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Analytical and Bioanalytical Chemistry (v.383, #1)
Quality Assurance Challenge 2 by Manfred Reichenbächer; Jürgen W. Einax (pp. 1-2).
We would like to invite you to participate in the Analytical Challenge, which is a series of puzzles designed to entertain and challenge our readers. The special ABC feature ‘Analytical Challenge’ has established itself as a truly unique quiz series, with a new scientific puzzle published every second month. Readers can access the complete collection of published problems with their solutions on the ABC homepage at www.springeronline.com/journal/00216. Test your knowledge and tease your wits in diverse areas of analytical and bioanalytical chemistry by viewing this collection. Quality assurance is the topic of the present challenge. And please note: there is a prize to be won (a Springer book from our catalogue up to a value of € 75,–). Please read on...
Quality Assurance Challenge 2 by Manfred Reichenbächer; Jürgen W. Einax (pp. 1-2).
We would like to invite you to participate in the Analytical Challenge, which is a series of puzzles designed to entertain and challenge our readers. The special ABC feature ‘Analytical Challenge’ has established itself as a truly unique quiz series, with a new scientific puzzle published every second month. Readers can access the complete collection of published problems with their solutions on the ABC homepage at www.springeronline.com/journal/00216. Test your knowledge and tease your wits in diverse areas of analytical and bioanalytical chemistry by viewing this collection. Quality assurance is the topic of the present challenge. And please note: there is a prize to be won (a Springer book from our catalogue up to a value of € 75,–). Please read on...
Surface complexation studied via combined grazing-incidence EXAFS and surface diffraction: arsenate on hematite (0001) and (10–12) by G. Waychunas; T. Trainor; P. Eng; J. Catalano; G. Brown; J. Davis; J. Rogers; J. Bargar (pp. 12-27).
X-ray diffraction [crystal-truncation-rod (CTR)] studies of the surface structure of moisture-equilibrated hematite reveal sites for complexation not present on the bulk oxygen-terminated surface, and impose constraints on the types of inner-sphere sorption topologies. We have used this improved model of the hematite surface to analyze grazing-incidence EXAFS results for arsenate sorption on the c (0001) and r (10–12) surfaces measured in two electric vector polarizations. This work shows that the reconfiguration of the surface under moist conditions is responsible for an increased adsorption density of arsenate complexes on the (0001) surface relative to predicted ideal termination, and an abundance of “edge-sharing” bidentate complexes on both studied surfaces. We consider possible limitations on combining the methods due to differing surface sensitivities, and discuss further analysis possibilities using both methods.
Keywords: Crystal truncation rod; GIXAFS; EXAFS; Surface structure; Hematite; Arsenate
Surface complexation studied via combined grazing-incidence EXAFS and surface diffraction: arsenate on hematite (0001) and (10–12) by G. Waychunas; T. Trainor; P. Eng; J. Catalano; G. Brown; J. Davis; J. Rogers; J. Bargar (pp. 12-27).
X-ray diffraction [crystal-truncation-rod (CTR)] studies of the surface structure of moisture-equilibrated hematite reveal sites for complexation not present on the bulk oxygen-terminated surface, and impose constraints on the types of inner-sphere sorption topologies. We have used this improved model of the hematite surface to analyze grazing-incidence EXAFS results for arsenate sorption on the c (0001) and r (10–12) surfaces measured in two electric vector polarizations. This work shows that the reconfiguration of the surface under moist conditions is responsible for an increased adsorption density of arsenate complexes on the (0001) surface relative to predicted ideal termination, and an abundance of “edge-sharing” bidentate complexes on both studied surfaces. We consider possible limitations on combining the methods due to differing surface sensitivities, and discuss further analysis possibilities using both methods.
Keywords: Crystal truncation rod; GIXAFS; EXAFS; Surface structure; Hematite; Arsenate
Combined LIBD and XAFS investigation of the formation and structure of Zr(IV) colloids by Hye-Ryun Cho; Clemens Walther; Jörg Rothe; Volker Neck; Melissa A. Denecke; Kathy Dardenne; Thomas Fanghänel (pp. 28-40).
The solubility of Zr(OH)4(am)—in other words hydrated Zr(IV) oxyhydroxide—is determined by means of coulometric titration (CT), and colloids are detected by laser-induced breakdown when the solubility limit is exceeded. Our results at pH 3–8 demonstrate that the solubility of Zr(OH)4(am) is several orders of magnitude higher than reported classical solubility data for acidic solutions, determined from undersaturation with a less soluble microcrystalline Zr(IV) oxide precipitate. Analysis of extended X-ray absorption fine structure (EXAFS) data shows that the microcrystalline colloids in a 0.1 mol l−1 Zr aqueous solution at pH 0.2 contain tetrameric units, similar to those present in the structure of ZrOCl2.8H2O. Characterization of the CT solutions by means of EXAFS shows that oligomeric species form as the solubility limit is approached. The current lack of data on equilibrium constants for polynuclear hydroxide complexes prohibits the use of a realistic speciation model to describe the solubility of pH-dependent Zr(OH)4(am). However, the solubility curve is obtained using the mononuclear hydrolysis constants estimated in the present paper, along with the solubility constant (logK′sp=−49.9±0.5 in 0.5 mol l−1 NaCl; logK°sp=−53.1±0.5 at I=0).
Keywords: LIBD; Colloids; EXAFS; Zirconium; Hydrolysis
Combined LIBD and XAFS investigation of the formation and structure of Zr(IV) colloids by Hye-Ryun Cho; Clemens Walther; Jörg Rothe; Volker Neck; Melissa A. Denecke; Kathy Dardenne; Thomas Fanghänel (pp. 28-40).
