Headspace solid-phase microextraction method for the study of the volatility of selected flavor compounds

Changes in the volatility of selected flavor compounds in the presence of nonvolatile food matrix components were studied using headspace solid-phase microextraction (HS-SPME) combined with GC-MS quantification. Time-dependent adsorption profiles to the SPME fiber and the partition coefficients between different phases were obtained for several individual volatiles, showing that HS-SPME analysis with a short sampling time can be used to determine the "true" headspace concentration at equilibrium between the headspace and a sample matrix. Equilibrium dialysis followed by HS-SPME/GC-MS was carried out to confirm the ability of HS-SPME extraction for monitoring the free volatile compounds in the presence of proteins. In particular, a short sampling time (1 min) avoided additional extraction of volatiles bound to the protein. Interactions between several selected flavor compounds and nonvolatile food matrix components [beta-lactoglobulin or (+)-catechin] were also studied by means of HS-SPME/GC-MS analysis. The volatility of ethyl hexanoate, heptanone, and hexanal was significantly decreased by the addition of beta-lactoglobulin compared to that of isoamyl acetate. Catechin decreased the volatility of ethyl hexanoate and hexanal by 10-20% and increased that of 2-heptanone by approximately 15%. This study indicates that HS-SPME can be a useful tool for the study of the interactions between volatile compounds and nonvolatile matrix components provided the kinetic and thermodynamic behavior of the volatiles in relation to the fiber chosen for the studies is carefully considered.     

Essential oil composition of sachalinmint from Norway detected by solid-phase microextraction and gas chromatography-mass spectrometry analysis

The essential oil of leaves and flowers of sachalinmint [Mentha sachalinensis (Briq.) Kud?] grown in Norway (Trondheim) has been studied by headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography-mass spectrometry analysis (GC-MS). The essential oil content increased linearly in acropetal direction from 1.08% (0-20 cm plant height) to 1.75% (60-80 cm; young leaves and flowers). The steam-distilled samples showed a minor complex matrix with a very high menthol and a much lower menthone content (87.89 and 4.05%, respectively). From testing of HS-SPME unequilibrated exposure times ranging from 10 s to 5 min, an extraction time of 30 s was found to be sufficient to detect both low- and high-eluting compounds. Comparison of HS-SPME and steam-distilled samples established that the same tendencies of increasing menthol/menthone content in the basipetal/acropetal direction could be detected by both analysis methods. With regard to the extraction efficiency, HS-SPME gave additional detailed information about less important terpenic compounds.     

Determination of 2-methylisoborneol and geosmin produced by Streptomyces sp. and Anabaena PCC7120

A new sample preparation and enrichment technique, headspace liquid-phase microextraction (HS-LPME) linked to gas chromatography-mass spectrometry (GC-MS), was developed for the determination of the off-flavor odorants, 2-methylisoborneol and geosmin, produced by Streptomyces sp. and Anabaena PCC7120. Some of the factors that influence the extraction efficiency of HS-LPME, such as the type of extraction solvent, ionic strength of sample solution, and sample agitation rate, were studied and optimized by a single factor test. Other factors, including extraction temperature, extraction time, microdrop volume, and headspace volume were optimized by orthogonal array design. Extraction of 2-methylisoborneol and geosmin was conducted by exposing 2.5 microL of 1-hexanol for 9 min at 50 degrees C in the headspace of a 20 mL vial with a 10 mL of sample solution saturated by NaCl and stirred at 800 rpm. The developed protocol demonstrated good repeatability (relative standard deviations (RSDs) < 5%), wide linear ranges (10-5000 ng/L, r2 > 0.999), and low limits of detection (LODs) for 2-methylisoborneol and geosmin (0.05 ng/L for both analytes). Subsequently, the method was successfully applied to extract the analytes in bacterial cultures with high recoveries (from 94% to 98%). Compared with headspace solid-phase microextraction (HS-SPME), HS-LPME demonstrates better linearity, precision, and recovery. Importantly, the sensitivity is about 1 order of magnitude higher than that of most HS-SPME. The results showed that HS-LPME coupled with GC-MS is a simple, convenient, rapid, sensitive, and effective method for the qualitative and quantitative analysis of 2-methylisoborneol and geosmin.     

