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Analytical and Bioanalytical Chemistry (v.383, #6)
Interfacing liquid chromatography with atmospheric pressure MALDI-MS by Jürg M. Daniel; Victor V. Laiko; Vladimir M. Doroshenko; Renato Zenobi (pp. 895-902).
Two different strategies for coupling liquid chromatography with atmospheric pressure matrix assisted laser desorption/ionization (AP MALDI) are presented. The first method is flow-injection liquid AP UV-MALDI. Compared with previous similar research, the detection limit was improved 10 times to 8.3 fmol using a solution of 50 nM peptide with 25 mM α-cyano-4-hydroxycinnamic acid. The applicability of this method to measure oligosaccharides, actinomycin antibiotics, antibiotics, phosphopeptides, and proteins is demonstrated. The upper mass limit achieved with the current instrumentation is 6,500 Da (doubly charged cytochrome c). The feasibility of a second strategy based on single-droplet IR AP MALDI is demonstrated here. Aqueous peptide solutions were successfully measured by this method.
Keywords: Matrix assisted laser desorption/ionization; IR ionization from solution; IR matrix assisted laser desorption/ionization; Liquid matrix; Atmospheric pressure matrix assisted laser desorption/ionization
Interfacing liquid chromatography with atmospheric pressure MALDI-MS by Jürg M. Daniel; Victor V. Laiko; Vladimir M. Doroshenko; Renato Zenobi (pp. 895-902).
Two different strategies for coupling liquid chromatography with atmospheric pressure matrix assisted laser desorption/ionization (AP MALDI) are presented. The first method is flow-injection liquid AP UV-MALDI. Compared with previous similar research, the detection limit was improved 10 times to 8.3 fmol using a solution of 50 nM peptide with 25 mM α-cyano-4-hydroxycinnamic acid. The applicability of this method to measure oligosaccharides, actinomycin antibiotics, antibiotics, phosphopeptides, and proteins is demonstrated. The upper mass limit achieved with the current instrumentation is 6,500 Da (doubly charged cytochrome c). The feasibility of a second strategy based on single-droplet IR AP MALDI is demonstrated here. Aqueous peptide solutions were successfully measured by this method.
Keywords: Matrix assisted laser desorption/ionization; IR ionization from solution; IR matrix assisted laser desorption/ionization; Liquid matrix; Atmospheric pressure matrix assisted laser desorption/ionization
Detection of testosterone, nandrolone and precursors in horse hair by P. Anielski; D. Thieme; A. Schlupp; J. Grosse; F. Ellendorff; R. K. Mueller (pp. 903-908).
Growing interest among several horse-breeder associations has initiated the development of a screening procedure to test for anabolic agents in hair, which has the advantage over blood and urine specimens of allowing long-term detection. An analytical method was established to monitor in tails or manes several anabolic substances available as veterinary medicines or as so-called nutritional supplements (clenbuterol, different esters or prohormones of nandrolone and testosterone). The analytical procedure to detect steroids in hair samples consists of the following steps: decontamination of the hair strand or segment with methanol/water (1:1), milling, extraction of the hair material in an ultrasonic bath using methanol, purification by liquid–liquid extraction (n-pentane/methanol, 25:1) and HPLC cleanup, derivatisation of the relevant LC fractions with MSTFA, and measurement using GC-MS/MS technique. The first objective of our study was the detection of exogenous nandrolone (nortestosterone, NT) in the horse hair; therefore nandrolone-associated compounds [nandrolone dodecanoate administered intramuscularly (i.m.) and a mixture of 4-estrenediol and 4-estrenedione, transdermal] were administered to four geldings. The highest concentrations of NT following i.m. treatment were measured after 10 days in a 2-cm hair segment (up to 18 pg/mg); NT was detectable for up to 120 days and in some cases up to 330 days in tail hair (limit of detection 0.3 pg/mg). Following transdermal application, nandrolone as well as the administered prohormones were identified in tail and mane until the latest sampling at 3 months. Furthermore, untreated stallions (128) were investigated to estimate the range of endogenous levels of NT and testosterone (T) in hair. Maximum values of 3 pg/mg (NT) and 1 pg/mg (T) were quantified originating from endogenous formation in the male horse. Additionally, a possible relationship between steroid concentrations in hair specimens and the age of stallions was appraised. NT and T were not detected in hair samples of control geldings. Following nandrolone treatment of geldings, highest values in hair exceeded the endogenous amount detected in untreated stallions. Therefore comparison of concentrations measured in control samples with the estimated endogenous levels could give a clue to exogenous application in cases of abnormally high amounts of NT or T. The possibility of the evaluation of threshold values is discussed as a means to verify an exogenous administration of NT and T in hair samples. Furthermore, the detection of a synthetic substance in hair, e. g. the parent steroid ester by itself, would be unequivocal proof of an exogenous origin of NT or T and the previous medication of the stallion.
Keywords: Hair analysis; Nandrolone; Testosterone; Horse; Anabolic steroids
Detection of testosterone, nandrolone and precursors in horse hair by P. Anielski; D. Thieme; A. Schlupp; J. Grosse; F. Ellendorff; R. K. Mueller (pp. 903-908).