The solubility of Zr(OH)4(am)—in other words hydrated Zr(IV) oxyhydroxide—is determined by means of coulometric titration (CT), and colloids are detected by laser-induced breakdown when the solubility limit is exceeded. Our results at pH 3–8 demonstrate that the solubility of Zr(OH)4(am) is several orders of magnitude higher than reported classical solubility data for acidic solutions, determined from undersaturation with a less soluble microcrystalline Zr(IV) oxide precipitate. Analysis of extended X-ray absorption fine structure (EXAFS) data shows that the microcrystalline colloids in a 0.1 mol l−1 Zr aqueous solution at pH 0.2 contain tetrameric units, similar to those present in the structure of ZrOCl2.8H2O. Characterization of the CT solutions by means of EXAFS shows that oligomeric species form as the solubility limit is approached. The current lack of data on equilibrium constants for polynuclear hydroxide complexes prohibits the use of a realistic speciation model to describe the solubility of pH-dependent Zr(OH)4(am). However, the solubility curve is obtained using the mononuclear hydrolysis constants estimated in the present paper, along with the solubility constant (logK′sp=−49.9±0.5 in 0.5 mol l−1 NaCl; logK°sp=−53.1±0.5 at I=0).
Keywords: LIBD; Colloids; EXAFS; Zirconium; Hydrolysis
Soft X-ray scanning transmission X-ray microscopy (STXM) of actinide particles by Hans J. Nilsson; Tolek Tyliszczak; Richard E. Wilson; Lars Werme; David K. Shuh (pp. 41-47).
A descriptive account is given of our most recent research on the actinide dioxides with the Advanced Light Source Molecular Environmental Science (ALS-MES) Beamline 11.0.2 soft X-ray scanning transmission X-ray microscope (STXM) at the Lawrence Berkeley National Laboratory (LBNL). The ALS-MES STXM permits near-edge X-ray absorption fine structure (NEXAFS) and imaging with 30-nm spatial resolution. The first STXM spectromicroscopy NEXAFS spectra at the actinide 4d5/2 edges of the imaged transuranic particles, NpO2 and PuO2, have been obtained. Radiation damage induced by the STXM was observed in the investigation of a mixed oxidation state particle (Np(V,VI)) and was minimized during collection of the actual spectra at the 4d5/2 edge of the Np(V,VI) solid. A plutonium elemental map was obtained from an irregular PuO2 particle with the dimensions of 650 × 650 nm. The Pu 4d5/2 NEXAFS spectra were collected at several different locations from the PuO2 particle and were identical. A representative oxygen K-edge spectrum from UO2 was collected and resembles the oxygen K-edge from the bulk material. The unique and current performance of the ALS-MES STXM at extremely low energies (ca. 100 eV) that may permit the successful measurement of the actinide 5d edge is documented. Finally, the potential of STXM as a tool for actinide investigations is briefly discussed.
Keywords: Actinide; STXM; NEXAFS; Spectromicroscopy; Plutonium; Neptunium
Soft X-ray scanning transmission X-ray microscopy (STXM) of actinide particles by Hans J. Nilsson; Tolek Tyliszczak; Richard E. Wilson; Lars Werme; David K. Shuh (pp. 41-47).
A descriptive account is given of our most recent research on the actinide dioxides with the Advanced Light Source Molecular Environmental Science (ALS-MES) Beamline 11.0.2 soft X-ray scanning transmission X-ray microscope (STXM) at the Lawrence Berkeley National Laboratory (LBNL). The ALS-MES STXM permits near-edge X-ray absorption fine structure (NEXAFS) and imaging with 30-nm spatial resolution. The first STXM spectromicroscopy NEXAFS spectra at the actinide 4d5/2 edges of the imaged transuranic particles, NpO2 and PuO2, have been obtained. Radiation damage induced by the STXM was observed in the investigation of a mixed oxidation state particle (Np(V,VI)) and was minimized during collection of the actual spectra at the 4d5/2 edge of the Np(V,VI) solid. A plutonium elemental map was obtained from an irregular PuO2 particle with the dimensions of 650 × 650 nm. The Pu 4d5/2 NEXAFS spectra were collected at several different locations from the PuO2 particle and were identical. A representative oxygen K-edge spectrum from UO2 was collected and resembles the oxygen K-edge from the bulk material. The unique and current performance of the ALS-MES STXM at extremely low energies (ca. 100 eV) that may permit the successful measurement of the actinide 5d edge is documented. Finally, the potential of STXM as a tool for actinide investigations is briefly discussed.
Keywords: Actinide; STXM; NEXAFS; Spectromicroscopy; Plutonium; Neptunium
Determination of actinide speciation in solution using high-energy X-ray scattering by L. Soderholm; S. Skanthakumar; J. Neuefeind (pp. 48-55).
High-energy X-ray scattering (HEXS) has been used to understand the coordination environment of the uranyl ion in a perchlorate solution. Assuming the two coordinating oxo ligands bound to U(VI) are represented in a peak in the pair distribution function (PDF) at 1.766(1) Å, integration of the peak intensity is used to quantify the charge located on the oxygens. The dioxo ligands are essentially neutral, as predicted by numerous published calculations, with a charge of −16.4(8) electrons. The peak in the PDF at 2.420(1) Å is consistent with equatorial ligating waters. The intensity of this peak is inconsistent with an integral coordination number and is used to propose a solution equilibrium of five and four waters coordinating to the uranyl(VI) ion favoring the five-coordinate species. This equilibrium is then used to experimentally determine that five-coordinate uranyl is 1.19±0.42 kcal/mol more stable than its four-coordinate counterpart under the conditions of the experiment. Further peaks in the Fourier transform of the scattering data at 4.50, 7, and 8.7 Å are attributed to uranium–solvent correlations.
