Check out our New Publishers' Select for Free Articles

Analytical and Bioanalytical Chemistry (v.397, #2)

Hans-Joachim Kuss and Stavros Kromidas (Eds.): Quantification in LC and GC. A practical guide to good chromatographic data by Yoshihiro Saito (pp. 407-408).

Stephen L.R. Ellison, Vicki J. Barwick, Trevor J. Duguid Farrant: Practical statistics for the analytical scientist. A bench guide, 2nd ed. by Jürgen W. Einax (pp. 409-410).

Twelfth international symposium on biological and environmental reference materials (BERM 12) by Mike Sargent (pp. 411-411).

is an analytical chemist by profession, with 40 years’ experience of research and development, laboratory and project management, analytical quality and reference materials. His current post at LGC Ltd, a private company formed from the UK’s Laboratory of the Government Chemist, is that of Chief Chemical Metrologist. He has played a leading role in developing and gaining acceptance for the UK’s chemical metrology infrastructure and in the development of international chemical metrology. Mike Sargent represents the UK at MetChem, the chemistry committee of EURAMET, and at the Consultative Committee on Amount of Substance (CCQM) of the International Committee of Weights and Measures. He has chaired the Inorganic Analysis Working Group of the CCQM since 1997. He has authored or contributed to an extensive range of scientific books and papers.

History of reference materials for food and nutrition metrology: as represented in the series of BERM symposia by Wayne R. Wolf (pp. 413-421).

joined the then Human Nutrition Division of the Agricultural Research Service, USDA, Beltsville, MD, USA, in 1971. He has been involved as a research chemist in analytical method development to determine components in foods and biological materials, mainly focusing on trace elements and vitamins. His present research activity focuses on the development of analytical methods for determining water-soluble vitamins in foods and dietary supplements. He initiated and founded a series of ten International Symposia on Biological and Environmental Reference Materials (BERM), commencing in 1983, and has developed significant recognition for these activities in the international analytical community. Figure Uncertainty interval percentages (UIP) in certified reference materials as a function of nutrient concentration. P proximates, C carbohydrates, M minerals, T trace elements, VS water soluble vitamins, VF fat-soluble vitamins, A amino acids, F fatty acids, O other

Marine mammal blubber reference and control materials for use in the determination of halogenated organic compounds and fatty acids by John R. Kucklick; Michele M. Schantz; Rebecca S. Pugh; Barbara J. Porter; Dianne L. Poster; Paul R. Becker; Teri K. Rowles; Stefan Leigh; Stephen A. Wise (pp. 423-432).

The National Institute of Standards and Technology (NIST) has a diverse collection of control materials derived from marine mammal blubber, fat, and serum. Standard Reference Material (SRM) 1945 Organics in Whale Blubber was recertified for polychlorinated biphenyl (PCB) congeners, organochlorine pesticides, and polybrominated diphenyl ether (PBDE) congeners. SRM 1945 has also been assigned mass fraction values for compounds not frequently determined in marine samples including toxaphene congeners, coplanar PCBs, and methoxylated PBDE congeners which are natural products. NIST also has assigned mass fraction values, as a result of interlaboratory comparison exercises, for PCB congeners, organochlorine pesticides, PBDE congeners, and fatty acids in six homogenate materials produced from marine mammal blubber or serum. The materials are available from NIST upon request; however, the supply is very limited for some of the materials. The materials include those obtained from pilot whale blubber (Homogenates III and IV), Blainville’s beaked whale blubber (Homogenate VII), polar bear fat (Homogenate VI), and California sea lion serum (Marine Mammal Control Material-1 Serum) and blubber (Homogenate V).

Keywords: Persistent organic pollutants; Reference materials; Marine mammal; Fatty acids

Determination of butoxyacetic acid (biomarker of ethylene glycol monobutyl ether exposure) in human urine candidate reference material by Ilona Šperlingová; Ludmila Dabrowská; Vladimír Stránský; Šárka Dušková; Jan Kučera; Monika Tvrdíková; Milon Tichy (pp. 433-438).

Ethylene glycol monobutyl ether (EGBE), an industrial solvent, is absorbed by the body not only by inhalation but also by dermal absorption (liquid or vapour). EGBE is metabolized to butoxyacetic acid (BAA). Pooled freeze-dried urine candidate reference material (RM) was prepared from urine obtained from persons occupationally exposed to EGBE. This material has the advantage of containing butoxyacetic acid in both the free and conjugated (glutamine and glycine) forms, as found in native urine. In all GC method modifications used, acid hydrolysis was used to release BAA from its conjugated form. The amount of butoxyacetic acid in homogeneity and stability testing was measured by GC after derivatisation with N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide. Detection was by MS in EI mode, in the authors’ laboratory. For interlaboratory comparison of the reference material GC methods with MS, FID, and ECD were used. Different extraction solvents (dichloromethane–isopropanol 2:1, ethyl acetate, or dichloromethane) and derivatisation reagents (trimethylsilyldiazomethane, N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide) were used. Using ANOVA (at the statistical level α = 0.05) no changes were found in the concentration of butoxyacetic acid during fifteen month isochronous stability testing, or in homogeneity testing. The uncertainty contributions were u _h = 8.8 mg L⁻¹ and u _s = 6.5 mg L⁻¹. The concentration of butoxyacetic acid in freeze-dried urine RM was evaluated from the results of eight laboratory data sets within an interlaboratory comparison by use of the interactive statistical software IPECA. The contribution to total uncertainty derived from interlaboratory comparison was u _i = 12.7 mg L⁻¹. The reference value (c = 273 ± 33 mg L⁻¹) is an unweighted arithmetic average of accepted results. The value is traceable to the pure butoxyacetic acid (98% w/w; Acros Organic #257760010) used as calibrant. The uncertainty given is combined expanded uncertainty derived from the results from interlaboratory comparison, and from homogeneity and stability tests (k = 2). The reference material will be used to verify method performance in the biological monitoring of occupational exposure to EGBE.

Keywords: Candidate reference material; Ethylene glycol monobutyl ether; Metabolite; Butoxyacetic acid; Human urine

Determination of perfluorinated alkyl acid concentrations in human serum and milk standard reference materials by Jennifer M. Keller; Antonia M. Calafat; Kayoko Kato; Mark E. Ellefson; William K. Reagen; Mark Strynar; Steven O’Connell; Craig M. Butt; Scott A. Mabury; Jeff Small; Derek C. G. Muir; Stefan D. Leigh; Michele M. Schantz (pp. 439-451).

Standard Reference Materials (SRMs) are certified reference materials produced by the National Institute of Standards and Technology that are homogeneous materials well characterized with values for specified properties, such as environmental contaminant concentrations. They can be used to validate measurement methods and are critical in improving data quality. Disagreements in perfluorinated alkyl acid (PFAA) concentrations measured in environmental matrices during past interlaboratory comparisons emphasized the need for SRMs with values assigned for PFAAs. We performed a new interlaboratory comparison among six laboratories and provided, for the first time, value assignment of PFAAs in SRMs. Concentrations for perfluorooctane sulfonate (PFOS), perfluorooctanoate (PFOA), and other PFAAs in two human serum and two human milk SRMs are reported. PFAA concentration measurements agreed for serum SRM 1957 using different analytical methods in six laboratories and for milk SRM 1954 in three laboratories. The interlaboratory relative standard deviation for PFOS in SRM 1957 was 7%, which is an improvement over past interlaboratory studies. Matrix interferences are discussed, as well as temporal trends and the percentage of branched vs. linear isomers. The concentrations in these SRMs are similar to the present-day average concentrations measured in human serum and milk, resulting in representative and useful control materials for PFAA human monitoring studies.

Keywords: Perfluorinated contaminants; Organic contaminants; Reference materials; Human samples; Blood; Intercomparison exercise

Degradation kinetics of the Alternaria mycotoxin tenuazonic acid in aqueous solutions by David Siegel; Stefan Merkel; Wolfram Bremser; Matthias Koch; Irene Nehls (pp. 453-462).

The degradation kinetics of the Alternaria mycotoxin tenuazonic acid (l-TA) in aqueous buffer were studied over a period of 4 months at different pH levels (3.5 and 7.0) and temperatures (4, 25 and 40 °C). l-TA and its degradation products were quantified by newly developed high-performance liquid chromatography methods with UV or electrospray multistage mass spectrometry detection. At pH 3.5, significant degradation occurred at 25 and 40 °C, the respective l-TA half-lives being 73.8 ± 0.4 and 14.0 ± 0.1 days. Two degradation processes, epimerization and hydrolysis, were evaluated kinetically. The hydrolytically formed iso-deacetyl TA (iso-DTA, epimeric mixture) was found to be the stable end product of l-TA degradation under the conditions of this study. This indicates that iso-DTA as well as the l-TA epimer u-TA are formed in aqueous beverage matrices.

Keywords: Tenuazonic acid; Alternaria ; Epimerization; Degradation; Kinetics

Characterization of NIES CRM No. 23 Tea Leaves II for the determination of multielements by Ikuko Mori; Miyuki Ukachi; Kimiyo Nagano; Hiroyasu Ito; Jun Yoshinaga; Masataka Nishikawa (pp. 463-470).