Growing interest among several horse-breeder associations has initiated the development of a screening procedure to test for anabolic agents in hair, which has the advantage over blood and urine specimens of allowing long-term detection. An analytical method was established to monitor in tails or manes several anabolic substances available as veterinary medicines or as so-called nutritional supplements (clenbuterol, different esters or prohormones of nandrolone and testosterone). The analytical procedure to detect steroids in hair samples consists of the following steps: decontamination of the hair strand or segment with methanol/water (1:1), milling, extraction of the hair material in an ultrasonic bath using methanol, purification by liquid–liquid extraction (n-pentane/methanol, 25:1) and HPLC cleanup, derivatisation of the relevant LC fractions with MSTFA, and measurement using GC-MS/MS technique. The first objective of our study was the detection of exogenous nandrolone (nortestosterone, NT) in the horse hair; therefore nandrolone-associated compounds [nandrolone dodecanoate administered intramuscularly (i.m.) and a mixture of 4-estrenediol and 4-estrenedione, transdermal] were administered to four geldings. The highest concentrations of NT following i.m. treatment were measured after 10 days in a 2-cm hair segment (up to 18 pg/mg); NT was detectable for up to 120 days and in some cases up to 330 days in tail hair (limit of detection 0.3 pg/mg). Following transdermal application, nandrolone as well as the administered prohormones were identified in tail and mane until the latest sampling at 3 months. Furthermore, untreated stallions (128) were investigated to estimate the range of endogenous levels of NT and testosterone (T) in hair. Maximum values of 3 pg/mg (NT) and 1 pg/mg (T) were quantified originating from endogenous formation in the male horse. Additionally, a possible relationship between steroid concentrations in hair specimens and the age of stallions was appraised. NT and T were not detected in hair samples of control geldings. Following nandrolone treatment of geldings, highest values in hair exceeded the endogenous amount detected in untreated stallions. Therefore comparison of concentrations measured in control samples with the estimated endogenous levels could give a clue to exogenous application in cases of abnormally high amounts of NT or T. The possibility of the evaluation of threshold values is discussed as a means to verify an exogenous administration of NT and T in hair samples. Furthermore, the detection of a synthetic substance in hair, e. g. the parent steroid ester by itself, would be unequivocal proof of an exogenous origin of NT or T and the previous medication of the stallion.
Keywords: Hair analysis; Nandrolone; Testosterone; Horse; Anabolic steroids
Stereospecific pharmacokinetic characterisation of phenprocoumon metabolites, and mass-spectrometric identification of two novel metabolites in human plasma and liver microsomes by Bernd Kammerer; Rainer Kahlich; Mike Ufer; Andreas Schenkel; Stefan Laufer; Christoph H. Gleiter (pp. 909-917).
Phenprocoumon belongs to the group of vitamin K antagonists (VKAs), for example warfarin and acenocoumarol. It is widely used for therapeutic anticoagulation and clinically administered as a racemate. Both enantiomers are partially metabolized by the polymorphic CYP2C9 enzyme. The pharmacokinetics are, however, substantially less dependent on CYP2C9 activity or genotype than for other CYP2C9-metabolised VKAs, and pharmacokinetic differences for the enantiomers are only minor. We have investigated the stereospecific pharmacokinetics of the monohydroxylated phenprocoumon metabolites in human plasma by achiral–chiral LC–LC–MS–MS coupling. In addition to the known metabolites, 4′-, 6-, and 7-hydroxyphenprocoumon, two other monohydroxylated metabolites (M1 and M2) were detected in plasma and human liver microsomal incubations. One of these was identified as 2′-hydroxyphenprocoumon by comparison with synthetic standards; the other seemed to be a side-chain-hydroxylated derivative. Analysis of enantiomeric metabolite ratios after a single oral dose of phenprocoumon revealed changes over time with an overall preponderance of the respective (R) enantiomers. The minor role of CYP2C9 in 4′-hydroxy-PPC formation and the effect of CYP2C9 genotype for (S)-6- and (S)-7-hydroxy-PPC were confirmed. M1 and M2 are formed highly stereoselectively, without dependence on CYP2C9 genotype. These may be interpreted as alternative metabolic pathways that render phenprocoumon less dependent on CYP2C9 activity or genotype.
Keywords: Phenprocoumon metabolism; Chiral discrimination effects; LC–MS coupling
Stereospecific pharmacokinetic characterisation of phenprocoumon metabolites, and mass-spectrometric identification of two novel metabolites in human plasma and liver microsomes by Bernd Kammerer; Rainer Kahlich; Mike Ufer; Andreas Schenkel; Stefan Laufer; Christoph H. Gleiter (pp. 909-917).
Phenprocoumon belongs to the group of vitamin K antagonists (VKAs), for example warfarin and acenocoumarol. It is widely used for therapeutic anticoagulation and clinically administered as a racemate. Both enantiomers are partially metabolized by the polymorphic CYP2C9 enzyme. The pharmacokinetics are, however, substantially less dependent on CYP2C9 activity or genotype than for other CYP2C9-metabolised VKAs, and pharmacokinetic differences for the enantiomers are only minor. We have investigated the stereospecific pharmacokinetics of the monohydroxylated phenprocoumon metabolites in human plasma by achiral–chiral LC–LC–MS–MS coupling. In addition to the known metabolites, 4′-, 6-, and 7-hydroxyphenprocoumon, two other monohydroxylated metabolites (M1 and M2) were detected in plasma and human liver microsomal incubations. One of these was identified as 2′-hydroxyphenprocoumon by comparison with synthetic standards; the other seemed to be a side-chain-hydroxylated derivative. Analysis of enantiomeric metabolite ratios after a single oral dose of phenprocoumon revealed changes over time with an overall preponderance of the respective (R) enantiomers. The minor role of CYP2C9 in 4′-hydroxy-PPC formation and the effect of CYP2C9 genotype for (S)-6- and (S)-7-hydroxy-PPC were confirmed. M1 and M2 are formed highly stereoselectively, without dependence on CYP2C9 genotype. These may be interpreted as alternative metabolic pathways that render phenprocoumon less dependent on CYP2C9 activity or genotype.
Keywords: Phenprocoumon metabolism; Chiral discrimination effects; LC–MS coupling
Biocomposites of covalently linked glucose oxidase on carbon nanotubes for glucose biosensor by Jian Li; Yue-Bo Wang; Jian-Ding Qiu; Da-cheng Sun; Xing-Hua Xia (pp. 918-922).
The formation of covalently linked composites of multi–walled carbon nanotubes (MWCNT) and glucose oxidase (GOD) with high-function density for use as a biosensing interface is described. The reaction intermediates and the final product were characterized by using FT–IR spectroscopy, and the MWCNT-coated GOD nanocomposites were examined by atomic force microscopy (AFM) and transmission electron microscopy (TEM). Interestingly, it was found that the GOD–MWCNT composites are highly water soluble. Electrochemical characterization of the GOD–MWCNT composites that were modified on a glassy carbon electrode shows that the covalently linked GOD retains its bioactivity and can specifically catalyze the oxidation of glucose. The oxidation current shows a linear dependence on the glucose concentration in the solution in the range of 0.5–40 mM with a detection limit of 30 μM and a detection sensitivity of 11.3 μA/mMcm2. The present method may provide a way to synthesize MWCNT related composites with other biomolecules and for the construction of enzymatic reaction-based biofuel cells and biosensors.