Keywords: X-ray scattering; Solution speciation; Uranyl ion; Synchrotron studies; Actinide ion; Uranium
Determination of actinide speciation in solution using high-energy X-ray scattering by L. Soderholm; S. Skanthakumar; J. Neuefeind (pp. 48-55).
High-energy X-ray scattering (HEXS) has been used to understand the coordination environment of the uranyl ion in a perchlorate solution. Assuming the two coordinating oxo ligands bound to U(VI) are represented in a peak in the pair distribution function (PDF) at 1.766(1) Å, integration of the peak intensity is used to quantify the charge located on the oxygens. The dioxo ligands are essentially neutral, as predicted by numerous published calculations, with a charge of −16.4(8) electrons. The peak in the PDF at 2.420(1) Å is consistent with equatorial ligating waters. The intensity of this peak is inconsistent with an integral coordination number and is used to propose a solution equilibrium of five and four waters coordinating to the uranyl(VI) ion favoring the five-coordinate species. This equilibrium is then used to experimentally determine that five-coordinate uranyl is 1.19±0.42 kcal/mol more stable than its four-coordinate counterpart under the conditions of the experiment. Further peaks in the Fourier transform of the scattering data at 4.50, 7, and 8.7 Å are attributed to uranium–solvent correlations.
Keywords: X-ray scattering; Solution speciation; Uranyl ion; Synchrotron studies; Actinide ion; Uranium
Three-dimensional modeling of EXAFS spectral mixtures by combining Monte Carlo simulations and target transformation factor analysis by Andre Rossberg; Andreas C. Scheinost (pp. 56-66).
We have developed a new method for the three-dimensional modeling of extended X-ray absorption fine structure (EXAFS) spectra which enables the extraction of the local structure of aqueous metal complexes from spectral mixtures of several components. The new method combines two techniques: Monte Carlo simulation and target transformation factor analysis (TFA). Monte Carlo simulation is used to create random arrangements between the X-ray absorbing metal ion and the ligand atoms, and to calculate the theoretical EXAFS spectrum of each arrangement. The theoretical EXAFS spectrum is then introduced as test spectrum in the TFA procedure, to test whether or not the test spectrum is likely to be a component of the spectral mixtures. This coupled procedure is repeated until the error in the test spectrum is minimized. The new method can thus be used to isolate and refine the structure of complexes from spectral mixtures and to determine their relative concentrations, solely on the basis of an estimate of a ligand structure. The performance of the proposed method is validated using uranium Liii-edge EXAFS spectra of binary mixtures of two uranium(VI) 3,4-dihydroxybenzoic acid complexes.
Keywords: EXAFS; Complexation; Speciation; Factor analysis; Monte Carlo; Multiple scattering
Three-dimensional modeling of EXAFS spectral mixtures by combining Monte Carlo simulations and target transformation factor analysis by Andre Rossberg; Andreas C. Scheinost (pp. 56-66).
We have developed a new method for the three-dimensional modeling of extended X-ray absorption fine structure (EXAFS) spectra which enables the extraction of the local structure of aqueous metal complexes from spectral mixtures of several components. The new method combines two techniques: Monte Carlo simulation and target transformation factor analysis (TFA). Monte Carlo simulation is used to create random arrangements between the X-ray absorbing metal ion and the ligand atoms, and to calculate the theoretical EXAFS spectrum of each arrangement. The theoretical EXAFS spectrum is then introduced as test spectrum in the TFA procedure, to test whether or not the test spectrum is likely to be a component of the spectral mixtures. This coupled procedure is repeated until the error in the test spectrum is minimized. The new method can thus be used to isolate and refine the structure of complexes from spectral mixtures and to determine their relative concentrations, solely on the basis of an estimate of a ligand structure. The performance of the proposed method is validated using uranium Liii-edge EXAFS spectra of binary mixtures of two uranium(VI) 3,4-dihydroxybenzoic acid complexes.
Keywords: EXAFS; Complexation; Speciation; Factor analysis; Monte Carlo; Multiple scattering
Characterization of rat liver microsomal and hepatocytal metabolites of brevetoxins by liquid chromatography–electrospray tandem mass spectrometry by Weiqun Wang; Yousheng Hua; Guangdi Wang; Richard B. Cole (pp. 67-75).