Chemotypical variation of tansy (Tanacetum vulgare L.) from 40 different locations in Norway

Between 2001 and 2002, plant collections from wild populations of Norwegian tansy (Tanacetum vulgare L.) were studied with a focus on essential oil (EO) yield and composition in order to characterize the chemotypical EO variability. Tansy collections of 40 different locations from North, Mid-, and South Norway were transplanted to the Apelsvoll Research Centre Div. Kise in 2000 and grown for 2 years before the aerial parts (leaves and flower buds) were harvested in June 2002. The EO from individual plants was isolated from dried plant material by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS) on a DB5 column at the Plant Biocenter. The EO yield ranged between 0.35 and 1.90% (v/w) (average: 0.81%); the most abundant thujone plants were especially rich in EO volatiles (0.95%). On the basis of GC-MS data, seven chemotypes could be identified as follows: A, alpha-thujone (two individuals); B, beta-thujone (22); C, camphor (six); D, chrysanthenyl acetate/chrysanthenol (three); E, chrysanthenone (two); F, artemisia ketone/artemisia alcohol (three); and G, 1,8-cineole (two). The thujone chemotype was dominated by beta-thujone (81%) associated with alpha-thujone, but tansy plants rich in alpha-thujone were also detected (61%). The chemotypical classification of Norwegian tansy genotypes was underscored by preliminary studies from 2001, indicating the genetic uniformity and biochemical stability of the domesticated plants.     

Influence of seed endophyte amounts on swainsonine concentrations in astragalus and oxytropis locoweeds

Locoism is a toxic syndrome of livestock caused by the ingestion of a subset of legumes known as locoweeds endemic to arid and semiarid regions of the western United States. Locoweeds contain the toxic alkaloid swainsonine, which is produced by the endophytic fungi Undifilum species. Two chemotypes of plants can coexist within toxic populations of locoweeds: chemotype 1 plants are defined as individuals containing swainsonine concentrations greater than 0.01% and quantitatively greater amounts of Undifilum, while chemotype 2 plants are defined as individuals containing less than 0.01% swainsonine and quantitatively smaller amounts of Undifilum. To elucidate the mechanisms that govern chemotypes, the amount of Undifilum in seeds/embryos was manipulated, thus altering subsequent swainsonine concentrations in three locoweed species: Astragalus mollissimus, Astragalus lentiginosus, and Oxytropis sericea. Chemotype 1 seeds that were fungicide-treated or had the seed coat removed resulted in plants with swainsonine concentrations comparable to those in chemotype 2 plants. Conversely, embryos from seeds of chemotypes 1 and 2 that were inoculated with the endophyte resulted in plants with swainsonine concentrations comparable to those of chemotype 1 plants. This reproducible interconversion between the two swainsonine chemotypes suggests that the quantity of endophyte present in the seed at the time of germination is a key determinant of the eventual chemotype. Additionally, this is the first report of the inoculation of locoweeds with the endophyte Undifilum species.     

Analysis of benzothiazole in Italian wines using headspace solid-phase microextraction and gas chromatography-mass spectrometry

Benzothiazoles are a part of the molecular structure of a large number of natural products, biocides, drugs, food flavors, and industrial chemicals. They also appear in the environment mainly as a result of their production and use as rubber vulcanization accelerators. A new headspace solid-phase microextraction (HS-SPME) method for analysis of benzothiazole (BTH) in wine is described. This method is fast, inexpensive, and does not require solvents. The detection limit of BTH in wine was 45 ppt with linearity up to 100 ppb. The quantification of BTH is performed by the standard additions method and does not require the use of an internal standard. We have analyzed 12 wines from different grape varieties grown in several regions, using SPME extraction and gas chromatography-mass spectrometry (GC-MS) detection. Under these experimental conditions, benzothiazole was found in all wines analyzed. Concentration levels in samples varied from 0.24 microg/L (Vermentino) to 1.09 microg/L (Franciacorta).     