Keywords: Carbon nanotubes (CNT); Enzyme; Biomaterials; Glucose; Biosensors
Biocomposites of covalently linked glucose oxidase on carbon nanotubes for glucose biosensor by Jian Li; Yue-Bo Wang; Jian-Ding Qiu; Da-cheng Sun; Xing-Hua Xia (pp. 918-922).
The formation of covalently linked composites of multi–walled carbon nanotubes (MWCNT) and glucose oxidase (GOD) with high-function density for use as a biosensing interface is described. The reaction intermediates and the final product were characterized by using FT–IR spectroscopy, and the MWCNT-coated GOD nanocomposites were examined by atomic force microscopy (AFM) and transmission electron microscopy (TEM). Interestingly, it was found that the GOD–MWCNT composites are highly water soluble. Electrochemical characterization of the GOD–MWCNT composites that were modified on a glassy carbon electrode shows that the covalently linked GOD retains its bioactivity and can specifically catalyze the oxidation of glucose. The oxidation current shows a linear dependence on the glucose concentration in the solution in the range of 0.5–40 mM with a detection limit of 30 μM and a detection sensitivity of 11.3 μA/mMcm2. The present method may provide a way to synthesize MWCNT related composites with other biomolecules and for the construction of enzymatic reaction-based biofuel cells and biosensors.
Keywords: Carbon nanotubes (CNT); Enzyme; Biomaterials; Glucose; Biosensors
Fluorometric determination of thiol redox state by Nikolaos Patsoukis; Christos D. Georgiou (pp. 923-929).
This work presents a fluorometric thiol redox state (TRS) method based on the thiol-specific fluorescent probe monobromobimane. It determines the concentrations of non-protein (NP) thiols glutathione, cysteine and N-acetylcysteine, the protein (P) thiols, as well as the contribution of these components to symmetric and mixed disulfides (NPSSR, NPSSC, NPSSCAc, PSSR, PSSC, PSSCAc, PSSP). The method is very sensitive since it measures as low as 30 pmol -SH groups in samples with a minimum of 1–5 mg total protein, making possible the measurement of oxidative stress–related TRS components even in biological fluids such as cerebrospinal.
Keywords: Monobromobimane; Thiols; Disulfides; Oxidative stress
Fluorometric determination of thiol redox state by Nikolaos Patsoukis; Christos D. Georgiou (pp. 923-929).
This work presents a fluorometric thiol redox state (TRS) method based on the thiol-specific fluorescent probe monobromobimane. It determines the concentrations of non-protein (NP) thiols glutathione, cysteine and N-acetylcysteine, the protein (P) thiols, as well as the contribution of these components to symmetric and mixed disulfides (NPSSR, NPSSC, NPSSCAc, PSSR, PSSC, PSSCAc, PSSP). The method is very sensitive since it measures as low as 30 pmol -SH groups in samples with a minimum of 1–5 mg total protein, making possible the measurement of oxidative stress–related TRS components even in biological fluids such as cerebrospinal.
Keywords: Monobromobimane; Thiols; Disulfides; Oxidative stress
A new affinity-HPLC packing for protein separation: Cibacron blue attached uniform porous poly(HEMA-co-EDM) beads by Ender Unsal; Aysun Durdu; Begum Elmas; Murvet Tuncel; Ali Tuncel (pp. 930-937).
In this study, a new affinity high-performance liquid chromatography (HPLC) stationary phase suitable for protein separation was synthesized. In the first stage of the synthesis, uniform porous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate), poly(HEMA-co-EDM), beads 6.2 μm in size were obtained. Homogeneous distribution of hydroxyl groups in the bead interior was confirmed by confocal laser scanning microscopy. The plain poly(HEMA-co-EDM) particles gave very low non-specific protein adsorption with albumin. The selected dye ligand Cibacron blue F3G-A (CB F3G-A) was covalently linked onto the beads via hydroxyl groups. In the batch experiments, albumin adsorption up to 60 mg BSA/g particles was obtained with the CB F3G-A carrying poly(HEMA-co-EDM) beads. The affinity-HPLC of selected proteins (albumin and lysozyme) was investigated in a 25 mm×4.0-mm inner diameter column packed with CB F3G-A carrying beads and both proteins were successfully resolved. By a single injection, 200 μg of protein was loaded and quantitatively eluted from the column. The protein recovery increased with increasing flow rate and salt concentration of the elution buffer and decreased with the increasing protein feed concentration. During the albumin elution, theoretical plate numbers up to 30,000 plates/m were achieved by increasing the salt concentration.
Keywords: Hydroxyethyl methacrylate; Affinity chromatography; High-performance liquid chromatography; Cibacron blue; Albumin; Lysozyme
A new affinity-HPLC packing for protein separation: Cibacron blue attached uniform porous poly(HEMA-co-EDM) beads by Ender Unsal; Aysun Durdu; Begum Elmas; Murvet Tuncel; Ali Tuncel (pp. 930-937).
In this study, a new affinity high-performance liquid chromatography (HPLC) stationary phase suitable for protein separation was synthesized. In the first stage of the synthesis, uniform porous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate), poly(HEMA-co-EDM), beads 6.2 μm in size were obtained. Homogeneous distribution of hydroxyl groups in the bead interior was confirmed by confocal laser scanning microscopy. The plain poly(HEMA-co-EDM) particles gave very low non-specific protein adsorption with albumin. The selected dye ligand Cibacron blue F3G-A (CB F3G-A) was covalently linked onto the beads via hydroxyl groups. In the batch experiments, albumin adsorption up to 60 mg BSA/g particles was obtained with the CB F3G-A carrying poly(HEMA-co-EDM) beads. The affinity-HPLC of selected proteins (albumin and lysozyme) was investigated in a 25 mm×4.0-mm inner diameter column packed with CB F3G-A carrying beads and both proteins were successfully resolved. By a single injection, 200 μg of protein was loaded and quantitatively eluted from the column. The protein recovery increased with increasing flow rate and salt concentration of the elution buffer and decreased with the increasing protein feed concentration. During the albumin elution, theoretical plate numbers up to 30,000 plates/m were achieved by increasing the salt concentration.