Brevetoxins are natural neurotoxins that are produced by “red tide” algae. This class of compounds can cause neurotoxic shellfish poisoning and other health problems. Brevetoxin-2 is the most abundant among the nine brevetoxins that have been characterized, whereas brevetoxin-1 is the most toxic. In this study, brevetoxin-1 and brevetoxin-2 were incubated with rat liver hepatocytes and rat liver microsomes, respectively. After clean-up steps were taken to remove the proteins, samples were analyzed by liquid chromatography (LC) coupled with electrospray mass spectrometry (LC-MS). After incubation of brevetoxin-1, two metabolites were found: brevetoxin-1-M1 (molecular weight = 900 Da), and brevetoxin-1-M2 (molecular weight = 884 Da). The increase in molecular weight combined with evidence from tandem mass spectrometry showing an increased tendency for loss of water molecules, along with considerations of established precedents for chemical transformations led to the conclusion that brevetoxin-1-M1 was formed by converting one double bond in the E or F ring of brevetoxin-1 into a diol. The second metabolite (brevetoxin-1-M2) is proposed to be a hydrolysis product of brevetoxin-1 involving opening of the lactone ring with the addition of a water molecule. The incubation study of the other starting compound, brevetoxin-2, found two metabolites in the LC-ES-MS selected ion chromatogram. Brevetoxin-2-M1 (molecular weight = 912 Da) gave a large [M−H]− peak at m/z 911, and its product ion mass spectrum allowed the deduction that this metabolite was the hydrolysis product of brevetoxin-2 involving conversion of the lactone to a carboxylic acid and an alcohol. The second metabolite (brevetoxin-2-M2, molecular weight = 896 Da) was deduced to have the same structure as that of brevetoxin-3 based on identical chromatographic retention times and similar mass spectra as those obtained for a brevetoxin-3 standard.
Keywords: Brevetoxin; Red tide; Metabolite; Electrospray; LC-MS; Mass spectrometry
Characterization of rat liver microsomal and hepatocytal metabolites of brevetoxins by liquid chromatography–electrospray tandem mass spectrometry by Weiqun Wang; Yousheng Hua; Guangdi Wang; Richard B. Cole (pp. 67-75).
Brevetoxins are natural neurotoxins that are produced by “red tide” algae. This class of compounds can cause neurotoxic shellfish poisoning and other health problems. Brevetoxin-2 is the most abundant among the nine brevetoxins that have been characterized, whereas brevetoxin-1 is the most toxic. In this study, brevetoxin-1 and brevetoxin-2 were incubated with rat liver hepatocytes and rat liver microsomes, respectively. After clean-up steps were taken to remove the proteins, samples were analyzed by liquid chromatography (LC) coupled with electrospray mass spectrometry (LC-MS). After incubation of brevetoxin-1, two metabolites were found: brevetoxin-1-M1 (molecular weight = 900 Da), and brevetoxin-1-M2 (molecular weight = 884 Da). The increase in molecular weight combined with evidence from tandem mass spectrometry showing an increased tendency for loss of water molecules, along with considerations of established precedents for chemical transformations led to the conclusion that brevetoxin-1-M1 was formed by converting one double bond in the E or F ring of brevetoxin-1 into a diol. The second metabolite (brevetoxin-1-M2) is proposed to be a hydrolysis product of brevetoxin-1 involving opening of the lactone ring with the addition of a water molecule. The incubation study of the other starting compound, brevetoxin-2, found two metabolites in the LC-ES-MS selected ion chromatogram. Brevetoxin-2-M1 (molecular weight = 912 Da) gave a large [M−H]− peak at m/z 911, and its product ion mass spectrum allowed the deduction that this metabolite was the hydrolysis product of brevetoxin-2 involving conversion of the lactone to a carboxylic acid and an alcohol. The second metabolite (brevetoxin-2-M2, molecular weight = 896 Da) was deduced to have the same structure as that of brevetoxin-3 based on identical chromatographic retention times and similar mass spectra as those obtained for a brevetoxin-3 standard.
Keywords: Brevetoxin; Red tide; Metabolite; Electrospray; LC-MS; Mass spectrometry
A new mass spectrometry electrospray tip obtained via precise mechanical micromachining by Yan-Zin Chang; Mu-Wen Yang; Gou-Jen Wang (pp. 76-82).
A new electrospray tip incorporating a beveled edge has been designed for use in plastic electrophoresis chip/electrospray mass spectrometry. Theoretical hydrodynamic analysis has been conducted to model the analytical sensitivity of the spray tip shape. A larger sample wall, that provides increased hydraulic pressure, is recommended in order to preserve the fluid stability at the tip outlet. A polymer with better hydrophobic characteristics than glass was used for the spray tip in order to restrict moisture accumulation at the spray tip outlet. Experimental results demonstrate that the analytical sensitivity of the proposed scheme is better than that obtained using the flat-head tip. Although a tapered capillary tip is commonly used in electrophoresis chip/electrospray mass spectrometry, the proposed tip offers a similar sensitivity while being more rugged and durable than the conventional tapered capillary tip. The cost of our design is also much lower than conventional spray tips.
Keywords: Mass spectrometry; Electrospray tip; Precise mechanical micromachining; Taylor cone
A new mass spectrometry electrospray tip obtained via precise mechanical micromachining by Yan-Zin Chang; Mu-Wen Yang; Gou-Jen Wang (pp. 76-82).
A new electrospray tip incorporating a beveled edge has been designed for use in plastic electrophoresis chip/electrospray mass spectrometry. Theoretical hydrodynamic analysis has been conducted to model the analytical sensitivity of the spray tip shape. A larger sample wall, that provides increased hydraulic pressure, is recommended in order to preserve the fluid stability at the tip outlet. A polymer with better hydrophobic characteristics than glass was used for the spray tip in order to restrict moisture accumulation at the spray tip outlet. Experimental results demonstrate that the analytical sensitivity of the proposed scheme is better than that obtained using the flat-head tip. Although a tapered capillary tip is commonly used in electrophoresis chip/electrospray mass spectrometry, the proposed tip offers a similar sensitivity while being more rugged and durable than the conventional tapered capillary tip. The cost of our design is also much lower than conventional spray tips.
Keywords: Mass spectrometry; Electrospray tip; Precise mechanical micromachining; Taylor cone
FluMag-SELEX as an advantageous method for DNA aptamer selection by R. Stoltenburg; C. Reinemann; B. Strehlitz (pp. 83-91).