Headspace sorptive extraction (HSSE), stir bar sorptive extraction (SBSE), and solid phase microextraction (SPME) applied to the analysis of roasted Arabica coffee and coffee brew.

Headspace sorptive extraction (HSSE) and stir bar sorptive extraction (SBSE), two recently introduced solventless enrichment techniques, have been applied to the analysis of the headspace of Arabica roasted coffee and of the headspace of the brew and of the brew itself. In both HSSE and SBSE enrichment is performed on a thick film of poly(dimethylsiloxane) (PDMS) coated onto a magnet incorporated in a glass jacket. Sampling is done by placing the PDMS stir bar in the headspace (gas phase extraction or HSSE) or by immersing it in the liquid (liquid phase extraction or SBSE). The stir bar is then thermally desorbed on-line with capillary GC-MS. The performance of HSSE and SBSE have been compared through the determination of the recoveries and relative abundances of 16 components of the coffee volatile fraction to classical static headspace (S-HS) and to headspace and in-sample solid phase microextraction (HS-SPME and IS-SPME, respectively) applying the fibers PDMS 100 microm, Carbowax/divinylbenzene 65 microm (CW/DVB), Carboxen/PDMS 75 microm(CAR/PDMS), polyacrylate 85 microm(PA), PDMS/divinylbenzene 65 microm(PDMS/DVB), and Carboxen/divinylbenzene/PDMS 50-30 microm(CAR/PDMS/DVB). In all cases, HSSE and SBSE gave higher recoveries, and this is entirely due to the high amount of PDMS applied.     

Comparison of static headspace, headspace solid phase microextraction, headspace sorptive extraction, and direct thermal desorption techniques on chemical composition of French olive oils

Static headspace (SHS), headspace solid phase microextraction (HS-SPME), headspace sorptive extraction (HSSE), and direct thermal desorption (DTD) were applied to the analysis of four French virgin olive oils from Corsica. More than 60 compounds were isolated and characterized by GC-RI and GC-MS. SHS was not suited to the characterization of olive oil volatile compounds because of low sensitivity. The SPME and HSSE techniques were successfully applied to olive oil headspace analysis. Both methods allow the characterization of volatile compounds (mainly C(6) aldehydes and alcohols), which contribute significantly to the "green" flavor note of virgin olive oils. The PDMS stir bar showed a higher concentration capacity than a DVB/CAR/PDMS SPME fiber due to the higher volume of polymeric coating. DTD was a very good tool for extracting volatile and especially semivolatile compounds, such as sesquiterpenes, but requires a significant investment like that for HSSE. Finally, SPME may be a more appropriate technique for routine quality control due to its operational simplicity, repeatability, and low cost.     

Yield and oil composition of 38 basil (Ocimum basilicum L.) accessions grown in Mississippi

A field experiment was conducted to assess yield, oil content, and composition of 38 genotypes of sweet basil ( Ocimum basilicum L.). Overall, biomass yields were high and comparable to those reported in the literature. However, basil genotypes differed significantly with respect to oil content and composition. Oil content of the tested accessions varied from 0.07% to 1.92% in dry herbage. On the basis of the oil composition, basil accessions were divided into seven groups: (1) high-linalool chemotype [19-73% (-)-linalool], (2) linalool-eugenol chemotype [six chemotypes with 28-66% (-)-linalool and 5-29% eugenol], (3) methyl chavicol chemotype [six accessions with 20-72% methyl chavicol and no (-)-linalool], (4) methyl chavicol-linalool chemotype [six accessions with 8-29% methyl chavicol and 8-53% (-)-linalool], (5) methyl eugenol-linalool chemotype [two accessions with 37% and 91% methyl eugenol and 60% and 15% (-)-linalool], (6) methyl cinnamate-linalool chemotype [one accession with 9.7% methyl cinnamate and 31% (-)-linalool], and (7) bergamotene chemotype [one accession with bergamotene as major constituent, 5% eucalyptol, and <1% (-)-linalool]. Our results demonstrated that basil could be a viable essential oil crop in Mississippi. The availability of various chemotypes offers the opportunity for production of basil to meet the market requirements of specific basil oils or individual compounds such as (-)-linalool, eugenol, methyl chavicol, methyl cinnamate, or methyl eugenol.     

