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Cobalt is an important element with magnetic properties used in various industrial applications, but is also needed for biological activity. Very little is known about the cellular response of living systems to cobalt stress. Towards investigating this mechanism, we isolated individual Saccharomyces cerevisiae cells resistant to high cobalt concentrations up to 8 mmoll(-1), by employing four different 'in vivo' evolutionary engineering strategies: selection under constant or gradually increasing stress levels, and selection under continuous or pulse exposure to cobalt stress. Selection under continuous exposure to gradually increasing cobalt stress levels yielded the most resistant cell population to cobalt. However, the resistance was highly heterogeneous within the mutant populations ranging from 3- to 3700-fold survival rate of isolated individuals to 8 mmoll(-1) CoCl2 in the most resistant population. Moreover, cobalt-resistant individual colonies were associated with 2-4-times lower intracellular cobalt contents as compared to wild-type, and with cross-resistance to metals such as nickel, zinc, manganese, but not to copper and chromium ions. Contrary to mutants evolved under continuous exposure to cobalt, those isolated by pulse exposure strategy also exhibited resistance to heat shock and hydrogen peroxide stress. Taken together, this study reinforced the fact that evolutionary engineering is useful in selecting strains with very specific phenotypes, and further illustrated the importance of the strategy chosen to isolate the best evolved strain.

A method has been developed for identification and quantification of the acaricide amitraz and its transformation products, 2,4-dimethylaniline (DMA), 2,4-dimethylformamidine (DMF) and N-2,4-dimethylphenyl-N-methylformamidine (DMPF) in pears. The analytes were extracted using ethyl acetate and anhydrous sodium sulphate. Analysis was performed by liquid chromatography-electrospray tandem mass spectrometry (LC-ESI-MS/MS) in the positive ion mode using a triple quadrupole (QqQ) instrument. Two precursor-product ion transitions were monitored for each compound in the selected reaction monitoring (SRM) mode. The method was validated with pears taken from the orchard before the amitraz treatment and spiked at the limit of quantification (LOQ), 10 times the LOQ and the maximum residue limit (MRL). Recoveries were between 70 and 106% and relative standard deviations were below 19% (n=5 at each spiked level). Excellent sensitivity resulted in limits of detection (LODs) for all the compounds below 10 microg kg(-1). Quantification was carried out using matrix-matched standards calibration, response was a linear function of the concentration from the LOQs to, at least, three orders of magnitude. Recoveries and standard deviations were comparable to those obtained after hydrolysis of amitraz and its metabolites to DMA. Occurrence of amitraz and its metabolites in field-treated pears showed that, seven days after the treatment, DMPF and DMF are the main degradation products. This work reports for the first time the use of a conventional pesticide multiresidue method and LC-ESI-MS/MS for determining amitraz and its metabolites in pears.

Liquid chromatography under high pressure in combination with quadrupole time-of-flight mass spectrometry (QqTOF-MS and MS/MS) has been used to detect amitraz degradation products in pears, to characterize their structures, and to evaluate their occurrences in samples of different origins. Using the proposed approach, the parent pesticide and four degradation products were identified. To this end, pear samples were extracted with ethyl acetate and anhydrous sodium sulphate. Amitraz was found to be rapidly decomposed into four related compounds, of which N-(2,4-dimethylphenyl)formamidine (DMPF) was the most abundant and persistent. N,N'-bisdimethylphenylformamidine (BDMPF), 2,4-dimethylformamidine (DMF) and 2,4-dimethyl aniline (DMA) were also main metabolites of amitraz. To our knowledge this is the first report that confirms the presence of BDMPF in pears. The method was validated using MS and MS/MS for those standard available (analytical or not). In MS, recoveries ranged from 83 to 101% with relative standard deviation (RSD) from 9 to 19% at the limit of quantification (LOQ) (between 5 and 20 microg kg(-1)). Using MS/MS, recoveries, linearity and precision were similar but LOQs were higher because the intense fragmentation of the protonated molecules in the product mass spectrum. BDMPF, as an approximation, was quantified based on the DMF metabolite. The results demonstrated that high-pressure LC-QqTOF-MS and MS/MS techniques enhance further the capabilities of LC-MS in the identification of polar species in complex food samples.