A candidate environmental certified reference material (CRM) for the determination of multielements in tea leaves and materials of similar matrix, NIES CRM No. 23 Tea Leaves II, has been developed and characterized by the National Institute for Environmental Studies (NIES), Japan. The origin of the material was tea leaves, which were ground, sieved through a 106-µm mesh, homogenized, and then subdivided into amber glass bottles. The results of homogeneity and stability tests indicated that the material was sufficiently homogeneous and stable for use as a reference material. The property values of the material were statistically determined based on chemical analyses by a network of laboratories using a wide range of methods. Sixteen laboratories participated in the characterization, and nine certified values and five reference values were obtained. These property values of the candidate CRM, which are expressed as mass fractions, were close to the median and/or mean values of the mass fractions of elements in various tea products. The candidate CRM is appropriate for use in analytical quality control and in the evaluation of methods used in the analysis of tea and materials of similar matrix. Figure New NIES CRM No. 23 Tea Leaves II.

Keywords: Certified reference material; Quality assurance/control; Trace elements; Aluminum; Fluoride; Water-soluble component

Simultaneous determination of water-soluble vitamins in SRM 1849 Infant/Adult Nutritional Formula powder by liquid chromatography–isotope dilution mass spectrometry by Robert J. Goldschmidt; Wayne R. Wolf (pp. 471-481).

Assessing dietary intake of vitamins from all sources, including foods, dietary supplements, and fortified foods, would be aided considerably by having analytical methodologies that are capable of simultaneous determination of several vitamins. Vitamins naturally present in foods may occur in different chemical forms, with levels ranging over several orders of magnitude. Vitamins in dietary supplements and fortified foods, however, are typically added in a single chemical form, and matrix issues are usually not as complex. These sources should thus be relatively amenable to approaches that aim for simultaneous determination of multiple vitamins. Our recent work has focused on development of liquid chromatography (LC)–UV/fluorescence and LC–tandem mass spectrometry methods for the simultaneous determination of water-soluble vitamins (thiamine, niacin, pyridoxine, pantothenic acid, folic acid, biotin, and riboflavin) in dietary supplement tablets and fortified foods, such as formula powders and breakfast cereals. As part of the validation of our methods and collaboration in characterization of a new NIST SRM 1849 Infant/Adult Nutritional Formula powder, we report data on SRM 1849 using isotope dilution mass spectrometric methods. Use of available NIST Standard Reference Materials® as test matrices in our method development and validation gives a benchmark for future application of these methods. We compare three chromatographic approaches and provide data on stability of vitamin standard solutions for LC-based multiple vitamin determinations. Figure Extracted ion chromatograms of seven vitamins using RP chromatography treatment

Keywords: Multivitamin analysis; Isotope dilution mass spectrometry; Liquid chromatography–mass spectrometry; Reference material

Separation of 26 toxaphene congeners and measurement in air particulate matter SRMs compared to technical toxaphene SRM 3067 by Stacy S. Vander Pol; John R. Kucklick; Stefan D. Leigh; Barbara J. Porter; Michele M. Schantz (pp. 483-492).

Toxaphene is a complex technical mixture that has been found ubiquitously in the environment but has caused issues for analysis, especially of individual congeners. This paper reports the elution order of 26 major toxaphene congeners on three gas chromatographic columns. The three different stationary phases generally had similar elution orders for the toxaphene congeners, but fewer co-elutions occurred on a low-bleed, low-polarity column. These congeners (except for two that co-eluted and were not added to the calibration mixture) were examined in air particulate matter standard reference materials (SRMs), 1648a, 1649a, and 1649b as well as SRM 3067 toxaphene in methanol for assignment of reference values. SRM 3067 had mass fractions an order of magnitude greater than the air particulate SRMs, which ranged from 0.568 ± 0.018 ng g⁻¹ dry mass (B9-2006 in SRM 1648a) to 12.9 ± 0.20 ng g⁻¹ dry mass (B9-715 (P 58) in SRM 1649a). The three air particulate SRMs all had different mass fractions and proportions of congeners relative to the sum of the toxaphene congeners. SRM 3067 may be useful as a technical mixture toxaphene congener calibrant. SRMs 1648a and 1649b will serve as reference materials for the analysis of 21 (three congeners were not included due to values below the detection limit or a potential polychlorinated biphenyl co-elution) toxaphene congeners in atmospheric particulate samples. Figure Comparison (mean±SD) of proportions of toxaphene congeners to the sum of the congeners in air particulate matter SRMs 1649a, 1649b, and 1648a and compared to SRM 3067 toxaphene in methanol. Among the air particulate matter SRMs, all congeners were significantly different (P≤0.0006) with most significantly different among all three SRMs (a indicates an SRM with greater proportion than the others for that congener). The comparisons between the air particulate matter SRMs and SRM 3067 were significant (P<0.002) except as indicated by n.s.

Keywords: Reference material; Air particulate; Toxaphene; Column separation

Preparation and certification of arsenate [As(V)] reference material, NMIJ CRM 7912-a by Tomohiro Narukawa; Takayoshi Kuroiwa; Izumi Narushima; Yasujiro Jimbo; Toshihiro Suzuki; Koichi Chiba (pp. 493-499).

Arsenate [As(V)] solution reference material, National Metrology Institute of Japan (NMIJ) certified reference material (CRM) 7912-a, for speciation of arsenic species was developed and certified by NMIJ, the National Institute of Advanced Industrial Science and Technology. High-purity As₂O₃ reagent powder was dissolved in 0.8 M HNO₃ solution and As(III) was oxidized to As(V) with HNO₃ to prepare 100 mg kg^-1 of As(V) candidate CRM solution. The solution was bottled in 400 bottles (50 mL each). The concentration of As(V) was determined by four independent analytical techniques—inductively coupled plasma mass spectrometry, inductively coupled plasma optical emission spectrometry, graphite furnace atomic absorption spectrometry, and liquid chromatography inductively coupled plasma mass spectrometry—according to As(V) calibration solutions, which were prepared from the arsenic standard of the Japan Calibration Service system and whose species was guaranteed to be As(V) by NMIJ. The uncertainties of all the measurements and preparation procedures were evaluated. The certified value of As(V) in the CRM is (99.53 ± 1.67) mg kg^-1 (k = 2).

Keywords: Certification; Certified reference material; As(V); Arsenic species; Speciation

Certification of drugs of abuse in a human serum standard reference material: SRM 1959 by Susan S.-C. Tai; Jocelyn L. Prendergast; Lorna T. Sniegoski; Michael J. Welch; Karen W. Phinney; Nien Fan Zhang (pp. 501-509).

A new standard reference material (SRM) for drugs of abuse in human serum (SRM 1959) has been developed. This SRM is intended to be used as a control material for laboratories performing analysis of drugs of abuse in blood to evaluate the accuracy of their methods. SRM 1959 is a frozen human serum material fortified with seven compounds for which analyses are performed to determine evidence of illegal drug use: benzoylecgonine (BZE), methadone (METH), methamphetamine (MAMP), morphine (MOR), nordiazepam (NOR), phencyclidine (PCP), and 11-nor-Δ⁹-tetrahydrocannabinol-9-carboxylic acid (THC-9-COOH). Two independent methods involving isotope dilution (ID)-gas chromatography/mass spectrometry (GC/MS) and ID-liquid chromatography/mass spectrometry (LC/MS) were used for the value assignment. For THC-9-COOH, an additional measurement using LC/tandem mass spectrometry (LC/MS/MS) was also included. All methods used isotopically labeled compounds as internal standards and solid-phase extractions to isolate the analytes from the serum. The GC/MS methods used different clean-up procedures from those used for the LC/MS-based methods. Repeatability with within-set coefficients of variation (CVs) ranged from 0.5% to 4.3% for the GC/MS methods and from 0.2% to 1.2% for the LC/MS-based methods. Intermediate precision with between-set CVs for all the methods ranged from 0.1% to 1.1%. Agreement between the GC/MS and LC/MS methods ranged from 0.8% to 8.8%. The results from the methods were combined to obtain the certified concentrations and their expanded uncertainties.

Keywords: Drugs of abuse; Standard reference material; SRM; Isotope dilution; Gas chromatography/mass spectrometry; GC/MS; Liquid chromatography/mass spectrometry; LC/MS; Liquid chromatography/tandem mass spectrometry; LC/MS/MS; Solid-phase extraction

Certification of NIST standard reference material 2389a, amino acids in 0.1 mol/L HCl—quantification by ID LC-MS/MS by Mark S. Lowenthal; James Yen; David M. Bunk; Karen W. Phinney (pp. 511-519).

