Production of the mycotoxins fusaproliferin and beauvericin by South African isolates in the Fusarium section Liseola

The production of fusaproliferin (FUS), a recently described mycotoxin, and beauvericin (BEA), a mycotoxin recently reported to co-occur with FUS in Fusarium-infected corn, by South African isolates in the Fusarium section Liseola, was investigated. Five isolates each of F. verticillioides, F. proliferatum, F. subglutinans, and F. globosum were cultured on corn kernels. Four each of the five South African isolates of F. proliferatum and F. subglutinans produced FUS (10-1725 and 330-2630 mg/kg, respectively). BEA was produced by four of the F. proliferatum strains (310-1130 mg/kg) and three of the F. subglutinans strains (140-700 mg/kg). The isolates of F. verticillioides failed to produce significant levels of either of these secondary metabolites. F. globosum was a weak producer of both in that one isolate of five produced 25 mg/kg FUS and five out of five produced BEA at levels ranging between 10 and 110 mg/kg. To further characterize these strains, their production of fumonisins B(1), B(2), and B(3), as well as moniliformin, was investigated. Of the four species investigated, fumonisins were produced by all except F. subglutinans, which in turn was the only species whose isolates in this study produced moniliformin (four of five isolates, ranging from 155 to 2095 mg/kg). Analysis of visibly Fusarium-infected home-grown corn collected in the Transkei region of the Eastern Cape Province of South Africa showed that nine of the ten samples contained low levels of FUS (up to 62 microg/kg), whereas all ten samples showed BEA contamination ranging from 8 to 1734 microg/kg with a mean of 258 microg/kg.     

Analysis of diacetyl in wine using solid-phase microextraction combined with gas chromatography-mass spectrometry

Analytical difficulties in the rapid and accurate determination of diacetyl (DA), an important flavor compound in wine, at low concentrations have been overcome by the use of solid-phase microextraction (SPME) with deuterated diacetyl-d(6) (d6-DA) as an internal standard followed by gas chromatography-mass spectrometry (GC-MS). The GC-MS analyses showed that the values of the ion response ratio of DA to d6-DA were consistent regardless of the conditions of SPME headspace and were not influenced by the presence of sulfur dioxide in wine. The quantitation value of DA was represented as the concentration of free plus bound with sulfur dioxide forms of DA. The detection limit of DA in wine was as low as 0.01 microg/mL with linearity through to 10 microg/mL.     

Modeling flavor release using inverse gas chromatography-mass spectrometry

Inverse gas chromatography (IGC)-mass spectrometry was used to determine the extent of flavor release from a food matrix as a function of moisture uptake. At the surface of a solid, components with higher binding affinities can exchange with and replace components with lower binding affinities. As a low moisture baked product absorbs moisture from the air, flavor is lost from the matrix as water molecules exchange with the flavor molecules. The amount of flavor lost over time can be modeled using this approach to determine the onset of flavor release and total amount of flavor release as well as the identity of the released components and their relative order of exclusion from the matrix.     

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).     

Analysis of cooking oil fumes by ultraviolet spectrometry and gas chromatography-mass spectrometry

This paper investigated the components, especially aldehydes, in the fume condensates from four kinds of cooking oil using ultraviolet (UV) spectrometry and gas chromatography-mass spectrometry (GC-MS). It was observed that there was a great change of the UV absorption spectra from the results of the unheated oil to the results of the fume after heat treatment (190-200, 230-240, and 270-280 degrees C). There was a strong peak within the wavelength range of 260-270 nm in each condensate sample. From the GC-MS results, it was tentatively deduced that there were some 2,4-dialkylenaldehydes and other conjugated compounds in the condensates. The results showed there were large amounts of hexanal and 2-heptenal in the cooking oil fume and that the total aldehyde peak areas of the condensates from four kinds of oil were around 30-50% of the total peak area at 270-280 degrees C.     