The formation of nanometer-sized gaps between silver nanoparticles is critically important for optimal enhancement in surface-enhanced Raman scattering (SERS). A simple approach is developed to generate nanometer-sized cavities in a silver nanoparticle thin film for use as a SERS substrate with extremely high enhancement. In this method, a submicroliter volume of concentrated silver colloidal suspension stabilized with cetyltrimethylammonium bromide (CTAB) is spotted on hydrophobic glass surfaces prepared by the exposure of the glass to dichloromethysilane vapors. The use of a hydrophobic surface helps the formation of a more uniform silver nanoparticle thin film, and CTAB acts as a molecular spacer to keep the silver nanoparticles at a distance. A series of CTAB concentrations is investigated to optimize the interparticle distance and aggregation status. The silver nanoparticle thin films prepared on regular and hydrophobic surfaces are compared. Rhodamine 6G is used as a probe to characterize the thin films as SERS substrates. SERS enhancement without the contribution of the resonance of the thin film prepared on the hydrophobic surface is calculated as 2 x 10(7) for rhodamine 6G, which is about one order of magnitude greater than that of the silver nanoparticle aggregates prepared with CTAB on regular glass surfaces and two orders of magnitude greater than that of the silver nanoparticle aggregates prepared without CTAB on regular glass surfaces. A hydrophobic surface and the presence of CTAB have an increased effect on the charge-transfer component of the SERS enhancement mechanism. The limit of detection for rhodamine 6G is estimated as 1.0 x 10(-8) M. Scanning electron microscopy and atomic force microscopy are used for the characterization of the prepared substrate.

In this study, in order to eliminate the drawbacks of elution step and to reach higher enrichment factors, a novel preconcentration/separation technique for the slurry analysis of sorbent loaded with lead prior to its determination by electrothermal atomic absorption spectrometry was described. For this purpose, at first, lead was collected on ethylene glycol dimethacrylate methacrylic acid copolymer (EGDMA-MA) treated with ammonium pyrolidine dithiocarbamate (APDC) by conventional batch technique. After separation of liquid phase, slurry of the sorbent was prepared and directly pipetted into graphite furnace of atomic absorption spectrophotometer. Optimum conditions for quantitative sorption and preparation of the slurry were investigated. A 100-fold enrichment factor could be easily reached. The analyte element in certified sea-water and Bovine-liver samples was determined in the range of 95% confidence level. The proposed technique was fast and simple and the risks of contamination and analyte loss were low. Detection limit (3delta) for Pb was 1.67 microg l(-1).

Manganese and copper in multivitamin-mineral supplements and standard reference materials were determined by slurry sampling electrothermal atomic absorption spectrometry. Slurries were prepared in an aqueous solution containing Triton X-100. The effects of different parameters such as ratio of solid to liquid phase volume, total slurry volume and addition of Triton X-100 as a dispersant on the analytical results were investigated. The graphite furnace programs were optimized for slurry sampling depending on the analytes and their concentrations in the samples. The linear calibration method with aqueous standard solutions was used for the quantification. At optimum experimental conditions, R.S.D. values were below 5%. The analytes were determined in the limits of 95% confidence level with respect to certified values in coal and soil standard reference materials and to those found by wet-digestion in multivitamin-mineral supplements. Detection limits (3delta) for Mn and Cu were 0.10 microg L(-1) and 1.82 microg L(-1) for 10 microL coal standard reference material slurry, respectively.

In this study, a water-soluble polymer, polyvinylpyrrolidinone (PVP) having chelating functionalities was used for the preconcentration and separation of traces of Pb, Cu, Ve and Mn prior to their determination by graphite furnace atomic absorption spectrometry. For this purpose, the sample and the PVP solutions were mixed and the metal bound polymer was precipitated by adding the mixture onto acetone. The precipitate was separated by decantation and dissolved with water. By increasing the ratio of the volumes of sample to water used in dissolving the precipitate, the analyte elements were concentrated as needed. The concentration of trace elements was determined using graphite furnace atomic absorption spectrometry. The analyte elements in matrix free aqueous solutions were quantitatively recovered. The validity of the proposed method was checked with a standard reference material (NIST SRM 1577b bovine liver) and spiked fruit juice, sea water and mineral water samples. The analytical results were found to be in good agreement with certified and added values. Detection limits (3delta) were 1.7, 3.6 and 4.1mugl(-1) for Pb, Cu and Mn, respectively, using 10mul of sample volume. The method is novel and can be characterized by rapidity, simplicity, quantitative recovery and high reproducibility.