An isotope-dilution liquid chromatography-tandem mass spectrometry (ID LC-MS/MS) measurement procedure was developed to accurately quantify amino acid concentrations in National Institute of Standards and Technology (NIST) Standard Reference Material (SRM) 2389a—amino acids in 0.1 mol/L hydrochloric acid. Seventeen amino acids were quantified using selected reaction monitoring on a triple quadrupole mass spectrometer. LC-MS/MS results were compared to gravimetric measurements from the preparation of SRM 2389a—a reference material developed at NIST and intended for use in intra-laboratory calibrations and quality control. Quantitative mass spectrometry results and gravimetric values were statistically combined into NIST-certified mass fraction values with associated uncertainty estimates. Coefficients of variation (CV) for the repeatability of the LC-MS/MS measurements among amino acids ranged from 0.33% to 2.7% with an average CV of 1.2%. Average relative expanded uncertainty of the certified values including Types A and B uncertainties was 3.5%. Mean accuracy of the LC-MS/MS measurements with gravimetric preparation values agreed to within |1.1|% for all amino acids. NIST SRM 2389a will be available for characterization of routine methods for amino acid analysis and serves as a standard for higher-order measurement traceability. This is the first time an ID LC-MS/MS methodology has been applied for quantifying amino acids in a NIST SRM material.

Keywords: Amino acids; Standard reference material; Mass spectrometry; Isotope dilution; LC-MS/MS

Standardization of ceruloplasmin measurements is still an issue despite the availability of a common reference material by Ilenia Infusino; Cristina Valente; Alberto Dolci; Mauro Panteghini (pp. 521-525).

The purpose of measurement standardization is to achieve closer comparability of results obtained using different commercial systems. Regarding serum protein immunoassays, a reference preparation (BCR-470) was released in 1993 and adopted by manufacturers across the world to value-assign their assay calibrators for routine methods to reduce method-dependent variation. Moving from nephelometric (Beckman Immage 800) to turbidimetric determination (Roche Cobas c 501) of seven serum proteins, we preliminarily checked the comparability of results between the two systems. The study was performed according to the CLSI EP9-A protocol on 30 fresh sera, tested on each system in duplicate, and subdivided on two different days, without recalibration and using manufacturers’ control materials to validate the runs. Both manufacturers’ package inserts provide statements that kit calibrators are traceable to BCR-470. Suggested reference intervals are also the same. Although a fairly good correlation was observed (r = 0.955), the comparison of ceruloplasmin methods produced evidence of highly significant proportional (regression slope, 0.572) and constant bias (intercept, 0.05 g/L). Absolute and percentage mean differences were −0.11 g/L (95% confidence interval (CI) −0.13 to −0.10 g/L) and −39.1% (CI −43.1 to −35.2%), respectively. No other evaluated proteins showed similar problems. Lacking a ceruloplasmin reference method, it is impossible to demonstrate that one of the two assays produces true ceruloplasmin values. The problem is, however, that results coming from the two assays are clearly not comparable. This may be either due to a lack of commutability of the reference material with biological samples in the evaluated assays or to calibration problems by manufacturers in one of the stages of the calibration hierarchy.

Keywords: Ceruloplasmin; Nephelometry; Immunologic techniques; Standardization; Reference materials; Biological samples

Colloquium Spectroscopicum Internationale XXXVI, Budapest (Hungary), August 30–September 3, 2009 by Gyula Záray; Jürgen W. Einax (pp. 527-528).

is professor at the Eötvös University of Budapest and Head of the Department of Analytical Chemistry. He has been director of the Hungarian Satellite Centre of Trace Element Institute for UNESCO since 2000. In 2005, he founded the Cooperative Research Centre of Environmental Sciences at Eötvös University. In 1994, he cofounded the Hungarian Spectrochemical Society and has been its president since 1999. He is the European Editor of Applied Spectroscopy Reviews and a member of the editorial board of Microchemical Journal. His research interests focus on the development of solid sampling spectrometric methods for the analytical investigation of ceramics, metal alloys and environmental matrices; investigation of heavy metal uptake, accumulation and translocation processes in plants; speciation of heavy metals in urban aerosols, soils, sediments and food materials; biomonitoring of surface waters; chemical characterisation of biofilms; determination of organic micropollutants (pharmaceutical residues, personal care products) in waste, surface and drinking water. is professor of Analytical Chemistry at the Friedrich Schiller University of Jena and Head of the Department of Environmental Analysis. His main areas of research include not only trace analytical problems in the environment, but also the application of chemometrics to environmental research. He is a member of the editorial board of Microchemical Journal. Furthermore, he is a member of the executive board of the Division Analytical Chemistry in the German Chemical Society and Chairman of the Working Group for Chemometrics and Laboratory Data Processing in this division.

Phthalates determination in pharmaceutical formulae used in parenteral nutrition by LC-ES-MS: importance in public health by Cristina Pérez-Feás; María Carmen Barciela-Alonso; Antía Sedes-Díaz; Pilar Bermejo-Barrera (pp. 529-535).

A method for determining a group of phthalate esters in pharmaceutical formulae used in parenteral nutrition samples (with and without vitamins) has been developed. The phthalic acid esters (PAEs) studied were dimethyl phthalate, diethyl phthalate, butyl benzyl phthalate, dibutyl phthalate, di-(2-ethylhexyl) phthalate, and dioctyl phthalate. This group of phthalates was determined by high performance liquid chromatography (HPLC)–electrospray ionization–mass spectrometry, working in positive ion mode. The phthalates analyzed were extracted from the sample using hexane and sodium hydroxide. The hexane was then evaporated, and the compounds were redissolved in acetonitrile. The compounds were separated by HPLC working in gradient mode with acetonitrile-ultrapure water starting from 5% to 75% acetonitrile in 5 min, followed by isocratic elution for 27 min. Standard calibration curves were linear for all the analytes over the concentration range 10–250 µg L⁻¹. The method was precise (with RSD from 3.3% to 12.9%) and sensitive. The proposed analytical method has been applied to the analysis of these compounds in different pharmaceutical formulae (with different compositions) for parenteral nutrition samples in order to check the presence of phthalates and determine their concentration. Figure Parenteral nutrition sample stored in an ethyl vinyl acetate bag.

Keywords: Phthalates; LC-ES-MS; Parenteral nutrition

Novel derivatisation technique for the determination of chlorophenoxy acid type herbicides by gas chromatography–mass spectrometry by Erik Maloschik; Mária Mörtl; András Székács (pp. 537-548).

The analytical detection of chlorophenoxycarboxylic-acid-type herbicides (2,4-D, dichloprop, MCPA, etc.) in environmental samples is often a problem in instrumental analysis, as these compounds containing free carboxylic groups require chemical derivatisation prior to gas chromatographic (GC) methods. Nine chlorophenoxy-acid-type herbicide active ingredients have been derivatised successfully with trimethylsilyl N,N-dimethyl carbamate and t-butyldimethylsilyl N,N-dimethyl carbamate by forming their trimethylsilyl (TMS) and t-butyldimethylsilyl (TBDMS) esters, respectively. The detection and determination of the derivatives were performed by capillary gas chromatography–mass spectrometry. The study included determination of retention indices, mass spectral properties and comparison of derivatives produced. The mass spectra of TBDMS derivatives are usually dominated by very characteristic ions [M-57]⁺ resulting from the cleavage of t-butyl moiety during electron impact (EI) ionisation in the mass spectrometer. Limits of detection were 5 to 100 pg applying GC with EI-MS detection in full scan mode. The method, using SPE sample preparation, was applied for the analysis of 115 ground water and surface water samples collected in Békés County, Hungary in 2009.

Keywords: Silylation; Chlorophenoxy acids; Residue analysis; Derivatisation; Gas chromatography; Mass spectrometry; Silyl carbamates

Superoxide dismutase mimicking Cu(II)–mixed amino acid complexes covalently grafted onto silica gel—an FT-IR study by Zita Csendes; Valéria Bugris; Linda Lackó; Imre Labádi; János T. Kiss; István Pálinkó (pp. 549-555).

Our recent work concerning the synthesis, characterisation and testing of bioinspired electron transfer catalysts is described in this contribution. The catalysts were various Cu(II) complexes having mixed C- or N-protected amino acids (l-histidine and l-tyrosine) as ligands covalently grafted onto surface-modified silica gel. The resulting materials were structurally characterised by FT-IR spectroscopy, and their superoxide dismutase activities were tested. The covalently anchored Cu(II) complexes displayed appreciable activities in the test reaction; thus, they may be considered as promising candidates as durable electron transfer catalysts approaching the efficiency of the enzyme mimicked. Figure Although the active site mimicking, covalently anchored Cu-mixed amino acid complexes are less effective catalysts than the superoxide dismutase enzyme, but they can work under harsh experimental conditions as well

Keywords: Enzyme mimics; Immobilisation; Structural characterisation; Superoxide dismutase activity; IR spectroscopy

Novel fabrication of silver-coated glass capillaries for ready SERS-based detection of dissolved chemical species by Ji Won Lee; Hyang Bong Lee; Kwan Kim; Kuan Soo Shin (pp. 557-562).

A novel method to deposit a highly surface-enhanced Raman scattering (SERS) active silver film onto the inside surface of a glass capillary is developed. Firstly, Ag sol was synthesized by the reaction of AgNO₃ with poly-(ethylenimine) (PEI), and then toluene and benzenethiol (BT) were added into the sol. The mixture was flowed through the glass capillary to obtain the SERS-active Ag film-coated glass capillary. The SERS activity of the Ag-coated capillary was dependent on the amount of PEI and BT used. In addition, BT could be easily desorbed from the Ag surface by treating it with a borohydride solution, maintaining the initial SERS activity. The SERS enhancement factor at 632.8-nm excitation was estimated to be on the order of 10⁶. The detection limits of adenine and dipicolinic acid were then as low as 1.0 × 10⁻⁸ and 1.0 × 10⁻⁷ M, respectively, based on an S/N ratio of 3. This clearly suggests that the Ag-coated capillary is an invaluable device for the analysis of effluent chemicals by SERS. Figure A highly SERS active Ag-coated glass capillary is developed that can be used in microanalysis of effluent chemicals.

