Bio-fortification and isotopic labelling of Se metabolites in onions and carrots following foliar application of Se and Se

The aims were to bio-fortify onions by foliar application of selenium (Se) and to intrinsically label bioactive Se-metabolites in onion and carrot by enriched, stable ⁷⁷Se for use in human physiological studies. Onion bulbs and leaves were enriched in Se by repeated foliar spraying of 10 or 100 μg Se ml⁻¹ solutions of sodium selenite (Se(IV)) or sodium selenate (Se(VI)). ICP-MS analysis of onion leaves and bulbs showed that the Se concentration was enhanced by up to a factor of approximately 50 and 200 in bulbs and leaves, respectively. HPLC–ICP-MS analysis of proteolytic plant extracts showed that foliar application of Se(IV) gave rise to bio-synthesis of a higher fraction of the desired organic Se species and was better tolerated by the plants than Se(VI). Based on these findings onions and carrots were bio-fortified by foliar application of a solution of ⁷⁷Se(IV) that was enriched to 99.7% as ⁷⁷Se. The ⁷⁷Se- labelled metabolites in onions were predominantly γ-glutamyl-⁷⁷Se-selenomethyl-selenocysteine (γ-glu-Me⁷⁷SeCys), ⁷⁷Se-methylselenocysteine (Me⁷⁷SeCys) and ⁷⁷Se-selenomethionine (⁷⁷SeMet). Furthermore, we report here for the first time the finding in carrots of the bioactive Me⁷⁷SeCys, the identity of which was verified by HPLC–ESI-MS/MS.     

Arsenic speciation in white wine by LC–ICP–MS

This study deals with As speciation in white wine. Arsenic species were selectively determined by liquid chromatography–inductively coupled plasma–mass spectrometry (LC–ICP–MS). Separation of As species was performed using an anion exchange column with ammonium phosphate solution (pH 6.00) as mobile phase. Samples of 14 white wine produced in South America were analysed. They were 10-fold diluted in the mobile phase prior to analysis by LC–ICP–MS. Accuracy was evaluated by recovery tests, whereas As species recovery ranged from 95% to 106%. Additionally, the sum of arsenic species concentration found by LC–ICP–MS was in agreement with the total arsenic concentration determined by ICP–MS after sample digestion. Arsenic species detected were arsenite [As(III)], arsenate [As(V)] and dimethylarsinic acid (DMA). As(III) and As(V) were detected in all analysed wine samples and DMA was detected only in wines produced in Argentina. Results for As determination in samples were from 2.9 to 10.3, 8.6 to 17.8, and <0.45 to 1.07 μg L⁻¹ for As(III), As(V) and DMA, respectively.     

Selenium speciation studies in Se-enriched chives (Allium schoenoprasum) by HPLC-ICP–MS

In this study, three liquid chromatographic techniques were employed to better understand selenium species distribution in Chives (Allium schoenoprasum) separately grown in three different supplementation media Se(IV), Se(VI), and SeMet. The highest selenium accumulation up to 700 μg Se g⁻¹ was observed in the case of the Se(VI)-enriched samples on the basis of total selenium measurements. Size-exclusion chromatography (SEC-HPLC) was performed for investigation of selenium containing proteins in chives. For speciation of selenium containing amino acids, reversed-phase ion-pairing chromatography (RP-IP-HPLC) and for enantiomeric separations, a crown ether column was used. In all three cases online detection with inductively coupled plasma mass spectrometry was performed for selenium specific detection. Two extractions (perchloric acid–ethanol and enzymatic) were carried out on chive samples. Speciation analysis on the chives grown in three different media revealed that selenium distribution among different forms of amino acids in the sample strongly depends on the type of enrichment employed. Enrichment with Se(VI) leads to accumulation of selenium in inorganic forms, while in case of Se(IV) and SeMet-enriched samples, methyl-selenocysteine and selenocystine were found to be present. Not surprisingly, chiral speciation revealed the presence of the l-enantiomeric forms of selenoamino acids in the sample. The major enantiomer found in the perchloric acid–ethanol extracts was l-MeSeCys, while in the enzymatic extracts l-SeMet was also detected.     