Lead and nickel were concentrated and separated after sorbing on Chromosorb-107 filled in a syringe prior to their determination by electrothermal atomic absorption spectrometry. To retain the analytes, the sample solution treated with or without ammonium pyrolidine dithiocarbamate (APDC) was drawn into the syringe filled with Chromosorb-107 and discharged back manually. Then the analyte elements were eluted by drawing and discharging the suitable eluent. The optimum experimental conditions for quantitative recoveries of analytes (amount of sorbent, pH of sample, concentration and kind of eluent, flow rates of sample and eluent and adding of the effect of complexing agent) were investigated. Nickel was quantitatively retained at pH6 irrespective of whether it was complexed with APDC while the quantitative sorption of lead was achieved at pH8 only if it was treated with APDC prior to passing through the sorbent. The lead and nickel retained on the sorbent were eluted drawing and discharging 4.5 M HNO(3), respectively. If the concentration of analyte elements in the sample were too low to be detected, then they were concentrated by increasing the ratio of sample value to eluent volume as needed. The lead and nickel in spiked sea-water samples and in certified reference Apple-Leaves (NIST SRM 1515) standards were quantitatively (95%) recovered with R.S.D. of around +/-2%.

In this study, the use of syringe filled with sorbent for the separation and enrichment of bismuth, lead and nickel prior to their analysis by graphite furnace atomic absorption spectrometry was described to substitute for batch and column techniques. The method proposed in this paper was compared with column technique with respect to easiness, fastness, simplicity, recovery and risk of contamination. The syringe was filled with 0.5 g of sorbent and in order to retain the analyte elements, 5 ml of sample solution (pH>/=5) was drawn into the syringe to 15 s and discharged again in 15 s. Then, 2.0 M of HCl, as the eluent, was drawn into the syringe and ejected back to desorb the analyte elements. At optimum conditions, the recoveries of Bi, Pb and Ni were 95-99% with relative standard deviations (RSDs) of around +/-2%. Detection limit (delta) was 0.5 mug l(-1) for Bi, Pb and Ni, respectively. The elements could be concentrated by drawing and discharging several portions of sample successively but eluting only one time. Bi, Pb and Ni added to a seawater sample were quantitatively recovered (>95%) with low RSD values of around +/-2-3%. The risk of contamination is less than that with the column technique. In addition, it is much faster, simpler, easier, more practical and handy compared with column technique.

In this study, a syringe was filled with silica gel loaded with 3-aminopropyltriethoxysilane, for the separation and preconcentration of copper, cadmium and chromium prior to their determination by graphite furnace atomic absorption spectrometry (GFAAS) in seawater. For this purpose, a syringe was filled with 0.5 g of modified silica gel and the sample solution was drawn into the syringe and ejected back again. The analyte elements were quantitatively retained at pH 5. Then, the elements sorbed by the silica gel were eluted with 2.0 M of HCl and determined by GFAAS. At optimum conditions, the recovery of Cu, Cd and Cr were 96-98%. Detection limits (3delta) were 6.6, 7.5 and 6.0 micro g L(-1) for Cu, Cd and Cr, respectively. The elements could be concentrated by drawing and discharging several portions of sample successively but eluting only once. Cu, Cd and Cr added to a seawater sample were quantitatively recovered (>95%) in the range of the 95% confidence level. The method proposed in this paper was compared with a column technique. Optimum experimental conditions, reproducibility, precision and recoveries of both techniques are the same, but the syringe technique is much faster, easier and more practical than the column technique. It is a portable system and allows one to make the sorption process in the source of sample. In addition, the risk of contamination is less than in the column technique.