Aptamers are ssDNA or RNA oligonucleotides with very high affinity for their target. They bind to the target with high selectivity and specificity because of their specific three-dimensional shape. They are developed by the so-called Systematic Evolution of Ligands by Exponential Enrichment (SELEX) process. We have modified this method in two steps—use of fluorescent labels for DNA quantification and use of magnetic beads for target immobilization. Thus, radioactive labelling is avoided. Immobilization on magnetic beads enables easy handling, use of very small amounts of target for the aptamer selection, rapid and efficient separation of bound and unbound molecules, and stringent washing steps. We have called this modified SELEX technology FluMag-SELEX. With FluMag-SELEX we have provided a methodological background for our objective of being able to select DNA aptamers for targets with very different properties and size. These aptamers will be applied as new biosensor receptors. In this work selection of streptavidin-specific aptamers by FluMag-SELEX is described. The streptavidin-specific aptamers will be used to check the surface occupancy of streptavidin-coated magnetic beads with biotinylated molecules after immobilization procedures.
Keywords: Aptamer; SELEX; Streptavidin; Magnetic beads
FluMag-SELEX as an advantageous method for DNA aptamer selection by R. Stoltenburg; C. Reinemann; B. Strehlitz (pp. 83-91).
Aptamers are ssDNA or RNA oligonucleotides with very high affinity for their target. They bind to the target with high selectivity and specificity because of their specific three-dimensional shape. They are developed by the so-called Systematic Evolution of Ligands by Exponential Enrichment (SELEX) process. We have modified this method in two steps—use of fluorescent labels for DNA quantification and use of magnetic beads for target immobilization. Thus, radioactive labelling is avoided. Immobilization on magnetic beads enables easy handling, use of very small amounts of target for the aptamer selection, rapid and efficient separation of bound and unbound molecules, and stringent washing steps. We have called this modified SELEX technology FluMag-SELEX. With FluMag-SELEX we have provided a methodological background for our objective of being able to select DNA aptamers for targets with very different properties and size. These aptamers will be applied as new biosensor receptors. In this work selection of streptavidin-specific aptamers by FluMag-SELEX is described. The streptavidin-specific aptamers will be used to check the surface occupancy of streptavidin-coated magnetic beads with biotinylated molecules after immobilization procedures.
Keywords: Aptamer; SELEX; Streptavidin; Magnetic beads
In vitro monitoring of GTPase activity and enzyme kinetics studies using capillary electrophoresis by Sandro Hillebrand; Wanius Garcia; Marcelo Delmar Cantú; Ana Paula Ulian de Araújo; Manami Tanaka; Tomoo Tanaka; Richard Charles Garratt; Emanuel Carrilho (pp. 92-97).
A capillary electrophoresis (CE)-based method for the in vitro detection and monitoring of nucleotide-triphosphatase activity is described. This robust and reproducible method was used to investigate GTPase activity of a recombinant protein construct containing the catalytic domain of Human SEPT4/Bradeion β (GST-rDGTPase). This example application demonstrates that the CE technique can replace classical radioactive methods for GTPase activity assays and may be used as a routine analytical tool. Enzyme kinetics of GST-rDGTPase was studied and yielded the following kinetic parameters: v max = 1.7 μM min−1 ± 0.1, Km = 1.0 mM ± 0.3, and apKcat = 9 × 10−3 s−1. In addition the effect of co-factors such as Mg2+ and Mn2+ on the catalytic activity was investigated. The described analytical method was also shown to be useful to analyze diphosphated and triphosphated forms of other nucleotides.
Keywords: SEPT4 Bradeion β; Septins; Guanosine triphosphate; Influence of magnesium (Mg2+); Recombinant GTPase
In vitro monitoring of GTPase activity and enzyme kinetics studies using capillary electrophoresis by Sandro Hillebrand; Wanius Garcia; Marcelo Delmar Cantú; Ana Paula Ulian de Araújo; Manami Tanaka; Tomoo Tanaka; Richard Charles Garratt; Emanuel Carrilho (pp. 92-97).
A capillary electrophoresis (CE)-based method for the in vitro detection and monitoring of nucleotide-triphosphatase activity is described. This robust and reproducible method was used to investigate GTPase activity of a recombinant protein construct containing the catalytic domain of Human SEPT4/Bradeion β (GST-rDGTPase). This example application demonstrates that the CE technique can replace classical radioactive methods for GTPase activity assays and may be used as a routine analytical tool. Enzyme kinetics of GST-rDGTPase was studied and yielded the following kinetic parameters: v max = 1.7 μM min−1 ± 0.1, Km = 1.0 mM ± 0.3, and apKcat = 9 × 10−3 s−1. In addition the effect of co-factors such as Mg2+ and Mn2+ on the catalytic activity was investigated. The described analytical method was also shown to be useful to analyze diphosphated and triphosphated forms of other nucleotides.
Keywords: SEPT4 Bradeion β; Septins; Guanosine triphosphate; Influence of magnesium (Mg2+); Recombinant GTPase
Long-term stability of hydrocarbons in NIST gas standard reference material (SRM) 1800 by George C. Rhoderick (pp. 98-106).
