Optimisation of a dispersive liquid-liquid microextraction method for the simultaneous determination of halophenols and haloanisoles in wines

A dispersive liquid-liquid microextraction (DLLME) method has been optimised for simultaneously extracting 2,4,6-trichloranisole (TCA), 2,3,4,6-tetrachloroanisole (TeCA), 2,4,6-tribromoanisole (TBA), pentachloroanisole (PCA), 2,4,6-trichlorophenol (TCP), 2,3,4,6-tetrachlorophenol (TeCP), 2,4,6-tribromophenol (TBP) and pentachlorophenol (PCP) from wine. The haloanisoles and halophenols were automatically determined using a gas chromatography-electron-capture detection (GC-ECD) system. Derivatisation of halophenols was performed at the same time as DLLME. Firstly, disperser and extraction solvents, salt addition and temperature conditions were selected. Then, the volume of disperser solvent, extraction solvent and derivatisation agent, and the percentage of base were optimised by means of a central composite design combined with desirability functions. The optimal extraction-derivatisation conditions found were 1.3 mL of acetone, 150 μL of carbon tetrachloride, 75 μL of acetic anhydride and a percentage of base of 0.7%; with no salt addition and at room temperature. Under these conditions, the proposed method showed satisfactory linearity (with correlation coefficients over 0.994), repeatability (below 9.7%) and reproducibility (below 9.9%). Moreover, detection limits were lower than the olfactory threshold of the compounds. The developed method was successfully applied to the analysis of red wine samples. To our knowledge, this is the first time that DLLME has been applied to determine cork taint responsible compounds in wine.     

Non-destructive method to determine halophenols and haloanisoles in cork stoppers by headspace sorptive extraction

The new solvent-free technique called headspace sorptive extraction (HSSE) was used to determine 2,4,6-trichlorophenol, 2,3,4,6-tetrachlorophenol, 2,4,6-trichloroanisole, 2,3,4,6-tetrachloroanisole, 2,4,6-tribromoanisole, pentachloroanisole in cork stoppers, without grinding them, as these may be responsible for the cork taint off-flavour in wine. The best HSSE sorption kinetics for the target analytes were obtained after submitting the spiked corks to 100 degrees C for 1h, followed by a 30 min stabilization time at room temperature. The stir bar was desorbed in a thermal desorption system coupled to a gas chromatograph-mass spectrometer. The method proposed showed good linearity over the concentration range tested, 1-70 ngg(-1), and correlation coefficients ranged from 0.90 to 0.99 for all the analytes. The reproducibility and repeatability of the method were estimated between 4.91 and 12.67%. The effect of the different cork matrixes on the extraction recovery of the target compounds was studied, with the natural corks showing the higher recovery percentage in relation to agglomerate ones.     

Quantification of chloroanisoles in cork using headspace solid-phase microextraction and gas chromatography with electron capture detection

Chloroanisoles can migrate from the cork stopper in wine bottles to the wine and give it a musty taint so it is important to find a method by which they can be determined. The aim of this paper is to develop a method for quantifying 2,4-dichloroanisole, 2,6-dichloroanisole, 2,4,6-trichloroanisole, 2,3,4,6-tetrachloroanisole and 2,3,4,5,6-pentachloroanisole in cork using headspace solid-phase microextraction and gas chromatography with electron capture detection. After we had prepared the cork standards that were so essential to the work we optimised the parameters that most influence headspace solid-phase microextraction: fibre coating, vial volume, cork, kind and volume of solvent to help the extraction, extraction temperature and time, ionic strength and stirring. The method quantifies the total amount of chloroanisoles in cork stoppers (natural, agglomerated, agglomerated with disks and sparkling wine stoppers), at suitable concentrations so that the capacity of these compounds to give wine a musty taint can be evaluated. The quantification limits are: 2,6-dichloroanisole (8.6 ng/g), 2,4,6-trichloroanisole (0.8 ng/g), 2,4-dichloroanisole (3.5 ng/g), 2,3,4,6-tetrachloroanisole (0.6 ng/g), 2,3,4,5,6-pentachloroanisole (0.8 ng/g). The other quality parameters are: recoveries (90.3-105.8%), repeatability (4-13% (RSD expressed)) and intermediate precision (5-14% (RSD expressed)).     