Trace element levels in honeys from different regions of Turkey

A survey of 25 honey samples from different botanical origin, collected all over the Turkey was conducted to assess their trace element contents. The aim of this study was to determine the levels of cadmium (Cd), lead (Pb), iron (Fe), manganese (Mn), copper (Cu), nickel (Ni), chromium (Cr), zinc (Zn), aluminium (Al) and selenium (Se) in honey samples from different regions of Turkey. Trace element contents were determined by a flame and graphite furnace atomic absorption spectrometry technique after dry-ashing, microwave digestion and wet-digestion. The accuracy of the method was corrected by the standard reference material, NIST-SRM 1515 Apple leaves. The contents of trace elements in honey samples were in the range of 0.23–2.41 μg g⁻¹, 0.32–4.56 μg g⁻¹, 1.1–12.7 μg g⁻¹, 1.8–10.2 μg g⁻¹, 8.4–105.8 μg kg⁻¹, 2.6–29.9 μg kg⁻¹, 2.4–37.9 μg kg⁻¹, 0.9–17.9 μg kg⁻¹, 83–325 μg kg⁻¹ and 38–113 μg kg⁻¹ for Cu, Mn, Zn, Fe, Pb, Ni, Cr, Cd, Al and Se, respectively. Iron was the most abundant element while cadmium was the lowest element in the Turkish honeys surveyed. The results showed that trace element concentrations in the honeys from different regions were generally correlated with the degree of trace element contamination of the environment.     

Identification and quantitative determination of glucosinolates in seeds and edible parts of Korean Chinese cabbage

Glucosinolates (GSLs) have attracted major interest due to the chemopreventive properties of some of their transformation products. GSLs in the seeds and edible parts of Korean Chinese cabbage (Brassicacampestris L. ssp. pekinensis) were identified and quantified by LC–ESI–MS and LC–UV. As a result, nine GSLs were identified: progoitrin, glucoraphanin, glucoalyssin, gluconapin, 4-hydroxy-3-indolylmethyl, glucobrassicanapin, glucoerucin, glucobrassicin, and 4-methoxyglucobrassicin. The total GSL levels were 268–198 and 23.0–15.8 μmol/g dry weight (DW) for seeds and edible parts, respectively. Gluconapin (197 μmol/g DW) was the highest individual GSL in seeds, whereas 4-methoxyglucobrassicin (6.08–4.94 μmol/g DW) and glucobrassicanapin (8.18–3.09 μmol/g DW) were found in the edible parts. In addition, LC–MS profiles of the nine GSLs identified from Korean Chinese cabbage were subjected to principal components analysis (PCA) to evaluate differences among samples. The metabolome among the four cultivar seed or edible parts was clearly separated by PCA.     

Simultaneous multi-channel hydride generation atomic fluorescence spectrometry determination of arsenic,bismuth, tellurium and selenium in tea leaves

A novel, rapid and sensitive method for the simultaneous multi-channel hydride generation atomic fluorescence spectrometry (HG-AFS) determination of total arsenic (As), total bismuth (Bi), total tellurium (Te) and total selenium (Se) in tea leaves was proposed. The operating parameters of self-made multi-channel HG-AFS were optimised, including negative high voltage of photo multiplier tube (PMT), the flow rates of carrier and shield gas, observation height and lamp currents. The conditions of hydride generation for As, Bi, Te and Se were studied in details. Under optimal conditions, the method detection limits (MDL) for As, Bi, Te and Se in tea leaves were 0.0152 μg g⁻¹, 0.0080 μg g⁻¹, 0.0022 μg g⁻¹ and 0.0068 μg g⁻¹, respectively. The proposed method was successfully applied to the simultaneous determination of As, Bi, Te and Se in various tea leaves and the spike recoveries were in the range of 90–103%. The accuracy of method was validated by analysing a tea certified reference material. The obtained values were consistent with the certified ones.     