Analysis of acrylamide in green tea by gas chromatography-mass spectrometry

Optimization of the solid-phase extraction cleanup procedure enabled the GC-MS analysis of acrylamide in tea samples without the interference of bromination by tea catechins. Although polyvinylpolypyrrolidone (PVPP) is available for removing tea catechins from tea extract, the peaks derived from PVPP had the same retention time as brominated acrylamide in mass chromatograms obtained by GC-MS. A considerable amount of acrylamide was formed at roasting temperatures of > or =120 degrees C; the highest acrylamide level was observed when tea samples were roasted at 180 degrees C for 10 min. Higher temperatures and longer processing times caused a decrease in the acrylamide content. Furthermore, an analysis of 82 tea samples showed that rather than the reducing sugar content, the asparagine content in tea leaves was a significant factor related to acrylamide formation in roasted products. The acrylamide level in roasted tea products was controlled by asparagine in the presence of reducing sugars.     

Analysis of glycated and ascorbylated proteins by gas chromatography-mass spectrometry

Proteins or poly-L-lysine which were incubated in the presence of ascorbic acid, dehydroascorbic acid (ascorbylation), or various sugars (glycation) were analyzed by gas chromatography-mass spectrometry (GC-MS). To also detect more labile reaction products, the Maillard modified proteins or poly-L-lysine were enzymatically hydrolyzed and reacted with N-methyl-N-(tert-butyldimethylsilyl)trifluoroacetamide to form the N(O)-tert-butyldimethylsilyl (tBDMS) derivatives prior to GC analysis. Under these conditions, the known Maillard products N (epsilon)-(carboxymethyl)lysine (1), oxalic acid mono-N (epsilon)-lysinylamide (2), and N (epsilon)-(carboxyethyl)lysine (3) could be simultaneously detected and quantified in glycated and ascorbylated proteins. Additionally, N (epsilon)-(1-carboxy-3-hydroxypropyl)-L-lysine (4) was identified for the first time as a Maillard product of proteins. Under the conditions applied here, 4 was found only in ascorbylated proteins or poly-L-lysine, but not in glycated proteins. Maillard-modified poly-L-lysine was further subjected to high-performance liquid chromatography (HPLC) analysis after enzymatic hydrolysis and formation of the phenyl isothiocyanate derivatized amino acids. Using this method, N (epsilon)-formyl-L-lysine (5), which cannot be distinguished from 2 by GC-MS analysis, was identified for the first time as a glycation product. Compound 5 is mainly formed from ribose, lactose, and fructose. The indicated Maillard products were quantified in beta-lactoglobulin (GC-MS) or poly-L-lysine (HPLC) which were glycated or ascorbylated using different precursors.     

Analysis of pesticides in honey by solid-phase extraction and gas chromatography-mass spectrometry

An analytical method for the simultaneous determination of 51 pesticides in commercial honeys was developed. Honey (10 g) was dissolved in water/methanol (70:30; 10 mL) and transferred to a C(18) column (1 g) preconditioned with acetonitrile and water. Pesticides were subsequently eluted with a hexane/ethyl acetate mixture (50:50) and determined by gas chromatography with electron impact mass spectrometric detection in the selected ion monitoring mode (GC-MS-SIM). Spiked blank samples were used as standards to counteract the matrix effect observed in the chromatographic determination. Pesticides were confirmed by their retention times, their qualifier and target ions, and their qualifier/target abundance ratios. Recovery studies were performed at 0.1, 0.05, and 0.025 microg/g fortification levels for each pesticide, and the recoveries obtained were >86% with relative standard deviations of <10%. Good resolution of the pesticide mixture was achieved in approximately 41 min. The detection limits of the method ranged from 0.1 to 6.1 microg/kg for the different pesticides studied. The developed method is linear over the range assayed, 25-200 microg/L, with determination coefficients of >0.996. The proposed method was applied to the analysis of pesticides in honey samples, and low levels of a few pesticides (dichlofluanid, ethalfluralin, and triallate) were detected in some samples.     