Determination of odour-causing volatile organic compounds in cork stoppers by multiple headspace solid-phase microextraction

Multiple headspace solid-phase microextraction (MHS-SPME) coupled with gas chromatography-mass spectrometry has been applied in order to determine 2,4,6-trichloroanisole (2,4,6-TCA), guaiacol, 1-octen-3-ol and 1-octen-3-one in three samples of cork stoppers. These compounds are responsible for cork taint in wine and can modify the organoleptic properties of bottled wine. Variables such as temperature, addition of water, extraction time, and amount of cork were studied. The extractions were performed with a 50/30 microm divinylbenzene-carboxen-polydimethylsiloxane (DVB-CAR-PDMS) fibre for 45 min at 100 degrees C using 20 mg of cork. For calibration, 50 microL of VOC aqueous solutions were used and the extraction were carried out for 45 min at 75 degrees C. The limits of detection of the method expressed as ng of VOC per g of cork were 0.3 for 2,4,6-TCA, 7.5 for guaiacol, 1.7 for 1-octen-3-one and 1.9 for 1-octen-3-ol. Relative standard deviation of replicate samples was less than 10%. Significant losses of analytes were observed when the samples were ground at room temperature. Finally, a recovery study was performed and the MHS-SPME results were validated using Soxhlet extraction results.     

Optimisation of a microwave-assisted extraction method for the simultaneous determination of haloanisoles and halophenols in cork stoppers

This study presents a method based on the use of microwave-assisted extraction (MAE) for the quantitative analysis of 2,4,6-trichloroanisole (TCA), 2,3,4,6-tetrachloroanisole (TeCA), pentachloroanisole (PCA), 2,4,6-tribromoanisole (TBA), 2,4,6-trichlorophenol (TCP), 2,3,4,6-tetrachlorophenol (TeCP), pentachlorophenol (PCP) and 2,4,6-tribromophenol (TBP) in cork stoppers. The influential parameters of the MAE procedure (extraction time, temperature and solvent volume) were optimised using a central composite experimental design combined with desirability functions. The optimal conditions identified were temperature 170 degrees C, solvent volume 35 mL and extraction time 90 min. MAE extracts were concentrated and derivatised prior to separation and quantification by gas chromatography with electron capture detection. To evaluate the applicability of the proposed MAE method, recovery results were compared with those obtained with the Soxhlet extraction method; the results were similar with both extraction methods. The new method was also satisfactorily applied to real cork stopper samples.     

Development of a dispersive liquid-liquid microextraction method for the simultaneous determination of the main compounds causing cork taint and Brett character in wines using gas chromatography-tandem mass spectrometry

A novel dispersive liquid-liquid microextraction (DLLME) method, coupled to gas chromatography-tandem mass spectrometry (GC-MS/MS), was developed for simultaneously determining the main compounds responsible for cork taint (2,4,6-trichloranisole (TCA), 2,3,4,6-tetrachloroanisole (TeCA), 2,4,6-tribromoanisole (TBA) and pentachloranisole (PCA)) and Brett character (4-ethylguaiacol (EG), 4-ethylphenol (EP), 4-vinylguaiacol (VG) and 4-vinylphenol (VP)) in wines. Optimisation of DLLME procedure was performed by evaluating the type of disperser and extraction solvents and the temperature and salt addition effects. The volumes of disperser and extraction solvents were also optimised by means of a central composite design combined with desirability functions. Under optimum conditions, 5 mL of wine were extracted with an extraction mixture consisting of 1.43 mL of acetone, and 173 μL of chloroform at room temperature. The analytical characteristics of the method were evaluated. Satisfactory linearity (with correlation coefficients over 0.992), repeatability (below 11.6%) and between-days precision (below 11.0%) were obtained for all target analytes. Detection limits attained were at similar levels or even lower than the olfactory threshold of the studied compounds. Finally, the developed method was successfully applied to the analysis of wine samples. To our knowledge, this is the first time that DLLME has been applied to simultaneously determine the compounds responsible for cork taint and Brett character in wine.     