Dispersive liquid–liquid microextraction for the determination of organochlorine pesticides residues in honey by gas chromatography-electron capture and ion trap mass spectrometric detection

A simple dispersive liquid–liquid microextraction (DLLME) protocol for the determination of 15 organochlorine pesticides residues in honey is proposed. The selected pesticides were separated using gas chromatography and detected by electron capture (ECD) or ion trap mass spectrometry (GC-IT/MS). Several parameters affecting the extraction efficiency namely type and volume of organic extraction solvent, type and volume of disperser solvent, sample pH, ionic strength, extraction time and centrifugation speed were systematically investigated. The final DLLME protocol involved the addition of 750 μL acetonitrile (disperser) and 50 μL chloroform (extraction solvent) into a 5 mL aqueous honey solution followed by centrifugation. The sedimented organic phase (chloroform) were analysed directly by GC-IT/MS or evaporated and reconstituted in acetonitrile prior to the GC-ECD analysis. The analytical performance of the GC-ECD and GC-IT/MS methods was compared and discussed. Under the selected experimental conditions, the enrichment factors varied between of 36 and 114. The limits of detection (LOD) were in the range of 0.02–0.15 μg L⁻¹ (0.4–3 ng g⁻¹) for GC-ECD and 0.01–0.2 μg L⁻¹ (0.2–4 ng g⁻¹) for GC-IT/MS which is adequate to verify compliance of products to legal tolerances. The proposed method was applied to the analysis of the selected organochlorine pesticides residues in various honey samples obtained from Greek region. Mean recoveries were ranged from 75% to 119% while the precision was better than 20% in both methodologies.     

Determination of minor carbohydrates in carrot (Daucus carota L.) by GC–MS

Minor carbohydrates present in carrot (Daucus carota L.) have been studied by GC–MS analysis of their trimethylsilyl derivatives because of their remarkable role in a variety of biological functions. Scyllo-inositol and sedoheptulose (d-altro-2-heptulose), identified for the first time in this paper were present in all the carrots analysed in concentrations ranging 1.5–5.8 and 1.4–24.6 mg g⁻¹ dried weight, respectively. Other minor carbohydrates detected in carrot were myo- inositol (2.2–9.8 mg g⁻¹) and mannitol (traces-1.3 mg g⁻¹). Whereas small amounts (close to 2 mg g⁻¹) of scyllo-inositol were experimentally determined in other vegetables from the Apiaceae family (parsley, coriander and fennel), sedoheptulose was only detected at trace levels.     

Residue levels of food-grade antioxidants in postharvest treated in-pod peanuts during five months of storage

In order to investigate residue levels of butylated hydroxyanisole (BHA), propyl paraben (PP) and butylated hydroxytoluene (BHT) during storage, eight-hundred kilograms of bulk peanuts were treated with the following antioxidant emulsions: BHA (1802 μg g⁻¹), BHA–PP (1802 μg g⁻¹ + 1802 μg g⁻¹) M1 and BHA–PP–BHT mixtures (1802 μg g⁻¹ + 901 μg g⁻¹ + 2204 μg g⁻¹) M2 and (1802 μg g⁻¹ + 1802 μg g⁻¹ + 2204 μg g⁻¹) M3. Residues were determined in peanut pod and seed tissues at 1-month intervals during the storage. While the reduction levels of BHA and PP in pods at the end of the storage period ranged from 66% to 76%, BHT levels were decreased extensively (86%). Twenty-four hours after peanuts were treated, antioxidant emulsions effectively seeped into the seeds and low levels of these chemicals were detected during the assay. Residues of PP in seeds were lower (62%) than the other antioxidants. Although the doses used were higher than those approved for food-grade antioxidants in stored peanuts, the residue levels in seeds (32.8–0.02 μg g⁻¹) did not exceed the maximum residue limits during the storage period.     