Analysis of ergosterol in single kernel and ground grain by gas chromatography-mass spectrometry

A method for analyzing ergosterol in a single kernel and ground barley and wheat was developed using gas chromatography-mass spectrometry (GC-MS). Samples were saponified in methanolic KOH. Ergosterol was extracted by "one step" hexane extraction and subsequently silylated by N-trimethylsilylimidazole/trimethylchlorosilane (TMSI/TMCS) reagent at room temperature. The recoveries of ergosterol from ground barley were 96.6, 97.1, 97.1, 88.5, and 90.3% at the levels of 0.2, 1, 5, 10, and 20 microg/g (ppm), respectively. The recoveries from a single kernel were between 93.0 and 95.9%. The precision (coefficient of variance) of the method was in the range 0.8-12.3%. The method detection limit (MDL) and the method quantification limit (MQL) were 18.5 and 55.6 ng/g (ppb), respectively. The ergosterol analysis method developed can be used to handle 80 samples daily by one person, making it suitable for screening cereal cultivars for resistance to fungal infection. The ability for detecting low levels of ergosterol in a single kernel provides a tool to investigate early fungal invasion and to study mechanisms of resistance to fungal diseases.     

Interaction of fusarium mycotoxins, fusaproliferin and fumonisin B1, with DNA studied by electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry (ESI-MS) in negative ion mode was used to monitor the possible noncovalent adduct formations between DNA analogue oligonucleotides and two Fusarium mycotoxins, fumonisin B1 and fusaproliferin. Using mild experimental ESI conditions specific noncovalent interactions were detected between both single- and double-stranded model oligonucleotides and fusaproliferin with 1:1 stoichiometry. Similar association complexes were observed for the deacetyl derivative of fusaproliferin. There were no peaks due to adduct formation present in the mass spectra of fumonisin B1, incubated with oligonucleotides in a wide concentration range, suggesting no specific interaction for this molecule. In a competitive complexation reaction, another mycotoxin, the beauvericin, forms more stable association complex with DNA than fusaproliferin. These findings can be of use in the understanding of molecular mechanisms of action during apoptosis and can be correlated with the teratogenic effect of fusaproliferin.     

Semiautomated determination of pesticides in water using solid phase extraction disks and gas chromatography-mass spectrometry

A method based on semiautomated solid phase extraction using octadecyl-bonded silica disks and gas chromatography-mass spectrometry, operated in selected ion monitoring mode, allows detection and quantification of approximately 100 pesticides and transformation products in drinking water. Samples (500 mL) were passed through the disk, and the retained pesticides were eluted with acetone and ethyl acetate. Typical recoveries for pesticides at 0.1 microg L(-1) in water were in the range of 72-120% with relative standard deviations less than 20%. Calibration curves were linear over the range of 0.025-0.5 microg mL(-1) (equivalent to a concentration range in drinking water of 0.05-1.0 microg L(-1)).     

Analysis of volatiles from Spanish honeys by solid-phase microextraction and gas chromatography-mass spectrometry

Headspace solid-phase microextraction (SPME), followed by gas chromatography (GC)-mass spectrometry (MS) determination, has been used for the analysis of honey volatiles. Two SPME fibers were employed to study the composition of volatiles from various types of Spanish honeys. The best results were obtained with the Carboxen/PDMS fiber, using a homogenization time of 1 h at 70 degrees C and a sampling period of 30 min. A total of 35 compounds were detected, most of them identified by GC-MS and quantified using external standards. Differences in the composition of honey volatiles were obtained, and these results allowed the differentiation of honeys. However, further studies are necessary to confirm the utility of this technique as an alternative tool for the characterization of the floral origin of honeys.     