Fast and sensitive method to determine chloroanisoles in cork using an internally cooled solid-phase microextraction fiber

A new generation of solid-phase microextraction (SPME) fiber, an internally cooled fiber (cold fiber with polydimethylsiloxane loading) that allows heating the sample matrix and simultaneously cooling the fiber coating, was used to determine 2,4-dichloroanisole, 2,6-dichloroanisole, 2,4,6-trichloroanisole and pentachloroanisole in cork. A comparison between the cold fiber and regular SPME fiber was performed. An automated headspace solid-phase microextraction (HS-SPME) using commercial fibers and an internally cooled SPME fiber (CF-HS-SPME) coupled to gas chromatography-time-of-flight mass spectrometry (GC-TOF-MS) was used. The extraction conditions for both CF-HS-SPME and HS-SPME were optimized using full factorial design and Doehlert matrix. The best extraction conditions for CF-HS-SPME were obtained using 10 min of incubation time, 10 min of extraction time, and sample and fiber temperature of 130 and 10 degrees C, respectively. For HS-SPME, polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber was used with 10 min of incubation time, 75 min of extraction time, 85 degrees C of sample temperature, 8 ml of water was added and agitated at 500 rpm. The quantification limits for the target compounds using CF-HS-SPME procedure were between 0.8 and 1.6 ng g(-1) of cork, while for HS-SPME were between 4 and 6 ng g(-1) of cork. Furthermore, the CF-HS-SPME procedure could be used as a non-destructive method after minor modification of the agitator for the autosampler.     

Analysis for chloroanisoles and chlorophenols in cork by stir bar sorptive extraction and gas chromatography–mass spectrometry

A complete methodology for the determination of chloroanisoles and chlorophenols in cork material is proposed. The determination is accomplished by means of a previous liquid–solid extraction followed by stir bar sorptive extraction (SBSE) coupled to gas chromatography–mass spectrometry (GC–MS). Two different liquid–solid extraction experiments were conducted and eight compounds considered (2,6-dichloroanisole, 2,4-dichloroanisole, 2,4,6-trichloroanisole, 2,4,6-trichlorophenol, 2,3,4,6-tetrachloroanisole, 2,3,4,6-tetrachlorophenol, pentachloroanisole and pentachlorophenol). From the results obtained we can conclude that high volume extraction extending extraction time up to 24 h is the best choice if we have to release compounds from the inner surfaces of cork stoppers. Recovery percentages ranged from 51% for pentachloroanisole to 81% for 2,4-dichloroanisole. This method allows the determination of an array of compounds involved in cork taint at very low levels from 1.2 ng g⁻¹ for 2,4,6-tricholoroanisole to 23.03 ng g⁻¹ for 2,3,4,6-tetrachlorophenol.     

Multiple headspace solid-phase microextraction for eliminating matrix effect in the simultaneous determination of haloanisoles and volatile phenols in wines

This paper proposes a multiple headspace solid-phase microextraction (MHS-SPME) method coupled to gas chromatography-tandem mass spectrometry detection (GC/MS/MS) for the simultaneous determination of 2,4,6-trichloroanisole, 2,3,4,6-tetrachloroanisole, pentachloroanisole, 2,4,6-tribromoanisole, 4-ethylphenol, 4-ethylguaiacol, 4-vinylphenol and 4-vinylguaiacol in wines. These compounds are involved in the presence of "cork taint" and Brett character in wines. The MHS-SPME method is a modification of SPME developed for quantitative analysis that avoids possible matrix effects based on an exhaustive analyte extraction from the sample. After demonstrating the existence of matrix effect in the analysis of the target compounds by HS-SPME with a divinylbenzene/Carboxen/polydimethylsiloxane (DVB/CAR/PDMS) fibre, the MHS-SPME method was developed and validated. The proposed method showed satisfactory linearity, precision and detection limits, all below the odour detection thresholds of the compounds in wine matrices. Good recoveries were observed for all compounds, always above 90%, and the repeatability obtained was considered acceptable, ranging between 2 and 11%. After checking the applicability of the method by comparing the results recorded with those obtained with the standard addition method, the method was applied successfully to the analysis of wine samples. To our knowledge, this is the first time that MHS-SPME combined with GC/MS/MS has been applied to simultaneously determine haloanisoles and volatile phenols in wine.     