Keywords: Hydroxyethyl methacrylate; Affinity chromatography; High-performance liquid chromatography; Cibacron blue; Albumin; Lysozyme
Determination of steryl sulphates in invertebrate tissue by liquid chromatography–tandem mass spectrometry by Renato R. Neto; Anu Thompson; George A. Wolff (pp. 938-946).
A method for the identification and quantification of underivatised steryl sulphates in invertebrates by liquid chromatography (LC) coupled with tandem mass spectrometry (MS) involving a single cleanup step has been developed. Negative electrospray ionisation and positive and negative atmospheric-pressure chemical ionisation (APCI) spectra of steryl sulphate showed pseudomolecular ions ([M+H–H2SO4]+or [M–H]−). Collision-induced dissociation (CID) was efficient only in positive APCI. LC-MS in negative APCI was least susceptible to interference and possible differences in response factors. The detection limits (signal-to-noise ratio of 3) based on cholest-5-enyl-3-sulphate in positive and negative APCI modes are 3.66 and 0.73 pmol μL−1, respectively. Calibration plots and response factors for cholest-5-enyl-3-sulphate relative to the internal standard, cholecalciferyl-3-sulphate, in both positive and negative polarities, were linear in the concentration range from 1.22 to 16.4 pmol μL−1 with good coefficients of determination (R 2>0.98). It is suggested that the structure elucidation of steryl sulphates is best achieved in CID positive APCI mode, whereas their quantification should be carried out using negative APCI.
Keywords: Steryl sulphates; Invertebrates; Atmospheric-pressure chemical ionisation; Electrospray ionisation; Liquid chromatography–tandem mass spectrometry
Determination of steryl sulphates in invertebrate tissue by liquid chromatography–tandem mass spectrometry by Renato R. Neto; Anu Thompson; George A. Wolff (pp. 938-946).
A method for the identification and quantification of underivatised steryl sulphates in invertebrates by liquid chromatography (LC) coupled with tandem mass spectrometry (MS) involving a single cleanup step has been developed. Negative electrospray ionisation and positive and negative atmospheric-pressure chemical ionisation (APCI) spectra of steryl sulphate showed pseudomolecular ions ([M+H–H2SO4]+or [M–H]−). Collision-induced dissociation (CID) was efficient only in positive APCI. LC-MS in negative APCI was least susceptible to interference and possible differences in response factors. The detection limits (signal-to-noise ratio of 3) based on cholest-5-enyl-3-sulphate in positive and negative APCI modes are 3.66 and 0.73 pmol μL−1, respectively. Calibration plots and response factors for cholest-5-enyl-3-sulphate relative to the internal standard, cholecalciferyl-3-sulphate, in both positive and negative polarities, were linear in the concentration range from 1.22 to 16.4 pmol μL−1 with good coefficients of determination (R 2>0.98). It is suggested that the structure elucidation of steryl sulphates is best achieved in CID positive APCI mode, whereas their quantification should be carried out using negative APCI.
Keywords: Steryl sulphates; Invertebrates; Atmospheric-pressure chemical ionisation; Electrospray ionisation; Liquid chromatography–tandem mass spectrometry
Development of a new biosensor for superoxide radicals by Emel Emregül (pp. 947-954).
A superoxide dismutase (SOD) biosensor for determination of superoxide radicals has been developed by immobilization of superoxide dismutase within gelatin (G) on a Pt electrode surface. The properties of the biosensor have been investigated and optimum conditions–enzyme concentration, glutaraldehyde concentration, and pH–were determined. The response of the G-SOD biosensor was proportional to $$O^{{ ullet - }}_{2} $$ concentration and the detection limit was 0.01 mmol L−1 at a signal-to-noise ratio of 3. The biosensor retained 89% and 60% of its sensitivity after use for three and four weeks, respectively. Immobilization of SOD on gelatin provides a biocompatible microenvironment around the enzyme and stabilizes the activity of the enzyme very efficiently. The superoxide dismutase biosensor was used to determine the antioxidant properties of acetylsalicylic acid-based drugs and the anti-radical activity of healthy and cancerous human brain tissues.
Keywords: Superoxide dismutase; Gelatin; Biosensor; Free radicals; Scavengers
Development of a new biosensor for superoxide radicals by Emel Emregül (pp. 947-954).
A superoxide dismutase (SOD) biosensor for determination of superoxide radicals has been developed by immobilization of superoxide dismutase within gelatin (G) on a Pt electrode surface. The properties of the biosensor have been investigated and optimum conditions–enzyme concentration, glutaraldehyde concentration, and pH–were determined. The response of the G-SOD biosensor was proportional to % MathType!Translator!2!1!AMS LaTeX.tdl!TeX -- AMS-LaTeX!% MathType!MTEF!2!1!+-% feaaeaart1ev0aaatCvAUfeBSjuyZL2yd9gzLbvyNv2CaerbbjxAHX% garmWu51MyVXgatuuDJXwAK1uy0HwmaeHbfv3ySLgzG0uy0Hgip5wz% aebbnrfifHhDYfgasaacH8qrps0lbbf9q8WrFfeuY-Hhbbf9v8qqaq% Fr0xc9pk0xbba9q8WqFfea0-yr0RYxir-Jbba9q8aq0-yq-He9q8qq% Q8frFve9Fve9Ff0dmeaabaqaciGacaGaaeqabaWaaeWaeaaakeaaie% aacaWFpbWaa0baaSqaaiaaikdaaeaacqGHIaYTcqGHsislaaaaaa!3BC4! $$O^{{ ullet - }}_{2} $$ concentration and the detection limit was 0.01 mmol L−1 at a signal-to-noise ratio of 3. The biosensor retained 89% and 60% of its sensitivity after use for three and four weeks, respectively. Immobilization of SOD on gelatin provides a biocompatible microenvironment around the enzyme and stabilizes the activity of the enzyme very efficiently. The superoxide dismutase biosensor was used to determine the antioxidant properties of acetylsalicylic acid-based drugs and the anti-radical activity of healthy and cancerous human brain tissues.
