Cabernet Sauvignon and Merlot wines principally by cask derivates (vanillin, (Z) 3-methyl-gamma-octalactone [(Z) wiskylactone], maltol and eugenol), some aldehydes and 3-isopropyl-2-methoxypyrazine.