Validation of antibiotics in catfish by on-line solid phase extraction coupled to liquid chromatography tandem mass spectrometry

For the first time automated on-line solid phase extraction coupled to liquid chromatography tandem mass spectrometry was developed for the simultaneous determination of 13 antibiotics (sulfonamides and tetracyclines) in catfish. The method proposed was validated according to Commission Decision 2002/657/EC, showing good linearity between 2 and 350 μg kg⁻¹, high recovery (80–99%) and reproducibility (13–20%) values, lower detection limits than 0.1 μg kg⁻¹, and quantification limits under 2.4 μg kg⁻¹ (between 39 and 84 times lower than the MRL fixed by the EU). Moreover, the proposed method was also used to determine sulfonamides and tetracyclines in 16 out of 107 samples, all previously analysed by microbiological screening that gave positive results. Five out of 13 antibiotics were found, having tetracycline the higher occurrence (10 samples); in all cases the concentrations were lower than the MRL established.     

Selenium species in buckwheat cultivated with foliar addition of Se(VI) and various levels of UV-B radiation

Common (Fagopyrum esculentum Moench) and tartary (Fagopyrum tataricum Gaertn.) buckwheat was treated by spraying the leaves with a water solution containing 15 mg Se per litre in the form of sodium selenate in the flowering period. The selenium content in all parts of plant was found to be less than 200 ng g⁻¹ in non-treated and in the range 2700–4650 ng g⁻¹ in selenium treated buckwheat. Exposure to UV-B radiation lead to higher Se accumulation in flowers of both Se enriched cultivars. For speciation analysis enzymatic hydrolysis was carried out, separation and detection of selenium species was performed by high performance liquid chromatography–ultraviolet treatment–hydride generation atomic fluorescence spectrometry (HPLC–UV–HG-AFS). In flowers and leaves, on average 11% of the Se content was soluble and in the form of Se(VI), representing between 0.6% (flowers) and 3% (leaves) of the Se content. The remaining soluble non-amino acid organic Se was not detected by HPLC–UV–HG-AFS. In seeds 93% of the selenium content was found in the extracts and the main selenium species was SeMet with 93 ± 5% relative to the selenium content.     

Selenium species in leaves of chicory,dandelion, lamb’s lettuce and parsley

Lamb’s lettuce, dandelion, parsley and four cultivars of chicory were cultivated aeroponically for 41 days with nutrient solution containing 7  mg Se/L in the form of Na₂SeO₄. Se compounds were determined by high performance liquid chromatography–ultraviolet treatment–hydride generation atomic fluorescence spectrometry (HPLC–UV–HG-AFS) in the green parts of the selected plants. Se species were extracted by water and by enzymatic hydrolysis with Protease XIV. Separation of Se^IV, Se^VI, SeMet, SeMeSeCys and SeCys₂ was made by a combination of anion and cation exchange chromatography in which the columns were connected on-line to a UV–HG-AFS detection system. Se accumulated efficiently in plant leaves up to 480 μg/g dry mass, mostly as Se^VI, i.e. the form of Se in the nutrient solution. Beside inorganic Se, selenomethionine (6–21%), selenomethylselenocysteine (0.5–4.4%) and selenocistine (<DL-0.8%) were determined in the extracts after enzymatic hydrolysis. Some unidentified peaks were also observed in the chromatograms.     

Development of a fast multiresidue method for the determination of pesticides in dry samples (wheat grains,flour and bran) using QuEChERS based method and GC–MS

In this study a multiresidue method for the determination of 24 pesticides in wheat, white flour and bran using gas chromatography coupled to mass spectrometry with negative chemical ionisation and selected ion monitoring (GC–MS (NCI–SIM)) was developed and validated. The QuEChERS method was used for the extraction of different pesticides. The method was validated evaluating the following parameters: linearity, limit of detention, limit of quantification, matrix effect as well as precision and accuracy, evaluating the percentage of recovery at four different spike levels. The linear range used in the calibration curves was from 1.0 to 100 μg L⁻¹ for wheat and 2.0 to 200 μg L⁻¹ for flour and bran, both with values of r² > 0.99. The recoveries had been considered satisfactory presenting values between 70% and 120% with RSD < 20% for the majority of compounds.     