Application of robust NiTi-ZrO2-PEG SPME fiber in the determination of haloanisoles in cork stopper samples

In this study, a novel solid-phase microextraction (SPME) fiber obtained using sol-gel technology was applied in the determination of off-flavor compounds (2,4,6-trichloroanisole (TCA), 2,4,6-tribromoanisole (TBA) and pentachloroanisole (PCA)) present in cork stopper samples. A NiTi alloy previously electrodeposited with zirconium oxide was used as the substrate for a poly(ethylene glycol) (PEG) coating. Scanning electronic microscopy showed good uniformity of the coating and allowed the coating thickness to be estimated as around 17 micarom. The optimization of the main parameters influencing the extraction efficiency, such as cork sample mass, sodium chloride mass, extraction temperature and extraction time were optimized using a full factorial design, followed by a Doehlert design. The optimum conditions were: 20 min of extraction at 70 degrees C using 60 mg of the cork sample and 10 mL of water saturated with sodium chloride in a 20 mL amber vial with constant magnetic stirring. Satisfactory detection limits between 2.5 and 5.1 ng g(-1) were obtained, as well as good precision (R.S.D. in the range of 5.8-12.0%). Recovery tests were performed on three different cork samples, and values between 83 and 119% were obtained. The proposed SPME fiber was compared with commercially available fibers and good results were achieved, demonstrating its applicability.     

Multiple response optimisation based on desirability functions of a microwave-assisted extraction method for the simultaneous determination of chloroanisoles and chlorophenols in oak barrel sawdust

A microwave-assisted extraction (MAE) method was optimised for extracting 2,4,6-trichloroanisole (TCA), 2,3,4,6-tetrachloroanisole (TeCA), pentachloroanisole (PCA), 2,4,6-trichlorophenol (TCP), 2,3,4,6-tetrachlorophenol (TeCP) and pentachlorophenol (PCP) from oak barrels. The method was optimised by using a central composite experimental design with extraction time, temperature and solvent volume as influential parameters. A desirability function was then employed in addition to the simultaneous optimisation of the compounds. The optimal conditions identified were temperature 130 degrees C, solvent volume 35 mL and extraction time 50 min. The compounds were determined by gas chromatography with electron-capture detection. MAE was compared with conventional Soxhlet extraction; the results obtained with MAE were in good agreement with those obtained by Soxhlet extraction.     

Optimisation of the derivatisation reaction and subsequent headspace solid-phase microextraction method for the direct determination of chlorophenols in red wine

An acetylation reaction for the derivatisation of the three chlorophenols involved in cork taint was optimised using a Doehlert design for direct application in wine samples. In this first step, the optimum reaction pH, by adding different amounts of KHCO3, and the required quantity of derivatisation reagent were fixed. Then a series of parameters relevant for the headspace solid-phase microextraction process, such as desorption conditions, salt addition and agitation sample were evaluated. A simultaneous study of the type of fibre and extraction temperature was performed at five levels and based on the results obtained the rest of factors (sample volume and exposition time) that could potentially affect the extraction yields were optimised by a central composite design. According to the validation of the method, we propose here, to our knowledge, the first application of solid-phase microextraction for the direct analysis of chlorophenols in red wine samples.     