A gas standard reference material (SRM) containing fifteen hydrocarbons in nitrogen at a nominal 5 nmol mol−1 was issued in 1993. The certification period for SRM 1800 was assigned as 2 years, because of limited stability data. Over a period of 10 years reanalysis of the lot standard (a sample chosen from the SRM lot to which all other lot samples are compared), SRM samples remaining in stock for sale, and SRMs returned to the National Institute of Standards and Technology (NIST) for recertification, were compared with primary standards to assess the stability of the hydrocarbons. New primary standards were periodically introduced into the original primary standard suite to assess the stability and consistency of the primary standards. Over this ten-year period 11 SRM 1800 samples were reanalyzed, resulting in 210 amount-of-substance fraction (concentration) determinations performed for quality-assurance purposes. Of these measurements 209 (99.5%) agreed within the original 95% confidence interval of the ±4% expanded uncertainty, demonstrating the stability of the standards. There was also agreement to within ±2% of the original concentration for 204 (97%) of the measurements. This is well within the original ±4% expanded uncertainty assigned to the hydrocarbon concentrations at the approximate 95% confidence interval demonstrating stability. These results will enable the expiry date to be increased for future restock issues of SRM 1800.
Keywords: Hydrocarbons; Long-term stability; nmol mol−1 ; SRM 1800; Gas standards; Stability
Long-term stability of hydrocarbons in NIST gas standard reference material (SRM) 1800 by George C. Rhoderick (pp. 98-106).
A gas standard reference material (SRM) containing fifteen hydrocarbons in nitrogen at a nominal 5 nmol mol−1 was issued in 1993. The certification period for SRM 1800 was assigned as 2 years, because of limited stability data. Over a period of 10 years reanalysis of the lot standard (a sample chosen from the SRM lot to which all other lot samples are compared), SRM samples remaining in stock for sale, and SRMs returned to the National Institute of Standards and Technology (NIST) for recertification, were compared with primary standards to assess the stability of the hydrocarbons. New primary standards were periodically introduced into the original primary standard suite to assess the stability and consistency of the primary standards. Over this ten-year period 11 SRM 1800 samples were reanalyzed, resulting in 210 amount-of-substance fraction (concentration) determinations performed for quality-assurance purposes. Of these measurements 209 (99.5%) agreed within the original 95% confidence interval of the ±4% expanded uncertainty, demonstrating the stability of the standards. There was also agreement to within ±2% of the original concentration for 204 (97%) of the measurements. This is well within the original ±4% expanded uncertainty assigned to the hydrocarbon concentrations at the approximate 95% confidence interval demonstrating stability. These results will enable the expiry date to be increased for future restock issues of SRM 1800.
Keywords: Hydrocarbons; Long-term stability; nmol mol−1 ; SRM 1800; Gas standards; Stability
Trace analysis of polycyclic aromatic hydrocarbons in suspended particulate matter by accelerated solvent extraction followed by gas chromatography–mass spectrometry by M. A. Olivella (pp. 107-114).
An analytical procedure based on extraction by accelerated solvent extraction (ASE) followed by gas chromatography–mass spectrometry (GC/MS) analysis has been developed for the determination of particulate polycyclic aromatic hydrocarbons (PAHs) from large-volume water samples (20 L). The effect of temperature and number of cycles on the efficiency of ASE was investigated: the best results were obtained by using a temperature of 100°C and one static cycle. A mixture of hexane/acetone 1:1 (v/v) was used as extraction solvent. Mean total method recovery under optimized conditions was 85%. The developed methodology was applied to the analysis of suspended particulate matter from Lake Maggiore waters (north of Italy). Mean PAH concentrations in suspended particulate matter from Lake Maggiore ranged from 0.2 ng L−1 for anthracene to 18.7 ng L−1 for naphthalene.
Keywords: Polycyclic aromatic hydrocarbons (PAHs); Accelerated solvent extraction (ASE); Suspended particulate matter (SPM); Lake Maggiore
Trace analysis of polycyclic aromatic hydrocarbons in suspended particulate matter by accelerated solvent extraction followed by gas chromatography–mass spectrometry by M. A. Olivella (pp. 107-114).
An analytical procedure based on extraction by accelerated solvent extraction (ASE) followed by gas chromatography–mass spectrometry (GC/MS) analysis has been developed for the determination of particulate polycyclic aromatic hydrocarbons (PAHs) from large-volume water samples (20 L). The effect of temperature and number of cycles on the efficiency of ASE was investigated: the best results were obtained by using a temperature of 100°C and one static cycle. A mixture of hexane/acetone 1:1 (v/v) was used as extraction solvent. Mean total method recovery under optimized conditions was 85%. The developed methodology was applied to the analysis of suspended particulate matter from Lake Maggiore waters (north of Italy). Mean PAH concentrations in suspended particulate matter from Lake Maggiore ranged from 0.2 ng L−1 for anthracene to 18.7 ng L−1 for naphthalene.
Keywords: Polycyclic aromatic hydrocarbons (PAHs); Accelerated solvent extraction (ASE); Suspended particulate matter (SPM); Lake Maggiore
Differential pulse voltammetric determination of tin in the presence of noble metals by Damiano Monticelli; Rinaldo Psaro; Andrea Pozzi; Carlo Dossi; Sandro Recchia (pp. 115-121).
A voltammetric method for the determination of tin is proposed to minimise interferences from noble metals that are commonly encountered with other analytical techniques. Strong distortions of voltammetric peaks are observed in the presence of platinum. On the basis of a full investigation, the formation of an intermediate Sn(II)–Pt mixed chloro-complex at the electrode surface is identified as being responsible for the platinum interference, as it competes with the normal Sn(IV)→Sn(0)Hg reduction. The use of a higher scan rate prevents the relatively low reaction kinetics and thus gets rid of this interference. No problems are encountered with other noble metals such as Pd, Ir, Re, Rh and Ru when using the modified method, although a baseline subtraction is necessary for the latter one. The proposed method is validated with real Pt–Sn catalysts.