Gas chromatographic–mass spectrometric characterisation of tri- and tetrasaccharides in honey

A GC–MS method has been used to characterize tri- and tetrasaccharides in honey after their derivatization into trimethylsilyloxime derivatives. Based on retention data and mass spectra, a total of 25 trisaccharides were characterized; 12 being unequivocally identified using standards and two of them detected for the first time in honey. Erlose and panose were the major trisaccharides in the 12 honeys under analysis, their concentrations ranging 30–1214 mg 100 g⁻¹ of honey and 17–863 mg 100 g⁻¹ of honey, respectively. The GC–MS method also allowed the analysis of tetrasaccharides. Besides nystose, another nine tetrasaccharides were characterized; six of them were sucrose derivatives. Tetrasaccharides were present in concentrations lower than 230 mg 100 g⁻¹ of honey.     

Simultaneous derivatisation and preconcentration of parabens in food and other matrices by isobutyl chloroformate and dispersive liquid–liquid microextraction followed by gas chromatographic analysis

A simple, rapid and economical method has been proposed for the quantitative determination of parabens (methyl, ethyl, propyl and butyl paraben) in different samples (food, cosmetics and water) based on isobutyl chloroformate (IBCF) derivatisation and preconcentration using dispersive liquid–liquid microextraction in single step. Under optimum conditions, solid samples were extracted with ethanol (disperser solvent) and 200 μL of this extract along with 50 μL of chloroform (extraction solvent) and 10 μL of IBCF was rapidly injected into 2 mL of ultra-pure water containing 150 μL of pyridine to induce formation of a cloudy state. After centrifugation, 1 μL of the sedimented phase was analysed using gas chromatograph-flame ionisation detector (GC–FID) and the peaks were confirmed using gas chromatograph-positive chemical ionisation-mass spectrometer (GC–PCI–MS). Method was found to be linear over the range of 0.1–10 μg mL⁻¹ with square of correlation coefficient (R²) in the range of 0.9913–0.9992. Limit of detection (LOD) and limit of quantification (LOQ) were found to be 0.029–0.102 μg mL⁻¹ and 0.095–0.336 μg mL⁻¹ with a signal to noise ratio of 3:1 and 10:1, respectively.     

Solid phase extraction with flame atomic absorption spectrometry for determination of traces of Ca,K, Mg and Na in quality control of white sugar

A fast and straightforward pre-concentration procedure based on solid phase extraction with a strongly acidic cation-exchanger Dowex 50 W × 8–400 was proposed to determine traces of Ca, K, Mg and Na in white sugar samples by means of flame atomic absorption spectrometry. For this purpose, 20% (m/v) white sugar solutions (100 ml) were driven through resin beds at 10 ml min⁻¹ to retain Ca, K, Mg and Na ions and to separate sucrose that passed through unretained. Thereafter, columns were rinsed with water and elements of interest were recovered prior to measurements using 5 ml of a 2 mol l⁻¹ HCl solution. Detection limits of 0.04, 0.05, 0.02 and 0.01 μg g⁻¹ for Ca, K, Mg and Na, respectively, and precision of measurements within 1–3% were achieved. The proposed method enabled to determine Ca, K, Mg and Na in samples of white sugar within corresponding ranges: 0.66–0.99 μg g⁻¹ (Ca), 2.9–12.2 μg g⁻¹ (K), 0.53–1.57 μg g⁻¹ (Mg) and 0.06–0.30 μg g⁻¹ (Na). Accuracy of this sample pre-treatment procedure and analysis method was assessed by performing spikes and recovery experiments. Recoveries of added Ca, K, Mg and Na were found to be within 97–102%, demonstrating good reliability of results.     