Assessment of the matrix effect on the headspace solid-phase microextraction (HS-SPME) analysis of chlorophenols in wines

In this paper, we propose a comparative study to check the matrix effect on the extraction of three chlorophenols, 2,4,6-trichlorophenol (TCP), 2,3,4,6-tetrachlorophenol and pentachlorophenol, direct precursors of 2,4,6-trichloroanisole, in synthetic and commercial wines (white and red wines). A rapid, simple and sensitive methodology based on solid-phase microextraction (SPME) and GC with electron capture detection (GC-ECD) and mass spectrometric detection (GC-MS) was developed and the variables affecting the extraction process (temperature, time and salt content) were examined employing a factorial design at two levels. Since GC-ECD does not allow the clear identification of target analytes in white wine, owing to overlapped interferences, GC-MS/MS was used for subsequent examinations. Calibration curves were constructed in synthetic, white and red wine. Significant differences between the slopes of synthetic and red wine, with the exception of TCP, were observed. Analytical parameters were evaluated and satisfactory results were obtained, showing the usefulness of the headspace SPME (HS-SPME) method for determining chlorophenolic compounds in wines.     

Determination of 2,4,6-trichloroanisole and 2,4,6-tribromoanisole on ng L-1 to pgL-1 levels in wine by solid-phase microextraction and gas chromatography-high-resolution mass spectrometry

A gas chromatography-high-resolution mass spectrometry (GC-HRMS) method using solid-phase microextraction (SPME) for the determination of 2,4,6-trichloroanisole (TCA) and 2,4,6-tribromoanisole (TBA) in wine at low ng L(-1) levels was developed. A robust SPME method was developed by optimizing several different parameters, including type of fiber, salt addition, sample volume, extraction and desorption time. The quantification limit for TCA and TBA in wine was lowered substantially using GC-HRMS in combination with the optimized SPME method and allowed the detection of low analyte concentrations (ng L(-1)) with good accuracy. Limits of quantification for red wine of 0.3 ng L(-1) for TCA and 0.2 ng L(-1) for TBA with gas chromatography-negative chemical ionization mass spectrometry and 0.03 ng L(-1) for TCA and TBA were achieved using GC-HRMS. The method was applied to 30 wines of which 4 wines were sensorically qualified as cork defected. TCA was found in three of these wines with concentrations in the range 2-25 ng L(-1). TBA was not detected in any of the samples.     

Determination of 2,4,6-trichloroanisole and guaiacol in cork stoppers by pressurised fluid extraction and gas chromatography-mass spectrometry

This paper describes the development of an analytical method consisting of pressurised fluid extraction (PFE) and gas chromatography-mass spectrometry (GC-MS) using experimental designs to determine two volatile compounds in naturally-tainted cork stoppers. The target analytes, 2,4,6-trichloroanisole (2,4,6-TCA) and guaiacol, are involved in the cork taint of wine. First, a Plackett-Burman experimental design was carried out in order to determine the significant experimental parameters affecting the PFE process, and then a central composite design was used to optimise these significant parameters. Once the method had been optimised, the influence of the number of extraction cycles was studied. The method was applied to determine the concentration of 2,4,6-TCA and guaiacol in three cork samples, and the results were compared with the ones obtained by multiple headspace-solid-phase microextraction (MHS-SPME) and by Soxhlet extraction.     

Flavour analysis of Greek white wine by solid-phase microextraction-capillary gas chromatography-mass spectrometry

Solid-phase microextraction (SPME) was optimised for the qualitative determination of the volatile flavour compounds responsible for the aroma of Greek Boutari wine. Several factors influencing the equilibrium of the aroma compounds between the sample and the SPME fiber were taken into account, including the extraction time, the extraction temperature, the sampling mode (headspace and direct immersion or liquid SPME), and the presence of salt. Four different SPME fibers were used in this study. namely poly(dimethylsiloxane) (PDMS), poly(acrylate), carbowax-divinylbenzene and divinylbenzene-carboxen on poly(dimethylsiloxane). The best results were obtained using the PDMS fiber during headspace extraction at 25 degrees C for 30 min after saturating the samples with salt. The optimised SPME method was then applied to investigate the qualitative aroma composition of three other Greek wines, namely Zitsa, Limnos and Filoni.     

