Brazil nuts have been classified as the foodstuffs that contain the highest level of unadulterated selenium, an essential trace element that appears to prevent cancer. To date, characterization of the selenium species in brazil nuts has not yet been investigated. In this work, various sample preparation approaches, including microwave extractions and enzymatic treatments, are examined with the goal of species preservation and subsequent selenium speciation; of these approaches, an enzymatic treatment with Proteinase K proved most effective. High-performance liquid chromatography (HPLC) separation strategies and inductively coupled plasma mass spectrometry (ICP-MS) detection schemes will also be presented. Extracts are evaluated against available standards for the commercially obtainable seleno-amino acids, selenomethionine (SeMet), selenoethionine (SeEt), and selenocystine (SeCys); selenomethionine was demonstrated to be the most abundant of these seleno-amino acids. Further characterization of unidentified selenium-containing peaks is attempted by the employment of several procedures, including electrospray-mass spectrometry (ES-MS). A peptide structure was identified; however, this was considered a tentative proposal due to the large background produced by the extremely complicated brazil nut matrix.
SUMMARY Biological systems contain highly-ordered macromolecular structures with diverse functions, inspiring their utilization in nanotechnology. A motor allows linear dsDNA viruses to package their genome into a preformed procapsid. The central component of the motor is the portal connector that acts as a pathway for the translocation of dsDNA. The elegant design of the connector and its channel motivates its application as an artificial nanopore. Herein, we demonstrate the robust characteristics of the connector of the bacteriophage phi29 DNA packaging motor by single pore electrophysiological assays. The conductance of each pore is almost identical and is perfectly linear with respect to the applied voltage. Numerous transient current blockade events induced by dsDNA are consistent with the dimensions of the channel and dsDNA. Furthermore, the connector channel is stable under a wide range of experimental conditions including high salt and pH 2–12. The robust properties of the connector nanopore made it possible to develop a simple reproducible approach for pore quantifications. Such quantifications led to a reliable real time counting of DNA passing through the channel. The fingerprint of DNA translocation in this system has provided a new tool for future biophysical and physicochemical characterizations of DNA transportation, motion, and packaging.
In addition to determination of total selenium in nuts, the element distribution among different fractions (lipid extract, low molecular weight, and protein fractions), and speciation analysis were studied. Improved precision for total selenium determination was observed after elimination of lipids. Because selenium was not detected in any of the lipid extracts obtained from the different types of nuts (ICP-MS), in each determination and/or speciation procedure used in this work lipids were extracted (chloroform-methanol, 2:1) and discarded before analysis. In agreement with previously reported data, high selenium levels were found in Brazil nuts (those purchased without shells contained approximately a quarter the content than those purchased with shells) and significantly lower levels in walnuts, cashews, and pecans nuts. Low-molecular-weight compounds were extracted with perchloric acid (0.4 mol L(-1)) to furnish a fraction containing 3 to 15% of the total selenium in different types of nuts. The proteins were isolated from nut samples by dissolution in 0.1 mol L(-1) sodium hydroxide and subsequent precipitation with acetone. They were then dissolved in phosphate buffer pH 7.5. Analysis of protein fractions focused on selenium in two possible states - weakly and firmly bound to proteins. Results obtained for Brazil nuts by size-exclusion chromatography with on-line ICP-MS detection, in the absence and in the presence of beta-mercaptoethanol, showed that approximately 12% of total selenium was weakly bound to proteins. To obtain information about firmly bound selenium, the protein extracts were hydrolyzed enzymatically with proteinase K. Speciation was performed by means of ion-pairing HPLC-ICP-MS. The primary species found in all types of nuts was Se-methionine (19-25% of total selenium for different types of nuts).