Keywords: Superoxide dismutase; Gelatin; Biosensor; Free radicals; Scavengers
A miniaturized active sampler for the assessment of personal exposure to nitrogen dioxide by Norbert Staimer; Ralph J. Delfino; Charles Bufalino; Philip M. Fine; Constantinos Sioutas; Michael T. Kleinman (pp. 955-962).
A personalized, miniaturized air sampling system was evaluated to estimate the daily exposure of pediatric asthmatics to nitrogen dioxide (NO2). The lightweight device (170 g) uses a sampling pump connected to a solid sorbent tube containing triethanolamine (TEA)-impregnated molecular sieve. The pump is powered by a 9 V battery and samples air over a 24 h period at a collection rate of 0.100 L/min. After exposure, the solid sorbent is removed from the tubes for spectrophotometric analysis (Griess Assay). The lower detection limit of the overall method for NO2 is 11 μg/m3. The linearity, precision and accuracy of the sampler was evaluated. Different NO2 concentrations generated in the laboratory (range: 50 to 340 μg/m3) were simultaneously measured by the TEA tube samplers and colocated continuous chemiluminescent NOx analyzers (reference method). The coefficient of determination for the laboratory test derived from ordinary linear regression (OLR) was r 2=0.99 (y OLR=0.94x−4.58) and the precision 3.6%. Further, ambient NO2 concentrations in the field (range: 10–120 μg/m3) were verified with continuous chemiluminescent monitors next to the active samplers. Reweighted least squares analysis (RLS) based on the least median squares procedure (LMS) resulted in a correlation of r 2=0.68 for a field comparison in Riverside, CA (y RLS=1.01x−0.94) and r 2=0.92 in Los Angeles, CA (y RLS=1.31x−7.12). The precision of the TEA tube devices was 7.4% (at 20–60 μg/m3 NO2) under outdoor conditions. Data show that the performance of this small active sampling system was satisfactory for measuring environmental concentrations of NO2 under laboratory and field conditions. It is useful for personal monitoring of NO2 in environmental epidemiology studies where daily measurements are desired.
Keywords: Air pollution; Exposure assessment; Nitrogen oxides; Active personal sampler; Asthma in children
A miniaturized active sampler for the assessment of personal exposure to nitrogen dioxide by Norbert Staimer; Ralph J. Delfino; Charles Bufalino; Philip M. Fine; Constantinos Sioutas; Michael T. Kleinman (pp. 955-962).
A personalized, miniaturized air sampling system was evaluated to estimate the daily exposure of pediatric asthmatics to nitrogen dioxide (NO2). The lightweight device (170 g) uses a sampling pump connected to a solid sorbent tube containing triethanolamine (TEA)-impregnated molecular sieve. The pump is powered by a 9 V battery and samples air over a 24 h period at a collection rate of 0.100 L/min. After exposure, the solid sorbent is removed from the tubes for spectrophotometric analysis (Griess Assay). The lower detection limit of the overall method for NO2 is 11 μg/m3. The linearity, precision and accuracy of the sampler was evaluated. Different NO2 concentrations generated in the laboratory (range: 50 to 340 μg/m3) were simultaneously measured by the TEA tube samplers and colocated continuous chemiluminescent NOx analyzers (reference method). The coefficient of determination for the laboratory test derived from ordinary linear regression (OLR) was r 2=0.99 (y OLR=0.94x−4.58) and the precision 3.6%. Further, ambient NO2 concentrations in the field (range: 10–120 μg/m3) were verified with continuous chemiluminescent monitors next to the active samplers. Reweighted least squares analysis (RLS) based on the least median squares procedure (LMS) resulted in a correlation of r 2=0.68 for a field comparison in Riverside, CA (y RLS=1.01x−0.94) and r 2=0.92 in Los Angeles, CA (y RLS=1.31x−7.12). The precision of the TEA tube devices was 7.4% (at 20–60 μg/m3 NO2) under outdoor conditions. Data show that the performance of this small active sampling system was satisfactory for measuring environmental concentrations of NO2 under laboratory and field conditions. It is useful for personal monitoring of NO2 in environmental epidemiology studies where daily measurements are desired.
Keywords: Air pollution; Exposure assessment; Nitrogen oxides; Active personal sampler; Asthma in children
High-performance liquid chromatography–tandem mass spectrometry method for quantifying sulfonylurea herbicides in human urine: reconsidering the validation process by Samuel E. Baker; Anders O. Olsson; Larry L. Needham; Dana B. Barr (pp. 963-976).
We have developed a method for measuring 17 sulfonylurea (SU) herbicides in human urine. Urine samples were extracted using solid phase extraction (SPE), preconcentrated, and analyzed by high-performance liquid chromatography–tandem mass spectrometry using turboionspray atmospheric pressure ionization. Carbon 13-labeled ethametsulfuron methyl was used as an internal standard. Chromatographic retention times were under 7 minutes. Total throughput was estimated as >100 samples per day. Because only one labeled internal standard was available for the analysis, we were forced to reconsider and restructure the validation process to include stringent stability tests and analyses of urine matrices of differing compositions. We describe our restructured validation process and the critical evaluation it provides for the method developed. The limits of detection (LOD) ranged from 0.05 μg/L to 0.10 μg/L with an average LOD of 0.06 μg/L. Average total relative standard deviations were 17%, 12% and 8% at 0.1 μg/L, 3.0 μg/L and 10 μg/L, respectively. Average extraction efficiencies of the SPE cartridges were 87% and 86% at 2.5 μg/L and 25 μg/L, respectively. Chemical degradation in acetonitrile and urine was monitored over 250 days. Estimated days for 10% and 50% degradation in urine and acetonitrile ranged from 0.7 days to >318 days. The influence of matrix effects on precision and accuracy was also explored.