Headspace‐single drop microextraction with a commercial capillary electrophoresis instrument

Automated coupling of headspace‐single drop microextraction (HS‐SDME) and CE has been demonstrated using a commercial CE instrument. When a drop hanging at the inlet tip of a capillary for CE is used as the acceptor phase, HS‐SDME becomes a simple but powerful sample pretreatment technique for CE before injection to facilitate sample cleanup and enrichment. By combining HS‐SDME with an on‐line sample preconcentration technique, large volume sample stacking using an electroosmotic flow pump, the sensitivity can be improved further. The overall enrichment factors for phenolic compounds were from 1900 to 3400. HS‐SDME large volume sample stacking using an electroosmotic flow pump was successfully applied to a red wine sample to obtain an LOD of 4 nM (0.8 ppb) for 2,4,6‐trichlorophenol which is a precursor for 2,4,6‐trichloroanisole causing the foul odor in wine called cork taint.     

Optimisation of a pressurised liquid extraction method for haloanisoles in cork stoppers

Cork taint is a musty off-flavour in wines mainly caused by 2,4,6-trichloroanisole, but other haloanisoles can contribute. In this work, a method for the extraction of 2,4,6-trichloroanisole, 2,4,6-tribromoanisole and 2,6-dichloroanisole has been developed. The procedure involves the extraction of the haloanisoles from cork by pressurised liquid extraction and the analysis of the extracts by both GC-μECD and GC-MS-MS. A central composite design was used to investigate the dependence of the recoveries of the analytes on the temperature, percentage pentane-diethyl ether ratio and the extraction time. Experimental data were then processed by using the multiple regression analysis in order to calculate a mathematical model representing the relationship between factors and responses and to determine the best experimental conditions for PLE method. These conditions corresponded to a temperature of 176 °C, an extraction time between 2.8 and 4 min and an 80:20 pentane:diethyl ether ratio.     

Determination of carbonyl compounds in beer by derivatisation and headspace solid-phase microextraction in combination with gas chromatography and mass spectrometry

Headspace solid-phase microextraction (SPME) followed by gas chromatography and mass spectrometry was applied for quantification of 41 chemically diverse carbonyl compounds in beer. Therefore, in-solution derivatisation with o-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine (PFBHA) combined with SPME was optimised for fibre selection, PFBHA concentration, extraction temperature and time and ionic strength. Afterwards, the method was calibrated and validated successfully and extraction efficiency was compared to sampling with on-fibre derivatisation. In-solution derivatisation enabled the detection of several compounds that were poorly extracted with on-fibre derivatisation such as 5-hydroxymethylfurfural, acrolein, hydroxyacetone, acetoin, glyoxal and methylglyoxal. Others, especially (E)-2-nonenal, were extracted better with on-fibre derivatisation.     

Analysis of trace levels of pesticides in rainwater by SPME and GC-tandem mass spectrometry after derivatisation with PFFBr

Solid-phase microextraction (SPME) was used for the analysis of some pesticides (bromoxynil, chlorotoluron, diuron, isoproturon, 2,4-MCPA, MCPP and 2,4-D) in rainwater after derivatisation with PFBBr and gas chromatography-ion trap mass spectrometry. The derivatisation procedure was optimized by testing different methods: direct derivatisation in the aqueous phase followed by SPME extraction, on-fibre derivatisation and derivatisation in the injector. The best result was obtained by headspace coating the PDMS/DVB fibre with PFBBr for 10 min followed by direct SPME extraction for 60 min at 68 °C (pH 2 and 75% NaCl). Good detection limits were obtained for all the compounds: these ranged between 10 and 1,000 ng L⁻¹ with a relatively high uncertainty due to the combination of derivatisation and SPME extraction steps. The optimized procedure was applied to the analysis of pesticides in rainwater and results obtained shows that this method is a fast and simple technique to assess the spatial and temporal variations of concentrations of pesticides in rainwater.