Polybrominated diphenyl ethers (PBDEs) are potentially harmful and persistent environmental pollutants. Despite evidence that soils are a major sink for PBDEs, little is known regarding their behavior in this medium. An environmentally relevant level of a commercial penta-BDE mixture (75 μg kg -1 ) was added to topsoil and the extractability of three congeners (BDE 47, 99, and 100) was monitored over 10 weeks in planted and unplanted treatments. The extractability of each congener decreased rapidly in the experimental soil due largely to abiotic sorption to soil particles, which was demonstrated by low PBDE recovery from sterilized and dry soils. Monoculture plantings of zucchini and radish did not affect the recovery of PBDEs from soil. However, PBDE recovery from mixed species plantings was nearly 8 times higher than that of unplanted and monoculture treatments, indicating that interspecific plant interactions may enhance PBDE bioavailablity in soil. Evidence for competitive interactions between the two species was revealed by reduced shoot biomass of zucchini plants in mixed treatments relative to pots containing only zucchini. Both plant species accumulated PBDEs in root and shoot tissue (< 5 μg kg -1 plant tissue). PBDE uptake was higher in zucchini and translocation of PBDEs to zucchini shoots was congener-specific. Our results suggest that although abiotic sorption may limit the potential for human exposure to PBDEs in soil, plants may increase the exposure risk by taking up and translocating PBDEs into aboveground tissues and by enhancing bioavailability in soil.
The occurrence and form of selenium and mercury were investigated in Indian Mustard, Brassica juncea, a selenium accumulating plant, which had been co-exposed to varying concentration levels of these two elements. Plants were grown and exposed in hydroponic solutions. Following exposure, root exudates were collected in fresh solutions and the head-space around the aerial portions of the plants was sampled. These samples and the harvested plant tissues were then processed for determination of Se and Hg-containing compounds. For the plant tissues, roots, stems and leaves were separated and extracted using a sequential procedure that removed watersoluble species, water-soluble proteins, and dodecyl sulfate-soluble proteins. Size exclusion chromatography allowed further fractionation. High molecular-weight selenium/mercurycontaining compounds were found primarily in the plant root extract. Evidence suggests that a Se and Hg complex of high molecular weight may be protein associated. For the analysis of exudate solutions, ion-pairing reversed phase chromatography coupled to ICP-MS was used. Multiple selenium and mercury species were detected, with one mercury-containing compound observed eluting near selenocystine. Plant head-space was sampled with solid phase microextraction and analyzed with GC-ICP-MS and GC-TOFMS. Apart from the primary selenium volatiles and elemental mercury, no volatile species simultaneously containing Se and Hg could be detected.
Sr appears as a radionuclide in the decay series of nuclear fission and can therefore be found in nuclear waste or released by nuclear accidents. Current methods for the detection of this radionuclide are time consuming and may be prone to a large variety of interferents. In this work, inductively coupled plasma mass spectrometry is explored for the determination of 90 Sr in the presence of stable zirconium in urine. Specific techniques are investigated to remove this as well as other contributions to the background at m/z~90. A quadrupole ICP-MS equipped with a hexapole collision cell is first explored (final LOD~2 ng L 21 for water samples), however, the desired limit of detection for 90 Sr in urine is quite low (0.02 pg L 21). The performance of a double-focusing sector field ICP mass spectrometer (ICP-SFMS) is further investigated, which allows measurement of 90 Sr at the ultratrace level. Other potential interferences were investigated and instrumental detection limits are calculated as 3 pg L 21 for water samples. Final parameters include the use of a cool plasma and medium mass resolution in ICP-SFMS. The method is applied to the analysis of 90 Sr extracted from urine using a crown ether extraction resin and concentrated (enrichment factor: 200); high levels of natural strontium in the separated fraction (of about 1 mg mL 21 ) equate to higher detection limits (80 pg L 21) due to 88 Sr 1 at m/z~90 and the relatively low abundance sensitivity of ICP-SFMS at medium mass resolution of 6 6 10 27 . This detection limit in the separated fraction corresponds to the detection limit of 0.4 pg L 21 in the original urine sample. The recovery of 90 Sr, determined with the developed analytical method in spiked urine samples, was in the range of 82-86%.
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