Screening of tropical fruit volatile compounds using solid-phase microextraction (SPME) fibers and internally cooled SPME fiber

In this study, the optimization and comparison of an internally cooled fiber [cold fiber with polydimethylsiloxane (PDMS) loading] and several commercial solid-phase microextraction (SPME) fibers for the extraction of volatile compounds from tropical fruits were performed. Automated headspace solid-phase microextraction (HS-SPME) using commercial fibers and an internally cooled SPME fiber device coupled to gas chromatography-mass spectrometry (GC-MS) was used to identify the volatile compounds of five tropical fruits. Pulps of yellow passion fruit (Passiflora edulis), cashew (Anacardium occidentale), tamarind (Tamarindus indica L.), acerola (Malphigia glabra L.), and guava (Psidium guajava L.) were sampled. The extraction conditions were optimized using two experimental designs (full factorial design and Doehlert matrix) to analyze the main and secondary effects. The volatile compounds tentatively identified included alcohols, esters, carbonyl compounds, and terpernes. It was found that the cold fiber was the most appropriate fiber for the purpose of extracting volatile compounds from the five fruit pulps studied.     

Optimization of multiple headspace solid-phase microextraction for the quantification of volatile compounds in dry fermented sausages

Multiple headspace solid-phase microextraction (HS-SPME) is a stepwise method that eliminates the influence of the matrix sample on the quantitative analysis of solid samples. The process was optimized for the analysis of volatile compounds in dry fermented sausages by gas chromatography and mass spectrometry. Different amounts of fermented sausages and different vial volumes were studied to obtain the theoretical exponential decay of the peak area of the four successive extractions in order to calculate the total area in the sausage. The highest number of volatile compounds analyzed by multiple HS-SPME in dry fermented sausages was obtained in a 10 mL headspace vial with 0.07 g of sample in the presence of water, 0.75 mg butylated hydroxytoluene, and 0.5 g sodium chloride. Finally, the method was characterized in terms of linearity and detection limits and applied to analyze the volatile compounds present in fermented sausages manufactured with either nitrate or nitrite.     

Optimization of headspace solid-phase Microextraction (SPME) for the odor analysis of surface-ripened cheese

Fifty volatile compounds of surface smear-ripened cheese were detected and identified using headspace solid-phase microextraction (HS-SPME) and vacuum distillation coupled to gas chromatography-mass spectrometry. Changes in the headspace of aroma compounds were monitored over the whole packaging period (47 days) using the HS-SPME method. Initially, the concentration of methanethiol increased before reaching a plateau. This evolution could be linked to the growth of Brevibacterium linens. During the shelf life of cheese, levels of acetic acid and 3-methylbutanoic acid remained constant, whereas butane-2,3-dione, 3-hydroxybutan-2-one, and hydroxypropan-2-one levels gradually declined and acetone and 3-methylbutanol levels dropped sharply to a plateau. Changes in odor could be related to changes of the rind, which behaved as a barrier, strongly influencing the distribution of volatile compounds in the headspace. Using a gas chromatography-olfactometry technique without separation, it was shown that the SPME extract was representative of the cheese odor.     