Cinnamaldehyde content in foods determined by gas chromatography-mass spectrometry

trans-Cinnamaldehyde, the principal component of cinnamon flavor, is a potent antimicrobial compound present in essential oils such as cinnamon. In the course of studies designed to discover its maximum microbial lethality under food-processing conditions, a gas chromatographic-mass spectrophotometric procedure was developed for the extraction and analysis of essential oil components such as cinnamaldehyde from commercial cinnamon-containing foods (several brands of cinnamon breads, cereals, cookies, puddings, applesauces, and fruit juices). The cinnamaldehyde content ranged from trace amounts in orange juice to 12.2 mg/100 g (122 ppm) in apple cinnamon cereals and 31.1 mg/100 g (311 ppm) for cinnamon swirl bread (highest value). To ascertain the heat stability of cinnamaldehyde, pure cinnamaldehyde, pure eugenol, cinnamon oil, and mixtures consisting of cinnamaldehyde plus eugenol or cinnamon oil were heated at graded temperatures up to 210 degrees C and 60 min, and then possible compositional changes were examined. Eugenol was stable to heat, as were the components of cinnamon oil: carvone, eugenol, and linalool. In contrast, starting at approximately 60 degrees C, pure cinnamaldehyde undergoes a temperature-dependent transformation to benzaldehyde under the influence of heat. Eugenol, both pure and in cinnamon oil, when added to pure cinnamaldehyde protected the aldehyde against heat destruction. The protection may due to an antioxidative action of eugenol. The possible mechanism of this effect and the significance of these findings for food chemistry and microbiology are discussed.     

Mass spectral investigations on trichothecene mycotoxins. II. Detection and quantitation of macrocyclic trichothecenes by gas chromatography/negative ion chemical ionization mass spectrometry

A general, sensitive gas chromatographic/negative ion chemical ionization mass spectrometric (GC/NICIMS) method of analysis was developed for the detection and quantitation of several polar, thermally labile, toxic macrocyclic trichothecenes. The procedure involves the conversion of the molecules to their corresponding alcohols (verrucarols) by alkaline hydrolysis, followed by derivatization of the hydrolysate with heptafluorobutyrylimidazole and analysis by GC/MS technique under negative ion chemical ionization conditions. Nanogram (250 ng) quantities of several macrocyclic trichothecenes with different verrucarol and ester moieties were analyzed successfully with good precision by this procedure. The method was applicable for the accurate determination of at least low ppb levels of these macrocyclic trichothecenes in environmental samples, such as fungal products, fermentation broths, and plant samples. This is the first reported, well developed, sensitive, and applicable method for the detection and quantitation of these compounds in naturally occurring samples.     

Pathway analysis of branched-chain ester biosynthesis in apple using deuterium labeling and enantioselective gas chromatography-mass spectrometry

The biosynthesis of volatile esters by Red Delicious apples was investigated by incubating fruit tissue with deuterated flavor precursors at various times after controlled atmosphere (CA) storage and measuring deuterium incorporation into branched-chain ester volatiles. 2-Methylbutyl acetate was the only volatile not significantly reduced by CA storage. Conversion of 2-methylbutanol to 2-methylbutyl acetate and of 2-methylbutanoic acid to ethyl 2-methylbutanoate and to hexyl 2-methylbutanoate was limited by the availability of 2-methylbutyl substrates but not by acetyl-CoA, ethanol, or hexanol, respectively. The enzymatic activity required for these reactions declined during CA storage. The conversion of 2-methylbutanoic acid to 2-methylbutanol was also substrate limited, but enzymic activity appeared stable in storage. Biosynthesis of both 2-methylbutanoic acid and 2-methylbutanol, from isoleucine, was severely depressed under CA storage. The reduced metabolism of isoleucine to 2-methylbutanoyl-CoA may be the primary reason for reduced branched-chain ester synthesis in CA-stored Red Delicious apples. Enantioselective gas chromatography-mass spectrometry confirmed that the chirality of (S)-2-methylbutyl acetate derives from l-isoleucine with the other enzymes in this pathway not being enantiospecific. Treatment of tissue samples with 2-methylbut-2E-enal gave only (S)-2-methylbutyl acetate, indicating that biosynthesis was not via tiglyl-CoA.     