Keywords: Validation; Sulfonyl urea; Mass spectrometry; Urine
High-performance liquid chromatography–tandem mass spectrometry method for quantifying sulfonylurea herbicides in human urine: reconsidering the validation process by Samuel E. Baker; Anders O. Olsson; Larry L. Needham; Dana B. Barr (pp. 963-976).
We have developed a method for measuring 17 sulfonylurea (SU) herbicides in human urine. Urine samples were extracted using solid phase extraction (SPE), preconcentrated, and analyzed by high-performance liquid chromatography–tandem mass spectrometry using turboionspray atmospheric pressure ionization. Carbon 13-labeled ethametsulfuron methyl was used as an internal standard. Chromatographic retention times were under 7 minutes. Total throughput was estimated as >100 samples per day. Because only one labeled internal standard was available for the analysis, we were forced to reconsider and restructure the validation process to include stringent stability tests and analyses of urine matrices of differing compositions. We describe our restructured validation process and the critical evaluation it provides for the method developed. The limits of detection (LOD) ranged from 0.05 μg/L to 0.10 μg/L with an average LOD of 0.06 μg/L. Average total relative standard deviations were 17%, 12% and 8% at 0.1 μg/L, 3.0 μg/L and 10 μg/L, respectively. Average extraction efficiencies of the SPE cartridges were 87% and 86% at 2.5 μg/L and 25 μg/L, respectively. Chemical degradation in acetonitrile and urine was monitored over 250 days. Estimated days for 10% and 50% degradation in urine and acetonitrile ranged from 0.7 days to >318 days. The influence of matrix effects on precision and accuracy was also explored.
Keywords: Validation; Sulfonyl urea; Mass spectrometry; Urine
Classifying wine according to geographical origin via quadrupole-based ICP–mass spectrometry measurements of boron isotope ratios by Paul P. Coetzee; Frank Vanhaecke (pp. 977-984).
The potential of quadrupole-based ICP–MS as a tool for B-isotopic analysis of wines and its usefulness in provenance determinations were assessed. A precision of 0.1–0.25% RSD (corresponding to a relative standard deviation of the mean of three replicate measurements of 0.06–0.12%) was sufficient to establish small differences in the B isotope ratios in wines from different geographical origins. Each sample measurement was bracketed by measurements of a standard and mass bias drift correction made by interpolation. Sample preparation was kept to a minimum to avoid possible fractionation. Dilution of the wine samples by a factor of 100 with 0.65% HNO3 was found to reduce matrix-induced mass discrimination substantially. Wines from three wine-producing regions, Stellenbosch, Robertson, and Swartland, in the Western Cape Province of South Africa, and wines from specific regions in France (Bergerac) and Italy (Valpolicella) were analyzed by ICP–QMS for their B-isotopic compositions. It was concluded that the 11B/10B ratios can be used to characterize wines from different geographical origins. Average 11B/10B ratios in red wines from South Africa (Stellenbosch), France (Bergerac), and Italy (Valpolicella) were found to differ by between 0.5 and 1.5%.
Keywords: ICP–mass spectrometry; Boron isotope ratio; Wine provenance
Classifying wine according to geographical origin via quadrupole-based ICP–mass spectrometry measurements of boron isotope ratios by Paul P. Coetzee; Frank Vanhaecke (pp. 977-984).
The potential of quadrupole-based ICP–MS as a tool for B-isotopic analysis of wines and its usefulness in provenance determinations were assessed. A precision of 0.1–0.25% RSD (corresponding to a relative standard deviation of the mean of three replicate measurements of 0.06–0.12%) was sufficient to establish small differences in the B isotope ratios in wines from different geographical origins. Each sample measurement was bracketed by measurements of a standard and mass bias drift correction made by interpolation. Sample preparation was kept to a minimum to avoid possible fractionation. Dilution of the wine samples by a factor of 100 with 0.65% HNO3 was found to reduce matrix-induced mass discrimination substantially. Wines from three wine-producing regions, Stellenbosch, Robertson, and Swartland, in the Western Cape Province of South Africa, and wines from specific regions in France (Bergerac) and Italy (Valpolicella) were analyzed by ICP–QMS for their B-isotopic compositions. It was concluded that the 11B/10B ratios can be used to characterize wines from different geographical origins. Average 11B/10B ratios in red wines from South Africa (Stellenbosch), France (Bergerac), and Italy (Valpolicella) were found to differ by between 0.5 and 1.5%.
Keywords: ICP–mass spectrometry; Boron isotope ratio; Wine provenance
Solid-phase extraction and purification for the quantification of polycyclic aromatic hydrocarbon metabolites in fish bile by Olivier Mazéas; Hélène Budzinski (pp. 985-990).
An analytical protocol including solid-phase extraction and purification is described for the individual quantification of polycyclic aromatic hydrocarbon metabolites (hydroxylated PAHs) in liquid biological matrices such as plasma and bile. The method consists in an enzymatic deconjugation followed by a solid-phase extraction on a C18 cartridge and by a cleanup on an NH2 cartridge. Extracts are then submitted to a derivatization step before gas chromatography/mass spectrometry (GC/MS) analysis. The quantification of PAH metabolites is ensured by adding an internal standard, 1-hydroxypyrene deuterated, at the beginning of the protocol. Recoveries obtained for the entire protocol were for the major part of the compounds between 96 and 70%. However, recoveries were not so satisfying concerning 2-hydroxybiphenyl and especially 3-hydroxybenzo(a)pyrene, with 62 and 36% respectively. Finally, the protocol was applied to different fish bile samples and showed its good applicability to fish bile samples. The NH2 cleanup step has been proved to be a very selective purification step, necessary to remove most of the bile pigments before GC/MS injection. Different PAH metabolites could be detected in those natural samples and their quantification allowed us to distinguish different levels of fish exposure.