Keywords: Adenine; Dipicolinic acid; Poly(ethylenimine); Silver nanoparticle; Surface-enhanced Raman scattering

Evaluation of sample preparation methods for elastomer digestion for further halogens determination by Diogo P. Moraes; Juliana S. F. Pereira; Liange O. Diehl; Márcia F. Mesko; Valderi L. Dressler; José Neri G. Paniz; Guenter Knapp; Érico M. M. Flores (pp. 563-570).

In this work, three sample preparation methods were evaluated for further halogen determination in elastomers containing high concentrations of carbon black. Samples of nitrile-butadiene rubber, styrene-butadiene rubber, and ethylene-propylene-diene monomer elastomers were decomposed using oxygen flask combustion and microwave-induced combustion (MIC) for further Br and Cl determination by ion chromatography (IC), inductively coupled plasma optical emission spectrometry (ICP OES), and inductively coupled plasma mass spectrometry (ICP-MS). Extraction assisted by microwave radiation in closed vessels was also evaluated using water or alkaline solution. Digestion by MIC was carried out using 50 mmol l⁻¹ (NH₄)₂CO₃ as the absorbing solution. The effect of the reflux step was also evaluated. Accuracy was evaluated using certified reference materials with polymeric matrix composition and by comparison of results using neutron activation analysis. Agreement for Br and Cl was better than 95% by MIC using 5 min of reflux, and no statistical difference was found using IC, ICP OES, and ICP-MS for determination of both analytes. For MIC, the relative standard deviation (RSD) was lower than 5%. Using extraction in closed vessels, a high amount of residues was observed, and recoveries were lower than 45% for both analytes. For oxygen flask combustion, the agreement was similar using MIC but RSD was higher (20%). The residual carbon content, an important parameter used to evaluate the digestion efficiency, was always below 1% for MIC. Using MIC, it was possible to digest elastomers with high efficiency, resulting in a single solution suitable for halogen determination by different techniques.

Keywords: Microwave-induced combustion; Elastomers; Halogen determination; Sample preparation; IC; ICP-MS

Layer-by-layer assembly of small interfering RNA and poly(ethyleneimine) for substrate-mediated electroporation with high efficiency by Hiroyuki Fujimoto; Koichi Kato; Hiroo Iwata (pp. 571-578).

Electroporation microarrays have been developed for the high-throughput transfection of expression constructs and small interfering RNAs (siRNAs) into living mammalian cells. These techniques have potential to provide a platform for the cell-based analysis of gene functions. One of the key issues associated with microarray technology is the efficiency of transfection. The capability of attaining reasonably high transfection efficiency is the basis for obtaining functional data without false negatives. In this study, we aimed at improving the transfection efficiency in the system that siRNA loaded on an electrode is electroporated into cells cultured directly on the electrode. The strategy we adopted here is to increase the surface density of siRNA loaded onto electrodes. For this purpose, the layer-by-layer assembly of siRNA and cationic polymers, branched or linear form of poly(ethyleneimine), was performed. The multilayer thus obtained was characterized by infrared reflection-adsorption spectroscopy and surface plasmon resonance analysis. Transfection efficiency was evaluated in a system that siRNA specific for enhanced green fluorescent protein (EGFP) was electroporated on the electrode into human embryonic kidney cells stably transformed with the EGFP gene. The suppression of EGFP expression was assessed by fluorescence microscopy and flow cytometry. Our data showed that the layer-by-layer assembly of siRNA with branched poly(ethyleneimine) facilitated to increase the surface density of loaded siRNA. As a result, the expression of EGFP gene in the electroporated cells was suppressed much more on the electrodes with the multilayer of siRNA than that with the monolayer. Figure LBL assembly of siRNA and PEI for surface-mediated electroporation

Keywords: RNA interference; Silencing; Gene transfer; Cell-based microarray; High-throughput screening

Pyridinium aldoxime analysis by HPLC: the method for studies on pharmacokinetics and stability by P. Szegi; H. Kalász; R. Laufer; K. Kuca; K. Tekes (pp. 579-586).

Reversed-phase separation of various pyridinium aldoximes requires a certain concentration of ion-pairing agent, as their chemical structures contain two quaternary amines in the pyridinium ring. Adequate mobile phase is scouted on the basis of retention of pyridinium aldoxime (using the graph of k′ versus concentration of an ion-pairing agent) compared to the chromatogram of the background peaks originated from the homogenate. Change in the ion-pairing agent concentration was more expressed for the elution of K-203 than that of the background peaks from the serum, brain and cerebrospinal fluid. Stability of K-203 was investigated using HPLC. Determination of K-203 in tissue samples requires homogenization using either trichloroacetic acid or perchloric acid. Fast degradation takes place at acidic pH. Adjusting pH to neutral in the possible shortest time frame helps to avoid degradation. Degradation of K-203 was easily followed by HPLC separation and monitoring the elution with an ultraviolet absorbance detector at 276 nm. Amperometric detection indicates only the decrease of K-203 content.

Keywords: Pyridinium aldoxime; HPLC; Ion-pairing agents; K-203

Novel molecular tumour classification using MALDI–mass spectrometry imaging of tissue micro-array by Marie-Claude Djidja; Emmanuelle Claude; Marten F. Snel; Simona Francese; Peter Scriven; Vikki Carolan; Malcolm R. Clench (pp. 587-601).

The development of tissue micro-array (TMA) technologies provides insights into high-throughput analysis of proteomics patterns from a large number of archived tumour samples. In the work reported here, matrix-assisted laser desorption/ionisation–ion mobility separation–mass spectrometry (MALDI–IMS–MS) profiling and imaging methodology has been used to visualise the distribution of several peptides and identify them directly from TMA sections after on-tissue tryptic digestion. A novel approach that combines MALDI–IMS–MSI and principal component analysis–discriminant analysis (PCA–DA) is described, which has the aim of generating tumour classification models based on protein profile patterns. The molecular classification models obtained by PCA–DA have been validated by applying the same statistical analysis to other tissue cores and patient samples. The ability to correlate proteomic information obtained from samples with known and/or unknown clinical outcome by statistical analysis is of great importance, since it may lead to a better understanding of tumour progression and aggressiveness and hence improve diagnosis, prognosis as well as therapeutic treatments. The selectivity, robustness and current limitations of the methodology are discussed.

Keywords: Tumour classification; Tissue micro-array; Pancreatic cancer; MALDI imaging; Ion mobility separation

Validation of a two-dimensional gas chromatography mass spectrometry method for the simultaneous quantification of cannabidiol, Δ⁹-tetrahydrocannabinol (THC), 11-hydroxy-THC, and 11-nor-9-carboxy-THC in plasma by Erin L. Karschner; Allan J. Barnes; Ross H. Lowe; Karl B. Scheidweiler; Marilyn A. Huestis (pp. 603-611).

A sensitive analytical method for simultaneous quantification of sub-nanogram concentrations of cannabidiol (CBD), Δ⁹-tetrahydrocannabinol (THC), 11-hydroxy-THC (11-OH-THC), and 11-nor-9-carboxy-THC (THCCOOH) in plasma is presented for monitoring cannabinoid pharmacotherapy and illicit cannabis use. Analytes were extracted from 1 mL plasma by solid-phase extraction, derivatized with N,O-bis(trimethylsilyl) trifluoroacetamide with 1% trimethylchlorosilane, and analyzed by two-dimensional gas chromatography mass spectrometry (2D-GCMS) with cryofocusing. The lower calibration curve was linear from 0.25–25 ng/mL for CBD and THC, 0.125-25 ng/mL for 11-OH-THC and 0.25-50 ng/mL for THCCOOH. A second higher linear range from 5–100 ng/mL, achieved through modification of injection parameters, was validated for THC, 11-OH-THC, and THCCOOH and was only implemented if concentrations exceeded the lower curve upper limit of linearity. This procedure prevented laborious re-extraction by allowing the same specimen to be re-injected for quantification on the high calibration curve. Intra- and inter-assay imprecision, determined at four quality control concentrations, were ≤7.8% CV. Analytical bias was within ±9.2% of target and extraction efficiencies were ≥72.9% for all analytes. Analytes were stable when stored at 22°C for 16 h, 4°C for 48 h, after three freeze–thaw cycles at −20°C and when stored on the autosampler for 48 h. This sensitive and specific 2D-GCMS assay provides a new means of simultaneously quantifying CBD, THC and metabolite biomarkers in clinical medicine, forensic toxicology, workplace drug testing, and driving under the influence of drugs programs.

Keywords: Δ⁹-Tetrahydrocannabinol; Cannabidiol; Sativex®; Two-dimensional chromatography; Plasma

Evaluation of the performance of sensors based on optical imaging of a chemically sensitive layer by Corrado Di Natale; Marco Santonico; Roberto Paolesse; Daniel Filippini; Arnaldo D’Amico; Ingemar Lundström (pp. 613-621).