Identification and quantification of trans fatty acids in bakery products by gas chromatography–mass spectrometry after focused microwave Soxhlet extraction

Trans fatty acids have been determined in fourteen bakery products using derivatisation by ester formation, gas chromatography–mass spectrometry for individual separation and identification/quantification following total fat isolation by Soxhlet extraction accelerated by focused microwave irradiation at the cartridge zone. The detection and quantification limits between 0.98 and 3.93 and 3.23–12.98 μg g⁻¹, respectively, and the linear dynamic ranges between LOQs values and 12,000 μg g⁻¹ thus obtained, demonstrated the utility of the approach for this type of analysis thanks to the wide determination range and high information level it provides. The proposed extraction method, which was validated by comparison with the Folch reference method – extraction under very mild conditions, shows that no changes of the original fat are produced under microwave-assisted extraction. The much shorter extraction time – 35 or 60 min vs. 3.5 h of the Folch method – and similar characteristics of the extract make this method an excellent alternative for the treatment of solid samples prior to trans fatty acids analysis. The target analytes were determined in fourteen bakery products; thus supporting the validity of the overall process.     

Distribution and migration study of pesticides between peel and pulp in grape by online gel permeation chromatography–gas chromatography/mass spectrometry

A multi-residue method for the analysis of 175 pesticides was developed by online gel permeation chromatography–gas chromatography/mass spectrometry (GPC–GC/MS) to study pesticide distribution and migration between peel and pulp in grape. The separated peel and pulp samples were extracted by acetonitrile after fortified with chlorpyrifos-d₁₀ isotope internal standard. The extract was first purified by solid phase distribution sorbent of primary secondary amine (PSA) and then detected by online GPC–GC/MS. At the spiking levels of 10, 50 and 200 μg kg⁻¹, 73.7%, 94.3% and 98.9% of the pesticides, respectively, presented recoveries between 70% and 120%. The ratios were 91.4%, 94.9% and 92.0%, respectively, for the relative standard deviations (RSDs) bellow 15%. Limits of detection (LODs) for the pesticides in pulp were below 10 μg kg⁻¹. Pesticides were separated to four groups according to the distribution ratios (peel/whole grape) of 100%, 80–99.9%, 50–80% and 0–50% in peel. Relationship between the pesticide distribution and corresponding regulation of EU maximum residue level (MRL) was discussed. Six factors influencing the pesticides distribution and migration between peel and pulp were discussed. Weak linear correlation between the pesticide solubility in water (20 °C) and the distribution ratios (lowest and average) in peel was found for most of the detected pesticides with solubility less than 200 mg L⁻¹.     

Determination of ultratrace levels of selenium in fruit and vegetable samples grown and consumed in Portugal

The selenium content in fruit and vegetable samples from two regions in Portugal were analysed using hydride generation atomic fluorescence spectrometry (HG-AFS) and radiochemical nuclear activation analysis (RNAA) – two analytical methods with very low limits of detection. The lower detection limits of HG-AFS, 3 μg kg⁻¹ and 8 μg kg⁻¹ (according to conditions used for digestion), and for RNAA, 10 μg kg⁻¹, meant that it was possible to determine selenium in samples previously analysed using the replicate sample instrumental nuclear activation analysis (RSINAA) with a higher detection limit associated.     

A validated matrix solid-phase dispersion method for the extraction of organophosphorus pesticides from bovine samples

A method based on matrix solid-phase dispersion (MSPD) was developed for the quantitative extraction of five organophosphorus (OPPs) pesticides from bovine samples. The determination was carried out by high performance liquid chromatography (HPLC) with diode array spectrophotometric UV detection. The MSPD extraction with octadecylsilyl (C18) sorbent combined with a silica gel clean-up and acetonitrile elution was optimised for chlorpyrifos, chlorfenvinphos, diazinon, fenitrothion, and parathion-methyl. The method was validated, yielding recovery values higher than 94%, except for chlorfenvinphos in liver (55%), and precision values, expressed as relative standard deviations (RSDs), which were less than or equal to 15% in liver and 11.5% in muscle at spiking levels of 0.25, 2.5 and 5 μg g⁻¹. Linearity was studied from 0.5 to 15 μg g⁻¹, and the limits of detection (LODs) were found to be lower than 0.1 μg g⁻¹_. This method was applied to the analysis of real samples with confirmative analyses performed using gas chromatography–mass spectrometry (GC–MS) in selected ion monitoring mode (SIM).