Keywords: Hydroxylated polycyclic aromatic hydrocarbons; Solid-phase extraction; Purification; Gas chromatography/mass spectrometry; Fish bile; Metabolites
Solid-phase extraction and purification for the quantification of polycyclic aromatic hydrocarbon metabolites in fish bile by Olivier Mazéas; Hélène Budzinski (pp. 985-990).
An analytical protocol including solid-phase extraction and purification is described for the individual quantification of polycyclic aromatic hydrocarbon metabolites (hydroxylated PAHs) in liquid biological matrices such as plasma and bile. The method consists in an enzymatic deconjugation followed by a solid-phase extraction on a C18 cartridge and by a cleanup on an NH2 cartridge. Extracts are then submitted to a derivatization step before gas chromatography/mass spectrometry (GC/MS) analysis. The quantification of PAH metabolites is ensured by adding an internal standard, 1-hydroxypyrene deuterated, at the beginning of the protocol. Recoveries obtained for the entire protocol were for the major part of the compounds between 96 and 70%. However, recoveries were not so satisfying concerning 2-hydroxybiphenyl and especially 3-hydroxybenzo(a)pyrene, with 62 and 36% respectively. Finally, the protocol was applied to different fish bile samples and showed its good applicability to fish bile samples. The NH2 cleanup step has been proved to be a very selective purification step, necessary to remove most of the bile pigments before GC/MS injection. Different PAH metabolites could be detected in those natural samples and their quantification allowed us to distinguish different levels of fish exposure.
Keywords: Hydroxylated polycyclic aromatic hydrocarbons; Solid-phase extraction; Purification; Gas chromatography/mass spectrometry; Fish bile; Metabolites
Feasibility of analyzing molecular pigments in paint layers using TOF S–SIMS by Rita Van Ham; Luc Van Vaeck; Freddy Adams; Annemie Adriaens (pp. 991-997).
A first attempt to measure the molecular compositions of pigments in paintings using static SIMS was made. An investigation of pellets of pure pigments such as auripigment and verdigris allowed the detection of numerous high m/z ions useful for molecular identification. Analysis of pigments in embedded paint fragments, on the other hand, only yielded elemental information because of charge build-up and contamination problems. Optimization of the form in which the sample is presented to the analysis method is obviously the price to pay for the ultimate sensitivity and information depth of S-SIMS.
Keywords: Static SIMS; FT LMMS; Pigments; Molecular; Cultural heritage
Feasibility of analyzing molecular pigments in paint layers using TOF S–SIMS by Rita Van Ham; Luc Van Vaeck; Freddy Adams; Annemie Adriaens (pp. 991-997).
A first attempt to measure the molecular compositions of pigments in paintings using static SIMS was made. An investigation of pellets of pure pigments such as auripigment and verdigris allowed the detection of numerous high m/z ions useful for molecular identification. Analysis of pigments in embedded paint fragments, on the other hand, only yielded elemental information because of charge build-up and contamination problems. Optimization of the form in which the sample is presented to the analysis method is obviously the price to pay for the ultimate sensitivity and information depth of S-SIMS.
Keywords: Static SIMS; FT LMMS; Pigments; Molecular; Cultural heritage
Determination of TNT and its metabolites in water samples by voltammetric techniques by York Zimmermann; J. A. C. Broekaert (pp. 998-1002).
Square-wave voltammetry with the hanging drop mercury electrode as the working electrode was used for the determination of ultratraces of explosives in aqueous solution. It was shown that the strong pressure dependence of the pneumatically controlled multimode electrode system of a conventional Metrohm apparatus could be compensated by an additional pressure regulation, through which the pressure variations could be decreased when switching from deaeration to the static measurements. By using square-wave voltammetry with this electrode system after this modification the limits of detection for 2,4,6-trinitrotoluene (TNT) and other TNT-metabolites could be decreased down to 0.2 μg L−1 when using a measurement time of 6 min. Also a simultaneous determination of TNT and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) was shown to be possible over a wide linear range and the detection limits then were 2.2 μg L−1 for TNT and 25 μg L−1 for RDX. By applying the highly stable and adjustable pressure as mentioned before, the calibrations could be kept stable over a period of up to 1 week.
Keywords: Square-wave voltammetry; 2,4,6-Trinitrotoluene; Hanging drop mercury electrode; 2-Amino-4,6-dinitrotoluene; Hexogen; Picric acid
Determination of TNT and its metabolites in water samples by voltammetric techniques by York Zimmermann; J. A. C. Broekaert (pp. 998-1002).
Square-wave voltammetry with the hanging drop mercury electrode as the working electrode was used for the determination of ultratraces of explosives in aqueous solution. It was shown that the strong pressure dependence of the pneumatically controlled multimode electrode system of a conventional Metrohm apparatus could be compensated by an additional pressure regulation, through which the pressure variations could be decreased when switching from deaeration to the static measurements. By using square-wave voltammetry with this electrode system after this modification the limits of detection for 2,4,6-trinitrotoluene (TNT) and other TNT-metabolites could be decreased down to 0.2 μg L−1 when using a measurement time of 6 min. Also a simultaneous determination of TNT and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) was shown to be possible over a wide linear range and the detection limits then were 2.2 μg L−1 for TNT and 25 μg L−1 for RDX. By applying the highly stable and adjustable pressure as mentioned before, the calibrations could be kept stable over a period of up to 1 week.
Keywords: Square-wave voltammetry; 2,4,6-Trinitrotoluene; Hanging drop mercury electrode; 2-Amino-4,6-dinitrotoluene; Hexogen; Picric acid
A comparison of accelerated solvent extraction, Soxhlet extraction, and ultrasonic-assisted extraction for analysis of terpenoids and sterols in tobacco by Jinchao Shen; Xueguang Shao (pp. 1003-1008).
The performance of accelerated solvent extraction in the analysis of terpenoids and sterols in tobacco samples was investigated and compared with those of Soxhlet extraction and ultrasonically assisted extraction with respect to yield, extraction time, reproducibility and solvent consumption. The results indicate that although the highest yield was achieved by Soxhlet extraction, ASE appears to be a promising alternative to classical methods since it is faster and uses less solvent, especially when applied to the investigation of large batch tobacco samples. However, Soxhlet extraction is still the preferred method for analyzing sterols since it gives a higher extraction efficiency than other methods.