Keywords: Catalyst; Tin; Noble metals; Differential pulse voltammetry; Chemical coupled reaction
Differential pulse voltammetric determination of tin in the presence of noble metals by Damiano Monticelli; Rinaldo Psaro; Andrea Pozzi; Carlo Dossi; Sandro Recchia (pp. 115-121).
A voltammetric method for the determination of tin is proposed to minimise interferences from noble metals that are commonly encountered with other analytical techniques. Strong distortions of voltammetric peaks are observed in the presence of platinum. On the basis of a full investigation, the formation of an intermediate Sn(II)–Pt mixed chloro-complex at the electrode surface is identified as being responsible for the platinum interference, as it competes with the normal Sn(IV)→Sn(0)Hg reduction. The use of a higher scan rate prevents the relatively low reaction kinetics and thus gets rid of this interference. No problems are encountered with other noble metals such as Pd, Ir, Re, Rh and Ru when using the modified method, although a baseline subtraction is necessary for the latter one. The proposed method is validated with real Pt–Sn catalysts.
Keywords: Catalyst; Tin; Noble metals; Differential pulse voltammetry; Chemical coupled reaction
Analysis of polycyclic aromatic hydrocarbons adsorbed on particles of atmospheric interest using pressurised fluid extraction by Emilie Perraudin; Hélène Budzinski; Eric Villenave (pp. 122-131).
Pressurised fluid extraction (PFE) was used for the measurement of 13 polycyclic aromatic hydrocarbons (PAHs) adsorbed on different types of particles: two model particles (PAH-coated silica, PAH-coated graphite) and two natural atmospheric particles (urban dust and diesel exhaust, from NIST reference materials). Samples were analysed by gas chromatography coupled to mass spectrometry. Extraction efficiency was evaluated with internal standard recovery yields and was shown to depend on the nature of the particle, on the structure of the analytes and on the PAH concentration. Extraction conditions (toluene, 130°C, 130 bar, 2 × 8-min static cycles) were optimised to extract PAHs when strongly interacting with solid matrices and were validated by the analysis of two PAH-certified materials.
Keywords: Pressurised fluid extraction; Polycyclic aromatic hydrocarbons; Atmospheric particles; Internal standard recovery yield
Analysis of polycyclic aromatic hydrocarbons adsorbed on particles of atmospheric interest using pressurised fluid extraction by Emilie Perraudin; Hélène Budzinski; Eric Villenave (pp. 122-131).
Pressurised fluid extraction (PFE) was used for the measurement of 13 polycyclic aromatic hydrocarbons (PAHs) adsorbed on different types of particles: two model particles (PAH-coated silica, PAH-coated graphite) and two natural atmospheric particles (urban dust and diesel exhaust, from NIST reference materials). Samples were analysed by gas chromatography coupled to mass spectrometry. Extraction efficiency was evaluated with internal standard recovery yields and was shown to depend on the nature of the particle, on the structure of the analytes and on the PAH concentration. Extraction conditions (toluene, 130°C, 130 bar, 2 × 8-min static cycles) were optimised to extract PAHs when strongly interacting with solid matrices and were validated by the analysis of two PAH-certified materials.
Keywords: Pressurised fluid extraction; Polycyclic aromatic hydrocarbons; Atmospheric particles; Internal standard recovery yield
Use of solid-phase extraction to enable enhanced detection of acyl homoserine lactones (AHLs) in environmental samples by Peter J. Schupp; Timothy S. Charlton; Michael W. Taylor; Staffan Kjelleberg; Peter D. Steinberg (pp. 132-137).
A challenge for understanding the role of bacterial cell–cell signalling in the environment is the detection of those signals, which are often present in low (nmol L−1) concentrations. We describe here a simple purification method, solid-phase extraction (SPE), for increasing the sensitivity of detection for one such group of signals, acyl homoserine lactones (AHLs), in environmental samples. Spiking of dried marine sponge tissue (Stylinos sp.) with AHLs resulted in detection down to 0.01 ppm for 3-oxo-hexanoyl homoserine lactone (3-oxo C6-HSL) and 1 ppm for hexanoyl homoserine lactone (C6-HSL). Compared with liquid extraction methods use of SPE resulted in twofold and tenfold improvements in sensitivity, respectively.
Keywords: Acyl homoserine lactone (AHL); Marine sponges; Solid-phase extraction; SPE
Use of solid-phase extraction to enable enhanced detection of acyl homoserine lactones (AHLs) in environmental samples by Peter J. Schupp; Timothy S. Charlton; Michael W. Taylor; Staffan Kjelleberg; Peter D. Steinberg (pp. 132-137).
A challenge for understanding the role of bacterial cell–cell signalling in the environment is the detection of those signals, which are often present in low (nmol L−1) concentrations. We describe here a simple purification method, solid-phase extraction (SPE), for increasing the sensitivity of detection for one such group of signals, acyl homoserine lactones (AHLs), in environmental samples. Spiking of dried marine sponge tissue (Stylinos sp.) with AHLs resulted in detection down to 0.01 ppm for 3-oxo-hexanoyl homoserine lactone (3-oxo C6-HSL) and 1 ppm for hexanoyl homoserine lactone (C6-HSL). Compared with liquid extraction methods use of SPE resulted in twofold and tenfold improvements in sensitivity, respectively.