Interest in the use of the optical properties of chemical indicators is growing steadily. Among the optical methods that can be used to capture changes in sensing layers, those producing images of large-area devices are particularly interesting for chemical sensor array development. Until now, few studies addressed the characterization of image sensors from the point of view of their chemical sensor application. In this paper, a method to evaluate such performance is proposed. It is based on the simultaneous measurement of absorption events in a metalloporphyrin layer with an image sensor and a quartz microbalance (QMB). Exploiting the well-known behaviour of QMB, comparison of signals enables estimation of the minimum amount of absorbed molecules that the image sensor can detect. Results indicate that at the single pixel level a standard image sensor (for example a webcam) can easily detect femtomoles of absorbed molecules. It should therefore be possible to design sensor arrays in which the pixels of images of large-area sensing layers are regarded as individual chemical sensors providing a ready and simple method for large sensor array development.

Keywords: Chemical sensing; Optical sensing; Computer screen photo-assisted technique; Metalloporphyrins

Complete chromatographic separation of steroids, including 17α and 17β-estradiols, using a carbazole-based polymeric organic phase in both reversed and normal-phase HPLC by Abul K. Mallik; Kaori Shingo; Usha Ghimire Gautam; Tsuyoshi Sawada; Makoto Takafuji; Hirotaka Ihara (pp. 623-629).

Poly(2-N-carbazolylethyl acrylate) with terminal trimethoxysilyl groups was prepared as an organic phase and immobilized onto silica. The retention behavior of the column packed with this carbazole-based polymer-immobilized silica (Sil-CEA) was investigated by using various estrogenic steroids and corticoids in both reversed-phase and normal-phase liquid chromatography. As a result, complete separation was confirmed for eight kinds of steroids with Sil-CEA. The most specific separation with Sil-CEA can be emphasized by the high separation factor (e.g., α = 1.39 in methanol–water (7:3, v/v) at 35 °C) for 17α and 17β-estradiols, one of the most difficult pairs of isomers in chromatographic separation, whereas for two kinds of commercially available polymeric ODS columns as references α = 1.01, only, under the same conditions. Because the excellent separation and retention order with Sil-CEA was maintained even in a normal-phase mobile phase such as a hexane–2-propanol, it is estimated that the CEA phase has multiple interaction mechanisms through stronger interactions such as dipole–dipole, carbonyl–π, and hydrogen bonding interactions than the hydrophobic effect expected with ODS.

Keywords: Carbazole-based polymer; Separation; Steroids; Solvent effect

Bovine liver slices combined with an androgen transcriptional activation assay: an in-vitro model to study the metabolism and bioactivity of steroids by S. Wang; J. C. W. Rijk; J. H. Riethoff-Poortman; S. Van Kuijk; A. A. C. M. Peijnenburg; T. F. H. Bovee (pp. 631-641).

Previously we described the properties of a rapid and robust yeast androgen bioassay for detection of androgenic anabolic compounds, validated it, and showed its added value for several practical applications. However, biotransformation of potent steroids into inactive metabolites, or vice versa, is not included in this screening assay. Within this context, animal-friendly in-vitro cellular systems resembling species-specific metabolism can be of value. We therefore investigated the metabolic capacity of precision-cut slices of bovine liver using 17β-testosterone (T) as a model compound, because this is an established standard compound for assessing the metabolic capacity of such cellular systems. However, this is the first time that slice metabolism has been combined with bioactivity measurements. Moreover, this study also involves bioactivation of inactive prohormones, for example dehydroepiandrosterone (DHEA) and esters of T, and although medium extracts are normally analyzed by HPLC, here the metabolites formed were identified with more certainty by ultra-performance liquid chromatography time-of-flight mass spectrometry (UPLC–TOFMS) with accurate mass measurement. Metabolism of T resulted mainly in the formation of the less potent phase I metabolites 4-androstene-3,17-dione (4-AD), the hydroxy-T metabolites 6α, 6β, 15β, and 16α-OH-T, and the phase II metabolite T-glucuronide. As a consequence the overall androgenic activity, as determined by the yeast androgen bioassay, decreased. In order to address the usefulness of bovine liver slices for activation of inactive steroids, liver slices were exposed to DHEA and two esters of T. This resulted in an increase of androgenic activity, because of the formation of 4-AD and T. Figure Bovine liver slices for exposure studies in a 6-well format.

Keywords: Bovine liver slices; Mass spectrometry; Metabolism; Histology; Yeast androgen bioassay

Qualitative screening of phenolic compounds in olive leaf extracts by hyphenated liquid chromatography and preliminary evaluation of cytotoxic activity against human breast cancer cells by Shaoping Fu; David Arráez-Roman; Antonio Segura-Carretero; Javier A. Menéndez; María P. Menéndez-Gutiérrez; Vicente Micol; Alberto Fernández-Gutiérrez (pp. 643-654).

In this work, high-performance liquid chromatography (HPLC) coupled to electrospray time-of-flight mass spectrometry (ESI-TOF-MS) and electrospray ion trap multiple-stage tandem mass spectrometry (ESI-IT-MS²) has been applied to screen phenolic compounds in olive leaf extracts. The use of a small particle size C18 column (1.8 μm) provided great resolution and made separation of a lot of isomers possible. The structural characterization was based on accurate mass data obtained by ESI-TOF-MS, and the nature of fragmentation ions were further confirmed by ESI-IT-MS² when possible. In addition, we employed tetrazolium salt (MTT)-based assays to assess the effects of olive leaf extracts on the growth of human tumor-derived cells. Upon this approach, we achieved an accurate profile of olive leaf phenolics along with the identification of several important isomers of secoiridoids and flavonoids. This will allow a better understanding of the complete composition of olive-leaf-bioactive compounds as well as their involvement in Olea europaea L. biochemical pathways. Importantly, olive leaf extracts exhibited dose-dependent inhibitory effects on the metabolic status (cell viability) of three breast cancer models in vitro. Since the tumoricidal activity of the extracts should be mainly attributed to the identified olive leaf phenolics, these findings warrant further investigation at the structure–function molecular level to definitely establish the anticancer value of these phytochemicals. OAF Survival plots obtained by using the MTT assay for SKBr3, MCF-7 and JIMT-1 breast cancer cell lines treated with olive leaf extract at increasing concentrations for 72 h.

Keywords: Olive leaf; Phenolic compounds; High-performance liquid chromatography; Time-of-flight mass spectrometry; Ion trap multiple-stage tandem mass spectrometry; Breast cancer

Modification of tricine–SDS–PAGE for online and offline analysis of phosphoproteins by ICP-MS by Syed R. Haider; Helen J. Reid; Barry L. Sharp (pp. 655-664).

This study describes a modification to tricine sodium dodecyl sulphate polyacrylamide gel electrophoresis to make it more effective for the separation of low molecular mass proteins and for coupling with inductively coupled plasma mass spectrometry (ICP-MS). The modified method employs low-percentage polyacrylamide gels (7–10%) (w/v) and low reagent concentrations that provide efficient separations, good quantitation and low matrix levels that are compatible with ICP-MS. Using phosphopeptides as a model system, and offline analysis, we obtained recoveries of 73% (w/v) in a 9% gel compared with 55% in a conventional 16% gel. Online coupling was achieved by modification of a standard commercially available gel electroelution apparatus and casting of the gel into a 7.3-cm-long tube. Online separation of a digest of β-casein was demonstrated with recovery of the mono- and tetraphosphopeptides, which were identified by comparison with peptide standards. A mass balance study with the standards yielded recoveries of 95% for tetraphosphopeptides and 48% for monophosphopeptides. The factors affecting the separations and recoveries are discussed in detail. The detection limits for 10-µL samples of the mono- and tetraphosphopeptides were 0.7 µM (7 pmol) and 0.2 µM (2 pmol) respectively.

Keywords: Inductively coupled plasma mass spectrometry; Tricine sodium dodecyl sulphate polyacrylamide gel electrophoresis; Phosphopeptide; Quantification; Online coupling; Gel percentage

Targeted LC–MS derivatization for aldehydes and carboxylic acids with a new derivatization agent 4-APEBA by Mark Eggink; Maikel Wijtmans; Ansgar Kretschmer; Jeroen Kool; Henk Lingeman; Iwan J. P. de Esch; Wilfried M. A. Niessen; Hubertus Irth (pp. 665-675).