Determination of 2,4,6-trichloroanisole and 2,4,6-tribromoanisole in wine using microextraction in packed syringe and gas chromatography-mass spectrometry

A selective and fast method for the quantitative determination of 2,4,6-trichloroanisole (TCA) and 2,4,6-tribromoanisole (TBA) in wine was developed. Microextraction in packed syringe (MEPS) was optimized for the extraction and preconcentration of the analytes using extremely small volume samples (0.1-1 mL). For GC-EI-MS, the limit of detection (LOD) for red and white wine was in the range 0.17-0.49 microg L(-1) for TCA and TBA. In addition to GC-EI-MS both GC-NCI-MS and GC-HRMS were used to further improve both selectivity and sensitivity. The lowest LODs were achieved using GC-HRMS in the EI mode. In red and white wine samples the LODs were between 0.22-0.75 ng L(-1) for TCA and TBA. The reproducibility and linearity for the GC-HRMS method was good, with RSD-values of 4-10% for spiked red wine samples at 1 ng L(-1) and linearity with R (2) > 0.962 over a concentration range of 1 to 100 ng L(-1).     

Analysis of veterinary drugs and metabolites in milk using quadrupole time-of-flight liquid chromatography-mass spectrometry

A quadrupole time-of-flight (Q-TOF) liquid chromatography-mass spectrometry (LC-MS) method was developed to analyze veterinary drug residues in milk. Milk samples were extracted with acetonitrile. A molecular weight cutoff filter was the only cleanup step in the procedure. Initially, a set of target compounds (including representative sulfonamides, tetracyclines, β-lactams, and macrolides) was used for validation. Screening of residues was accomplished by collecting TOF (MS(1)) data and comparing the accurate mass and retention times of found compounds to a database containing information for veterinary drugs. The residues included in the study could be detected in samples fortified at the levels of concern with this procedure 97% of the time. Although the method was intended to be qualitative, an evaluation of the MS data indicated a linear response and acceptable recoveries for a majority of target compounds. In addition, MS/MS data were also generated for the [M + H](+) ions. Product ions for each compound were identified, and their mass accuracy was compared to theoretical values. Finally, incurred milk samples from cows dosed with veterinary drugs, including sulfamethazine, flunixin, cephapirin, or enrofloxacin, were analyzed with Q-TOF LC-MS. In addition to monitoring for the parent residues, several metabolites were detected in these samples by TOF. Proposed identification of these residues could be made by evaluating the MS and MS/MS data. For example, several plausible metabolites of enrofloxacin, some not previously observed in milk, are reported in this study.     

Multiresidue determination of pesticides in soil by gas chromatography-mass spectrometry detection

An analytical multiresidue method for the simultaneous determination of various classes of pesticides in soil was developed. Pesticides were extracted from soil with ethyl acetate. Soil samples were placed in small columns, and the extraction was carried out assisted by sonication. Pesticides were determined by gas chromatography with electron impact mass spectrometric detection in the selected ion monitoring mode. Spiked blank samples were used as standards to counteract the matrix effect observed in the chromatographic determination. Pesticides were confirmed by their retention times, their qualifier and target ions, and their qualifier/target abundance ratios. Recovery studies were performed at 0.2, 0.1, and 0.05 microg/g fortification levels of each pesticide, and the recoveries obtained ranged from 87.0 to 106.2% with a relative standard deviation between 2.4 and 10.6%. Good resolution of the pesticide mixture was achieved in approximately 41 min. The detection limits of the method ranged from 0.02 to 1.6 microg/kg for the different pesticides studied. The developed method is linear over the range assayed, 25-1000 microg/L, with determination coefficients >0.999. The proposed method was used to determine pesticide levels in real soil samples, taken from different agricultural areas of Spain, where several herbicides and insecticides were found.