Keywords: Acyl homoserine lactone (AHL); Marine sponges; Solid-phase extraction; SPE
Determination of phenols in waters using micro-pumped multicommutation and spectrophotometric detection: an automated alternative to the standard procedure by Eva Ródenas-Torralba; Ángel Morales-Rubio; Miguel de la Guardia (pp. 138-144).
An automated and greener spectrophotometric procedure has been developed for the determination of phenol in water at 700 nm. The method uses the reaction between phenol, sodium nitroprusside, and hydroxylamine hydrochloride in a buffered medium at pH 12.3. The flow manifold comprises four solenoid micro-pumps employed for sample and reagent introduction into the reaction coil and to transport the colored product formed to the detector. The linear dynamic range was 50–3,500 ng mL−1 (R = 0.99997; n = 6) and the method provided a limit of detection (3σ) of 13 ng mL−1. The sampling throughput was estimated to be 65 measurements per hour and the coefficient of variation was 0.5% (n = 10) for a 1.0 μg mL−1 phenol concentration. Recoveries of 92–105% were obtained for phenol determination in spiked water samples at concentration levels from 50 to 5,000 ng mL−1. The use of multicommutation reduced the reagent consumption 25-fold, the sample consumption 225-fold, and the waste generation 30-fold compared with the batch procedure. The proposed method is an environmentally friendly alternative to the official 4-aminoantipyrine method since it avoids the use of chloroform.
Keywords: Phenol; Solenoid micro-pumps; Multicommutation; Spectrophotometry; Waters
Determination of phenols in waters using micro-pumped multicommutation and spectrophotometric detection: an automated alternative to the standard procedure by Eva Ródenas-Torralba; Ángel Morales-Rubio; Miguel de la Guardia (pp. 138-144).
An automated and greener spectrophotometric procedure has been developed for the determination of phenol in water at 700 nm. The method uses the reaction between phenol, sodium nitroprusside, and hydroxylamine hydrochloride in a buffered medium at pH 12.3. The flow manifold comprises four solenoid micro-pumps employed for sample and reagent introduction into the reaction coil and to transport the colored product formed to the detector. The linear dynamic range was 50–3,500 ng mL−1 (R = 0.99997; n = 6) and the method provided a limit of detection (3σ) of 13 ng mL−1. The sampling throughput was estimated to be 65 measurements per hour and the coefficient of variation was 0.5% (n = 10) for a 1.0 μg mL−1 phenol concentration. Recoveries of 92–105% were obtained for phenol determination in spiked water samples at concentration levels from 50 to 5,000 ng mL−1. The use of multicommutation reduced the reagent consumption 25-fold, the sample consumption 225-fold, and the waste generation 30-fold compared with the batch procedure. The proposed method is an environmentally friendly alternative to the official 4-aminoantipyrine method since it avoids the use of chloroform.
Keywords: Phenol; Solenoid micro-pumps; Multicommutation; Spectrophotometry; Waters
Matrix effects during phosphorus determination with quadrupole inductively coupled plasma mass spectrometry by Miroslav Kovačevič; Walter Goessler; Nevenka Mikac; Marjan Veber (pp. 145-151).
A quadrupole inductively coupled plasma mass spectrometer was evaluated for use in the detection of phosphorus. The influences of nitric acid and methanol (simulating the composition of a sample solution after nitric acid digestion) on phosphorus determination were studied using two different measuring methods at different plasma conditions: detection of phosphorus ions at m/z 31 and detection of phosphorus oxide ions at m/z 47. The existence of polyatomic interferences at m/z 31 and 47 was explored. Nitric acid and methanol are shown to be the sources of polyatomic ions and therefore cause poorer detection limits. Better detection limits were achieved in such matrices when phosphorus was detected as 31P+. The presence of methanol improves the system sensitivity towards phosphorus sevenfold; however, this positive effect is hindered by the high background signal due to carbon-based polyatomic ions. For samples with an organic matrix an appropriate mineralization procedure should be applied (high excess of nitric acid and high temperature) to quantitatively oxidize organic compounds to carbon dioxide, which is easily removed from the sample, in order to achieve correct results.
Keywords: Quadrupole inductively coupled plasma mass spectrometry; Phosphorus; Matrix effects; Polyatomic interferences; Signal enhancement
Matrix effects during phosphorus determination with quadrupole inductively coupled plasma mass spectrometry by Miroslav Kovačevič; Walter Goessler; Nevenka Mikac; Marjan Veber (pp. 145-151).
A quadrupole inductively coupled plasma mass spectrometer was evaluated for use in the detection of phosphorus. The influences of nitric acid and methanol (simulating the composition of a sample solution after nitric acid digestion) on phosphorus determination were studied using two different measuring methods at different plasma conditions: detection of phosphorus ions at m/z 31 and detection of phosphorus oxide ions at m/z 47. The existence of polyatomic interferences at m/z 31 and 47 was explored. Nitric acid and methanol are shown to be the sources of polyatomic ions and therefore cause poorer detection limits. Better detection limits were achieved in such matrices when phosphorus was detected as 31P+. The presence of methanol improves the system sensitivity towards phosphorus sevenfold; however, this positive effect is hindered by the high background signal due to carbon-based polyatomic ions. For samples with an organic matrix an appropriate mineralization procedure should be applied (high excess of nitric acid and high temperature) to quantitatively oxidize organic compounds to carbon dioxide, which is easily removed from the sample, in order to achieve correct results.
Keywords: Quadrupole inductively coupled plasma mass spectrometry; Phosphorus; Matrix effects; Polyatomic interferences; Signal enhancement