Based on the template of a recently introduced derivatization reagent for aldehydes, 4-(2-(trimethylammonio)ethoxy)benzeneaminium dibromide (4-APC), a new derivatization agent was designed with additional features for the analysis and screening of biomarkers of lipid peroxidation. The new derivatization reagent, 4-(2-((4-bromophenethyl)dimethylammonio)ethoxy)benzenaminium dibromide (4-APEBA) contains a bromophenethyl group to incorporate an isotopic signature to the derivatives and to add additional fragmentation identifiers, collectively enhancing the abilities for detection and screening of unknown aldehydes. Derivatization can be achieved under mild conditions (pH 5.7, 10 °C). By changing the secondary reagent (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide instead of sodium cyanoborohydride), 4-APEBA is also applicable to the selective derivatization of carboxylic acids. Synthesis of the new label, exploration of the derivatization conditions, characterization of the fragmentation of the aldehyde and carboxylic acid derivatives in MS/MS, and preliminary applications of the labeling strategy for the analysis of aldehydes in urine and plasma are described. Figure Structure and MS/MS fragmentation spectrum of 4-APEBA reagents derivatized with octanoic acid

Keywords: Derivatization; Aldehydes; Carboxylic acids; 4-APEBA; 4-APC; LC–MS/MS; Lipid peroxidation

Experimental design for optimization of microwave-assisted extraction of benzodiazepines in human plasma by P. Fernández; C. Vázquez; R. A. Lorenzo; A. M. Carro; I. Álvarez; P. Cabarcos (pp. 677-685).

A simple and fast microwave-assisted-extraction (MAE) method has been evaluated as an alternative to solid-phase extraction (SPE) for the determination of six benzodiazepines widely prescribed in European countries (alprazolam, bromazepam, diazepam, lorazepam, lormetazepam and tetrazepam) in human plasma. For MAE optimization a Doehlert experimental design was used with extraction time, temperature and solvent volume as influential parameters. A desirability function was employed in addition to the simultaneous optimization of the MAE conditions. The analysis of variance showed that the solvent volume had a positive influence on the extraction of all the analytes tested, achieving a statistically significant effect. Also, the extraction time had a statistically significant effect on the extraction of four benzodiazepines. The selected MAE conditions—89 °C, 13 min and 8 mL of chloroform/2-propanol (4:1, v/v)—led to recoveries between 89.8 ± 0.3 and 102.1 ± 5.2% for benzodiazepines using a high performance liquid chromatography method coupled with diode-array detection. The comparison of MAE and SPE shows better results for MAE, with a lower number of steps in handling the sample and greater efficiency. The applicability of MAE was successfully tested in 27 plasma samples from benzodiazepine users.

Keywords: Benzodiazepines; Human plasma; Microwave-assisted extraction; Experimental design; High-performance liquid chromatography–diode-array detection

Development and validation of a dried blood spot–HPLC assay for the determination of metronidazole in neonatal whole blood samples by Maysa Faisal Suyagh; Godwill Iheagwaram; Prashant Laxman Kole; Jeff Millership; Paul Collier; Henry Halliday; James C. McElnay (pp. 687-693).

A selective and sensitive high-performance liquid chromatography method with UV detection for the determination of metronidazole in dried blood spots (DBS) has been developed and validated. DBS samples [spiked or patient samples] were prepared by applying blood (30 µL) to Guthrie cards. Discs (6 mm diameter) were punched from the cards and extracted using water containing the internal standard, tinidazole. The extracted sample was chromatographed without further treatment using a reversed phase system involving a Symmetry® C18 (5 µm, 3.9 × 150 mm) preceded by a Symmetry® guard column of matching chemistry and a detection wavelength of 317 nm. The mobile phase comprised acetonitrile/0.01 M phosphate solution (KH₂PO₄), pH 4.7, 15:85, v/v, with a flow rate of 1 mL/min. The calibration was linear over the range 2.5–50 mg/mL. The limits of detection and quantification were 0.6 and 1.8 µg/mL, respectively. The method has been applied to the determination of 203 DBS samples from neonatal patients for a phamacokinetic/pharmacodynamic study.

Keywords: Dried blood spots; Guthrie card; Metronidazole; HPLC

Electrochemical probing into cytochrome c modification with homocysteine-thiolactone by Jing Zhao; Wei Zhu; Tao Liu; Jinghua Yang; Genxi Li (pp. 695-701).

Homocysteine thiolactone modification is a unique process of posttranslational protein modification as well as a significant clinical indicator of cardiovascular and neurovascular diseases, so we report a new method in this paper to sensitively monitor such a modification using horse heart cytochrome c as a model protein. After the modification has been confirmed by UV–vis spectroscopy and ESI-MS, N-linked cytochrome c is then covalently assembled onto the surface of a gold electrode via the resulted homocysteine thiol group, thus electrochemical techniques, especially differential pulse voltammetry, have been employed and proven to provide an efficient way to probe into the modification of the protein. While the immobilized protein can exhibit well-defined voltammetric response, the signal of the modified cytochrome c is positively correlated to the concentration of homocysteine-thiolactone. The detectable electrochemical signal can be attained with the minimum concentration of 5 × 10⁻⁵ M homocysteine-thiolactone. Furthermore, screening of N-homocysteinylation inhibitors can be also feasible since the electrochemical waves linearly decrease with the concentration of an inhibitor pyridoxal 5-phosphate. The limit of detection for the inhibitors can be about 1 × 10⁻⁵ M. Figure Schematic illustration of the electrochemical method to probe into the modification of cytochrome c with homocysteine-thiolactone

Keywords: Homocysteine thiolactone; Cytochrome c; Inhibitor; Cyclic voltammetry; Differential pulse voltammetry

Accurate detection of on-state quantum dot and biomolecules in a microfluidic flow with single-molecule two-color coincidence detection by Chun-yang Zhang; Kun Yang (pp. 703-708).

Due to their unique optical and electronic properties, quantum dots (QDs) have been widely used in a variety of biosensors for sensitive detection of biomarkers and small molecules. However, single QD exhibits dynamic fluctuation of fluorescence intensity (i.e., blinking) with the transition between on and off states, which adversely influences the development of QD-based optical biosensors. Therefore, the methods for efficient evaluation of on-state QD are especially important and highly desirable. In this paper, a novel and unique approach based on single-molecule two-color coincidence detection is developed to simply and accurately evaluate the on-state QDs in a microfluidic flow. Our results demonstrate that improved QDs in the on state are detected in a microfluidic flow in comparison with that in the Brownian motion state, thus paving the way to the development of single QD-based biosensors for sensitive detection of low-abundance biomolecules. This single-molecule two-color coincidence detection has been applied for the homegeneous detection of nucleic acids in a microfluidic flow with the detection sensitivity of 5.0 fM. Figure Representative traces of fluorescence bursts from the 525Bead–655QD complex in the Brownian motion state. The fluorescence signals of the 655QDs were shown in red; the fluorescence signals of the 525Beads were shown in blue

Keywords: Single-molecule detection; Two color; Coincidence; Quantum dot; Fluorescence; Microfluidic flow

Absolute quantification of semicarbazide-sensitive amine oxidase in human umbilical artery by single-reaction monitoring with electrospray tandem mass spectrometry by Yongqian Zhang; Shengyuan Xiao; Lin Wang; Hongbin Wang; Yong Zhu; Yujuan Li; Yulin Deng (pp. 709-715).

Semicarbazide-sensitive amine oxidase (SSAO; E.C.1.4.3.6.) is widely distributed in different tissues, particularly in vascular smooth muscle and adipose tissue. Its physiological function remains unclear. Up to now, the common method to determine SSAO is based on enzymatic activity measurement. However, enzymatic activity could be easily influenced by the temperature, pH, and circumstance. In the present study, we have developed the single-reaction monitoring (SRM) approach for measuring the absolute amount of SSAO expression in human umbilical artery based on LC-ESI-MS/MS. The measurement of protein was converted to the measurement of a unique peptide of SSAO from Homo sapiens. The peptide (YQLAVTQR) was confirmed to be unique to the SSAO in human using the ExPasy blast tools, and thus the synthetic peptide was used as the standard which can produce abundant parent ion (m/z = 490.0) and daughter ion (m/z = 687.4) in the mass spectrometry. Trap drive and fragmentation energy of MS/MS of the unique peptide was 60 V and 0.6 V, respectively. The calibration curve was linear over the range of 1.99–127.8 fmol/μL, with 1.99 fmol/μl of the lower limit of quantification. The inter- and intra-day precisions and recoveries for all samples were satisfactory. The results demonstrated SSAO protein concentration was 7.75 fmol/g wet weight. It proved that the novel assay was sensitive and selective to measure the amount of SSAO protein originated from H. sapiens. Figure The workflow of absolute quantification of SSAO by LC-SRM analysis

Keywords: Semicarbazide-sensitive amine oxidase; Absolute quantification; Single-reaction monitoring; ESI/MS/MS

Alternative instrumentation for the analysis of total carbon dioxide (TCO₂) in equine plasma by Mark Jarrett; D. Brynn Hibbert; Roy Osborne; E. Bruce Young (pp. 717-722).

The analysis of total carbon dioxide (TCO₂) in equine plasma is conventionally done in Australia and elsewhere using Beckman Synchron EL-ISE® analysers. This instrument is no longer being manufactured and has not been supported by the supplier since December 2008. For testing for TCO₂ to continue, it is necessary to evaluate and commission alternative instrumentation. In this paper, we compare the Beckman Synchron EL-ISE®, the Beckman Synchron CX®5, the Beckman UniCel DxC®600 and the Randox Daytona™ instruments. Results indicate that all four instruments perform in accordance with the manufacturer’s specifications. The Beckman CX 5, DxC 600 and Randox Daytona instruments are therefore all suitable alternatives for routine screening in a laboratory environment. Only the Randox Daytona instrument is sufficiently ‘portable’, i.e. it can be readily transported and used on-site at a racecourse (typically in a purpose-built modest-size laboratory vehicle). The three Beckman instruments are suitable for ‘confirmatory analysis’ using the quality-accredited method (Racing Science Centre), but the principle of operation of the Randox Daytona instrument may preclude its use in confirmatory analysis. Instrument costs may affect purchase decisions.