Correlations between pulp properties of eucalyptus clones and leaf volatiles using automated solid-phase microextraction

Analysis of biogenic volatile organic compounds (BVOC) of 14 Eucalyptus clones has been performed using an automated headspace solid-phase microextraction (HS-SPME) coupled to gas chromatography (GC)/ion trap mass spectrometry (ITMS) method. Correlations between pulp properties of Eucalyptus clones and the BVOC of their leaf headspaces were studied. The compounds alpha-terpineol and the sesquiterpene beta-eudesmol were positively correlated with S5, a property related to the hemicelluose content in the pulp. Qualitative results obtained with automated HS-SPME were sufficient to group together the same species and related hybrids through cluster analysis and were confirmed through principal component analysis. A preliminary separation of the essential oils of Eucalyptus dunnii through comprehensive two-dimensional gas chromatography (GC x GC) showed approximately 580 peaks compared to approximately 60 in a typical GC/ITMS first-dimension chromatogram. The potential of HS-SPME coupled to GC x GC to improve the separation of Eucalyptus volatiles and other plant essential oils looks extremely promising for new applications of unsupervised learning methods.     

Solid-phase microextraction of volatile compounds from the chopped leaves of three species of Eucalyptus

Headspace solid-phase microextraction (HS-SPME) coupled with gas chromatography and ion-trap mass spectrometry has been used to identify biogenic volatile organic compounds present in the headspace of chopped leaves of Eucalyptus (E.) dunnii, E. citriodora, and E. saligna. A simple HS-SPME method entailing 30 min of extraction at 30 degrees C was developed for this purpose. Thirty compounds were identified in the headspace of 60 juvenile chopped Eucalyptus leaves, and another 30 were tentatively identified. The presence of compounds such as (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), (E,E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene (TMNT), (E,E)-alpha-farnesene, (E,E,E)-3,7,11,15-tetramethyl-1,3,6,10,14-hexadecapentaene (TMHP), beta-caryophyllene, alpha-humulene, germacrene D, and beta-cubebene in the headspace of the leaves but not in the essential oils from the same Eucalyptus trees and information about the infochemical roles of some of these compounds in other living plant systems suggest they might play a bioactive role in Eucalyptus leaves.     

Characterization of aroma compounds in apple cider using solvent-assisted flavor evaporation and headspace solid-phase microextraction

The aroma-active compounds in two apple ciders were identified using gas chromatography-olfactometry (GC-O) and GC-mass spectrometry (MS) techniques. The volatile compounds were extracted using solvent-assisted flavor evaporation (SAFE) and headspace solid-phase microextraction (HS-SPME). On the basis of odor intensity, the most important aroma compounds in the two apple cider samples were 2-phenylethanol, butanoic acid, octanoic acid, 2-methylbutanoic acid, 2-phenylethyl acetate, ethyl 2-methylbutanoate, ethyl butanoate, ethyl hexanoate, 4-ethylguaiacol, eugenol, and 4-vinylphenol. Sulfur-containing compounds, terpene derivatives, and lactones were also detected in ciders. Although most of the aroma compounds were common in both ciders, the aroma intensities were different. Comparison of extraction techniques showed that the SAFE technique had a higher recovery for acids and hydroxy-containing compounds, whereas the HS-SPME technique had a higher recovery for esters and highly volatile compounds.     

Natural product chemistry meets genetics: when is a genotype a chemotype?

The chemotype of a microbial or plant species has traditionally been defined as its profile of natural products, and the genotype has been defined as its genetic constitution or DNA sequence. The purpose of this perspective is to discuss applications of DNA genotyping, particularly by polymerase chain reaction (PCR)-amplification methods, to predicting natural product chemotypes of fungi and plants of importance in food and agriculture. Development of PCR genotyping for predicting chemotypes will require collaboration between molecular biologists and natural product chemists, as well as community standards for reporting data. PCR genotyping should be validated by chemical analysis of individuals that represent the allelic diversity of the target gene in the population. To avoid misinterpretation, it is critical to differentiate data obtained by genotyping from data obtained by chemical analysis. The obvious and appropriate solution is to retain the established meanings of genotype and chemotype, both of which have been in use for half a century in the fields of genetics and natural product chemistry.     