Keywords: Accelerated solvent extraction; Terpenoids; Sterols; Tobacco
A comparison of accelerated solvent extraction, Soxhlet extraction, and ultrasonic-assisted extraction for analysis of terpenoids and sterols in tobacco by Jinchao Shen; Xueguang Shao (pp. 1003-1008).
The performance of accelerated solvent extraction in the analysis of terpenoids and sterols in tobacco samples was investigated and compared with those of Soxhlet extraction and ultrasonically assisted extraction with respect to yield, extraction time, reproducibility and solvent consumption. The results indicate that although the highest yield was achieved by Soxhlet extraction, ASE appears to be a promising alternative to classical methods since it is faster and uses less solvent, especially when applied to the investigation of large batch tobacco samples. However, Soxhlet extraction is still the preferred method for analyzing sterols since it gives a higher extraction efficiency than other methods.
Keywords: Accelerated solvent extraction; Terpenoids; Sterols; Tobacco
Voltammetric analysis using a self-renewable non-mercury electrode by Peter Surmann; Hanan Zeyat (pp. 1009-1013).
Galinstan is a new kind of electrode material and the galinstan electrode is a promising alternative to the commonly used mercury electrodes. The eutectic mixture of gallium, indium and tin is liquid at room temperature (m.p. −19°C) and its voltammetric behaviour is similar to that of mercury. The potential windows of use were determined for different pH values and are similar to those obtained with conventional mercury electrodes. Furthermore, the high hydrogen overpotential, which is characteristic for mercury, can be observed when galinstan is used as electrode material. Galinstan can be employed as a liquid electrode in the voltammetric analysis of different metal ions, such as lead and cadmium, in different supporting electrolytes. Our results indicate that the non-toxic liquid alloy galinstan could therefore become immensely important in electrochemical research as a potential surrogate material for mercury.
Keywords: Galinstan; Voltammetry; Potential window
Voltammetric analysis using a self-renewable non-mercury electrode by Peter Surmann; Hanan Zeyat (pp. 1009-1013).
Galinstan is a new kind of electrode material and the galinstan electrode is a promising alternative to the commonly used mercury electrodes. The eutectic mixture of gallium, indium and tin is liquid at room temperature (m.p. −19°C) and its voltammetric behaviour is similar to that of mercury. The potential windows of use were determined for different pH values and are similar to those obtained with conventional mercury electrodes. Furthermore, the high hydrogen overpotential, which is characteristic for mercury, can be observed when galinstan is used as electrode material. Galinstan can be employed as a liquid electrode in the voltammetric analysis of different metal ions, such as lead and cadmium, in different supporting electrolytes. Our results indicate that the non-toxic liquid alloy galinstan could therefore become immensely important in electrochemical research as a potential surrogate material for mercury.
Keywords: Galinstan; Voltammetry; Potential window
Determination of paralytic shellfish poisoning toxins in cultured microalgae by high-performance liquid chromatography with fluorescence detection by Hong-Zhi He; Hua-Bin Li; Yue Jiang; Feng Chen (pp. 1014-1017).
A novel method for the determination of paralytic shellfish poisoning (PSP) toxins using high-performance liquid chromatography with fluorescence detection was developed. The fluorescent derivates of neosaxitoxin (neoSTX), saxitoxin (STX), gonyautoxins 1 and 4 (GTX1+4), and gonyautoxins 2 and 3 (GTX2+3) were separated on a μBondapak NH2 column (300 mm × 3.9 mm, 10 μm) using water and acetate buffer (pH 6.5) as the mobile phase (1.00 mL min−1) in gradient mode with fluorescence detection at 390 nm (excitation at 330 nm). The linear ranges of neoSTX, STX, GTX1+4 and GTX2+3 were 3.31–331, 0.952–95.2, 3.78–378 and 0.124–12.4 ng mL−1, respectively. The detection limits of neoSTX, STX, GTX1+4 and GTX2+3 were 1.10, 0.32, 1.26 and 0.041 ng mL−1, respectively. The method was successfully applied to the determination of PSP toxins in microalgae. The recoveries ranged from 88±2% to 107±4% and the relative standard deviations were 0.16% to 4.4%. The procedure is also environmentally friendly because no organic solvent is used in the mobile phase.
Keywords: Paralytic shellfish poisoning toxins; High performance liquid chromatography; Fluorescence detection; Microalgae
Determination of paralytic shellfish poisoning toxins in cultured microalgae by high-performance liquid chromatography with fluorescence detection by Hong-Zhi He; Hua-Bin Li; Yue Jiang; Feng Chen (pp. 1014-1017).
A novel method for the determination of paralytic shellfish poisoning (PSP) toxins using high-performance liquid chromatography with fluorescence detection was developed. The fluorescent derivates of neosaxitoxin (neoSTX), saxitoxin (STX), gonyautoxins 1 and 4 (GTX1+4), and gonyautoxins 2 and 3 (GTX2+3) were separated on a μBondapak NH2 column (300 mm × 3.9 mm, 10 μm) using water and acetate buffer (pH 6.5) as the mobile phase (1.00 mL min−1) in gradient mode with fluorescence detection at 390 nm (excitation at 330 nm). The linear ranges of neoSTX, STX, GTX1+4 and GTX2+3 were 3.31–331, 0.952–95.2, 3.78–378 and 0.124–12.4 ng mL−1, respectively. The detection limits of neoSTX, STX, GTX1+4 and GTX2+3 were 1.10, 0.32, 1.26 and 0.041 ng mL−1, respectively. The method was successfully applied to the determination of PSP toxins in microalgae. The recoveries ranged from 88±2% to 107±4% and the relative standard deviations were 0.16% to 4.4%. The procedure is also environmentally friendly because no organic solvent is used in the mobile phase.
Keywords: Paralytic shellfish poisoning toxins; High performance liquid chromatography; Fluorescence detection; Microalgae