Keywords: Horseracing; TCO₂ ; Equine plasma; Instrumentation; Compliance testing

Chiral separation of (+)/(−)-catechin from sulfated and glucuronidated metabolites in human plasma after cocoa consumption by Christina Ritter; Benno F. Zimmermann; Rudolf Galensa (pp. 723-730).

Cocoa is well-known to be rich in flavan-3-ols. Previous analyses have established that alkaline treatment of cocoa beans results in epimerization of (−)-epicatechin to (−)-catechin and (+)-catechin to (+)-epicatechin. Now, the question is whether both epimers can be absorbed by the human organism. This paper describes sample preparation and an HPLC method for chiral determination of (+)/(−)-catechin from sulfated and glucuronidated metabolites in human plasma. The sample preparation includes enzymatic hydrolysis of the catechin metabolites, and solid-phase extraction (SPE). A PM-γ-cyclodextrin column is used with a coulometric electrode-array detection (CEAD) system. The recovery of catechin ranges from 89.9 to 96.8%. The limit of detection is 5.9 ng mL⁻¹ for (−)-catechin and 6.8 ng mL⁻¹ for (+)-catechin, and the limit of quantification is 12.8 ng mL⁻¹ for (−)-catechin and 16.9 ng mL⁻¹ for (+)-catechin. The relative standard deviation of the method ranges from 0.9 to 1.5%. This method was successfully applied to human plasma after consumption of a cocoa drink. In one human self-experiment, (+)-catechin and (−)-catechin were found in human plasma, but metabolism of the two enantiomers differed. Figure Separation of (+)/(-)-catechin in human plasma

Keywords: Flavanols; Cocoa; Chiral; Human plasma; Electrochemical detection; HPLC

Determination of selenomethionine, selenocysteine, and inorganic selenium in eggs by HPLC–inductively coupled plasma mass spectrometry by Elżbieta Lipiec; Grzegorz Siara; Katarzyna Bierla; Laurent Ouerdane; Joanna Szpunar (pp. 731-741).

A method for the simultaneous determination of selenomethionine (SeMet), selenocysteine (SeCys), and selenite [Se(IV)] in chicken eggs was developed. A sample preparation protocol including defatting, protein denaturation, and carbamidomethylation was optimized in order to achieve complete protein digestion and to avoid SeCys losses. Quantification was carried out by reversed-phase HPLC–inductively coupled plasma mass spectrometry (ICP MS) after quantitative isolation of the selenium-containing fraction by size-exclusion liquid chromatography. The detection limits were 0.06, 0.003, and 0.01 µg g⁻¹ (dry weight) for SeCys, Se(IV) and SeMet, respectively, and the precision was 5–10%. The end products of carbamidomethylation of the different selenium species were identified for the first time by electrospray QTOF MS after custom-designed 2D HPLC purification. Differences in selenium speciation in egg yolk and white were highlighted, the yolk containing more SeCys and the white more SeMet. An insight into selenium bioaccessibility in eggs was obtained by digestion with simulated gastric and gastrointestinal juices and size-exclusion HPLC-ICP MS. Figure Selenium speciation in chicken eggs

Keywords: Chicken egg; Supplementation; Selenium; Selenium speciation

Potential use of gamma irradiation in the production of mussel and oyster reference materials for paralytic shellfish poisoning toxins by Andrew D. Turner; Robert G. Hatfield; Andy L. Powell; Wendy Higman (pp. 743-749).

Bivalve shellfish samples containing paralytic shellfish poisoning toxins were subjected to gamma irradiation dosage trials in order to assess the potential suitability of the technique in the production of toxin reference materials. Two candidate reference materials of tissue homogenates, mussels (Mytilus sp.) and native oysters (Ostrea edulis), were prepared in-house. Both were subjected to gamma irradiation at four different dose levels, 3.0, 6.0, 13.0 and 18.1 kGy. Bacterial levels were shown to be eliminated in the mussels and significantly reduced in the oysters following irradiation at all four dose levels. Paralytic shellfish poisoning (PSP) toxin concentrations were not significantly reduced in any of the samples indicating the treatment had no adverse affect on the initial stability of any of the PSP toxins monitored. Chromatographic results showed near-identical profiles for treated and non-treated samples inferring that no fluorescent toxin degradation products or matrix interferences were produced during the irradiation process. Results therefore proved that gamma irradiation treatment reduced bacterial levels within paralytic shellfish poisoning reference materials without compromising analyte content, with the subsequent potential to enhance the stability of future candidate reference materials treated in this manner. Fig. 1 Cefas tubular bag photobioreactor used for mass culture of Alexandrium

Keywords: Paralytic shellfish poisoning; Reference materials; Gamma irradiation; Stability; Shellfish toxins

A sensitive and efficient procedure for the high throughput determination of banned aromatic amines in textiles and leather products aided by advanced sample composition by J. García-Lavandeira; C. Salgado-Petinal; E. Blanco; R. Cela (pp. 751-763).

A highly sensitive procedure for the efficient routine control of aromatic amines derived from banned azo dyes in textile and leather products was developed and optimized. The procedure involves the extraction and reduction of azo dyes from solid samples following the sample preparation protocols outlined in EN 14362-1:2003, EN 14362-2:2003, and ISO/TS 17234:2003 standards, and cleanup and concentration of aromatic amines by solid-phase extraction, with Oasis HLB and ENVI-Carb sorbents in series. The elution was carried out with the cartridges connected in series, although the positions of the cartridges were reversed and the ENVI-Carb cartridge was placed in the backflush mode. Extracted and concentrated aromatic amines were separated and analyzed by liquid chromatography (LC)–tandem mass spectrometry (MS). The chromatographic separation was optimized by means of computer-assisted method development with a special chemometric tool (the PREGA LC-MS module), developed specifically for LC-MS systems. This system enables the unattended optimization of separations after a few priming isocratic and gradient experiments. The optimized separation program enables accurate detection and measurement of all the 23 aromatic amines considered, at very low quantification limits and without any notable matrix effects. Strategic sample composition was applied as an efficient means of reducing the costs and work involved in the control of aromatic amines in finished textile and leather products. The benefits of strategic sample composition are demonstrated by means of a case study of 20 sample specimens.

Keywords: Aromatic amines; Textiles; Liquid chromatography–mass spectrometry; Sample composition

Multi-mycotoxin analysis of maize silage by LC-MS/MS by R. R. Rasmussen; I. M. L. D. Storm; P. H. Rasmussen; J. Smedsgaard; K. F. Nielsen (pp. 765-776).

This paper describes a method for determination of 27 mycotoxins and other secondary metabolites in maize silage. The method focuses on analytes which are known to be produced by common maize and maize-silage contaminants. A simple pH-buffered sample extraction was developed on the basis of a very fast and simple method for analysis of multiple pesticide residues in food known as QuEChERS. The buffering effectively ensured a stable pH in samples of both well-ensiled maize (pH < 4) and of hot spots with fungal infection (pH > 7). No further clean-up was performed before analysis using liquid chromatography–tandem mass spectrometry. The method was successfully validated for determination of eight analytes qualitatively and 19 quantitatively. Matrix-matched calibration standards were used giving recoveries ranging from 37% to 201% with the majority between 60% and 115%. Repeatability (5–27% RSD_r) and intra-laboratory reproducibility (7–35% RSD_IR) was determined. The limit of detection (LOD) for the quantitatively validated analytes ranged from 1 to 739 µg kg⁻¹. Validation results for citrinin, fumonisin B₁ and fumonisin B₂ were unsatisfying. The method was applied to 20 selected silage samples and alternariol monomethyl ether, andrastin A, alternariol, citreoisocoumarin, deoxynivalenol, enniatin B, fumigaclavine A, gliotoxin, marcfortine A and B, mycophenolic acid, nivalenol, roquefortine A and C and zearalenone were detected. Maize silage fed to cows can be contaminated with mycotoxins from pre- and post-harvest fungi. Several metabolites were detected by LC-MS/MS; including zearalenone and mycophenolic acid from Fusarium (red) and Penicillium (green) infections, respectively.

Keywords: Mycotoxins; Maize; Silage; LC-MS/MS; Validation; QuEChERS

Simultaneous quantification of carotenoids, retinol, and tocopherols in forages, bovine plasma, and milk: validation of a novel UPLC method by B. Chauveau-Duriot; M. Doreau; P. Nozière; B. Graulet (pp. 777-790).