基于HS-SPME-GC-MS法比较瓢鸡和盐津乌骨鸡不同部位挥发性风味成分

为了研究优质地方鸡种瓢鸡和盐津乌骨鸡不同部位的主体风味成分,以300日龄瓢鸡和盐津乌骨鸡的胸肌和腿肌作为试验对象,利用顶空固相微萃取(HS-SPME)技术提取,采用气相色谱质谱联用(GC-MS)技术分离和鉴定鸡肉中的挥发性物质,结合相对活度值(ROAV)确定主体风味活性物质。结果表明,鸡肉样品中共检出76种挥发性化合物,主要包括醛类、醇类、酮类、酯类、酸类、烃类化合物,不同品种不同部位之间挥发性风味物质的组分和含量存在差异。瓢鸡主体风味物质由2-甲基丁醛、戊醛、己醛、庚醛、辛醛、反-2-辛烯醛、壬醛、1-辛烯-3-醇、辛烷构成;盐津乌骨鸡主体风味物质主要由2-甲基丁醛、己醛、壬醛、1-辛烯-3-醇构成。主体风味物质对不同部位不同品种鸡肉样品的贡献程度不同,其中醛类化合物对鸡肉的整体风味贡献最大。本研究结果为瓢鸡和盐津乌骨鸡的风味特性研究和开发利用提供了理论依据。     

Headspace solid-phase microextraction gas chromatography-mass spectrometry analysis of volatiles in orujo spirits from a defined geographical origin

A headspace solid-phase microextraction (HS-SPME) and gas chromatography-selective ion monitoring/mass spectrometry (GC-SIM/MS) method was optimized for analysis of 22 volatile compounds in orujo spirit samples from the Geographic Denomination "Orujo de Galicia/Augardente de Galicia". HS-SPME experimental conditions, such as fiber coating, extraction temperature, extraction and pre-equilibrium time, sample volume, and the presence of salt, were studied to improve the extraction process. The best results were obtained using a 65 microm Carbowax-divinylbenzene fiber during a headspace extraction at 40 degrees C with constant magnetic stirring for 15 min and after a 5 min period of pre-equilibrium time. The sample volume was 6 mL of orujo containing 25% of NaCl, placed in 12 mL glass vials equipped with a screw cap and PTFE/silicone septum. Desorption was performed directly in the gas chromatograph injector port for 5 min at 250 degrees C using the splitless mode. The proposed method is sensible (with detection limits between 0.0045 and 0.2399 mg/L), precise (with coefficients of variation in the range 0.99-8.18%), and linear over more than 1 order of magnitude. The developed method presented recoveries comprised between 76.0 and 112.4%. The applicability of the new method was demonstrated by determining the considered 22 volatile compounds in nine orujo commercial samples with quality and origin brands.     

Gas chromatography-mass spectrometry determination of phosphine residues in stored products and processed foods

A gas chromatography-mass spectrometry (GC-MS) method was used for the quantitative confirmation of phosphine residues in stored products and processed foods. An established extraction technique was utilized for the preparation of headspace samples, which were analyzed by GC-MS and gas chromatography-nitrogen-phosphorus detection (GC-NPD). Wheat, oats, maize, white rice, brown rice, cornflakes, tortilla cornchips, groundnuts, and raisins were validated, showing excellent agreement between detectors when spiked at levels equivalent to 0.001 and 0.01 mg/kg phosphine and for samples containing incurred residues. The GC-MS method was reproducible and accurate when compared to the GC-NPD method and allowed five samples to be quantified in a working day. Subambient GC-MS oven temperatures were most suitable for phosphine residues ranging from 0.001 to 0.005 mg/kg, and a GC oven temperature of 100 degrees C was appropriate for residues >0.005 mg/kg. The method was sufficiently robust to be evaluated for other similar commodities as the need arises.