Simultaneous quantification of various liposoluble micronutrients is not a new area of interest since these compounds participate in the nutritional quality of feeds that is largely explored in human, and also in animal diet. However, the development of related methods is still under concern, especially when the carotenoid composition is complex such as in forages given to ruminants or in lipid-rich matrices like milk. In this paper, an original method for simultaneous extraction and quantification of all carotenoids, vitamins E, and A in milk was proposed. Moreover, a new UPLC method allowing simultaneous determination of carotenoids and vitamins A and E in forage, plasma and milk, and separation of 23 peaks of carotenoids in forages was described. This UPLC method using a HSS T3 column and a gradient solvent system was compared to a previously published reverse-phase HPLC using two C18 columns in series and an isocratic solvent system. The UPLC method gave similar concentrations of carotenoids and vitamins A and E than the HPLC method. Moreover, UPLC allowed a better resolution for xanthophylls, especially lutein and zeaxanthin, for the three isomers of β-carotene (all-E-, 9Z- and 13Z-) and for vitamins A, an equal or better sensitivity according to gradient, and a better reproducibility of peak areas and retention times, but did not reduce the time required for analysis. Figure Chromatogram of a mix of standard carotenoids obtained with UPLC at 450 nm

Keywords: Carotenoids; Tocopherol; Retinol; Liquid chromatography; Milk; Feed; Bioanalytical methods; Biological samples; Foods/Beverages; HPLC; UV/VIS

Preparation of a chitosan-coated C₁₈-functionalized magnetite nanoparticle sorbent for extraction of phthalate ester compounds from environmental water samples by Xiao Le Zhang; Hong Yun Niu; Sheng Xiao Zhang; Ya Qi Cai (pp. 791-798).

Novel superparamagnetic chitosan-coated C₁₈-functionalized magnetite nanoparticles (MNPs) were successfully synthesized and applied as an effective sorbent for the preconcentration of several typical phthalate ester compounds from environmental water samples. The MNPs were 20 nm in diameter and had a high magnetic saturation value (52 emu g⁻¹), which endowed the sorbent with a large surface area and the convenience of isolation from water samples. Phthalate esters could be extracted by the interior octadecyl groups through hydrophobic interaction. The hydrophilic porous chitosan polymer coating promoted the dispersion of MNPs in water samples, and improved the anti-interference ability of the sorbent without influencing the adsorption of analytes. The main factors affecting the adsorption of phthalate esters, including the pH of the solution, humic acid, sample loading volume, adsorption time, and desorption conditions, were investigated and optimized. Under the conditions selected (pH 11, adsorption time 20 min, elution with 10 mL of acetonitrile, and concentration to 0.5 mL), concentration factors of 1,000 were achieved by extracting 500 mL of several environmental water samples with 0.1 g of MNP sorbent. The method detection limits obtained for di-n-propyl phthalate, di-n-butyl phthalate, dicyclohexyl phthalate, and di-n-octyl phthalate were 12.3, 18.7, 36.4, and 15.6 ng L⁻¹, respectively. The recoveries of spiked samples ranged from 60 to 100%, with a low relative standard deviation (1–8%), which indicated good method precision. Figure

Keywords: Chitosan-coated C₁₈-functionalized magnetite nanoparticles; Solid-phase extraction; Phthalate esters

Determination of organophosphorous flame retardants in fish tissues by matrix solid-phase dispersion and gas chromatography by Luca Campone; Anna Lisa Piccinelli; Conny Östman; Luca Rastrelli (pp. 799-806).

Organophosphate esters (OPEs), utilized as flame retarding agents and/or plasticizers, are almost ubiquitous in environmental compartments, and biota and foods could be contaminated by bioaccumulation or during the treatment processes. A multiresidue method is proposed for the determination of 13 OPEs in fish tissues: analytes were simultaneously extracted and purified using the matrix solid phase dispersion technique and then determined by gas chromatography with nitrogen–phosphorus detection. The main parameters affecting extraction yield and selectivity, such as the type of dispersant material, clean-up co-sorbent, rinse and elution solvents, were evaluated to obtain lipid-free extracts and quantitative recoveries for OPEs. Under optimal conditions, 0.5 g of samples was dispersed with 2 g Florisil and 1 g anhydrous sodium sulphate and transferred to a solid phase extraction cartridge containing 1 g alumina. The lipids were removed using 5 mL n-hexane/dichloromethane (1:1) and analytes were recovered with 10 mL n-hexane/acetone (6:4) and directly analysed. The method developed provided recoveries between 70 and 110% for different kinds of fish, and the day-to-day variability was between 1 and 9%. This procedure constitutes the first analytical method for the analysis of OPEs in a food matrix and it is applicable to analyse a large number of samples to evaluate the occurrence and sources of OPEs in biota and foods. Figure GC-NPD chromatograms of the MSPD extracts from spiked salmon sample

Keywords: Organophosphates ester; Flame retardants; Matrix solid-phase dispersion; Fish tissues; Gas chromatography with nitrogen–phosphorus detection

Studies on the extraction of sulfonamides from agricultural soils by J. Raich-Montiu; J. L. Beltrán; M. D. Prat; M. Granados (pp. 807-814).

The extraction of six sulfonamides (sulfadiazine, sulfadimidine, sulfathiazole, sulfachloropiridazine, sulfadimethoxine, and sulfaquinoxaline) from soils with different physicochemical characteristics and at several aging times was investigated. Conventional mechanical shaking, microwave-assisted extraction, ultrasound probe-assisted extraction and pressurized liquid extraction techniques were evaluated. The four techniques provided similar results when applied to freshly contaminated soils. However, microwave-assisted extraction was the most suitable to extract sulfonamide aged residues from soils. Microwave-assisted extraction was applied to eight soils aged for 3 months, using acetonitrile:buffer pH 9 (20:80) as the extraction solvent, and recoveries ranged from 15–25% for STZ to 42–64% for SDM.

Keywords: Sulfonamides; Soil; Aged residues; Recovery; Microwave-assisted extraction

Molecularly imprinted polymer/cryogel composites for solid-phase extraction of bisphenol A from river water and wine by Claudio Baggiani; Patrizia Baravalle; Cristina Giovannoli; Laura Anfossi; Gianfranco Giraudi (pp. 815-822).

Superporous monolithic hydrogels (cryogel monoliths) are elastic, sponge-like materials that can be prepared in an aqueous medium through a cryotropic gelation technique. These monoliths show interesting properties for the development of high-throughput solid-phase extraction supports to treat large volumes of aqueous samples. In this work, a cryogel-supported molecularly imprinted solid-phase extraction approach for the endocrine disruptor bisphenol A (BPA) from river water and wine samples is presented. An imprinted polymer with molecular recognition properties for BPA was prepared in acetonitrile by thermal polymerization of a mixture of 4,4′-dihydroxy-2,2-diphenyl-1,1,1,3,3,3-trifluoropropane as a mimic template of BPA, 4-vinylpyridine and trimethylolpropane trimethacrylate in a molar ratio of 1 + 6 + 6. Fine imprinted particles (<10 μm) were embedded in a poly-acrylamide-co-N,N′-methylenbisacrylamide cryogel obtained by ammonium persulfate-induced cryopolymerization at −18 °C. The resulting monolithic gel was evaluated for its use as a sorbent support in an off-line solid-phase extraction approach to recover BPA from dilute aqueous samples with minimum pre-loading work-up. The optimized extraction protocol resulted in a reliable MISPE method suitable to selectively extract and preconcentrate BPA from river water and red wine samples, demonstrating the practical feasibility of cryogel-trapped imprinted polymers as solid-phase extraction materials Figure Superporous poly-acrylamide-co-N,N'-methylen-bis-acrylamide cryogels embedding micron-sized imprinted particles were used to set up a method for efficient solid phase extraction of the endocrine disruptor bisphenol A from river water and wine samples with minimum pre-loading work-up

Keywords: Bisphenol A; Molecularly imprinted polymer; Cryogel; Solid-phase extraction; Water analysis; Wine analysis

Evaluation of the novel passive sampler for cyanobacterial toxins microcystins under various conditions including field sampling by Jiří Kohoutek; Blahoslav Maršálek; Luděk Bláha (pp. 823-828).

In the present study, we have evaluated the effectiveness of a passive sampler for polar organic chemicals to accumulate a group of widespread and hazardous tumor-promoting toxins produced in cyanobacterial water blooms—microcystins (MC). The previously optimized configuration of the sampler based on polycarbonate membrane and Oasis HLB sorbent (2.75 mg/cm²) was validated under various exposure scenarios in laboratory and field. Calibration of the passive sampler conducted under variable conditions and concentrations of MC revealed linearity of the sampling up to 4 weeks. The sampling rates of microcystins for two different exposure scenarios were derived (e.g., MC-LR: R _s = 0.017 L/day under static and 0.087 L/d under turbulent conditions). R _s values were further used for calculations of time-weighted average concentrations in natural water. Improved sensitivity and selectivity of the in-house-made sampler was observed in comparison with the commercially available Polar Organic Compound Integrative Sampler (POCIS). Comparisons of grab and passive sampling methods were performed during cyanobacterial water bloom season in the Brno reservoir, Czech Republic in 2008. Data obtained by passive sampling provided a more relevant picture of the situation and enabled better assessment of potential risks. The present study demonstrated that the modification of POCIS is suitable for monitoring of occurrence and retrospective estimations of microcystin water concentrations, especially with respect to the control of drinking water quality.

Keywords: Passive sampling; POCIS; Microcystin; Cyanobacteria

Headspace solid-phase microextraction followed by gas chromatography tandem mass spectrometry for the sensitive determination of benzotriazole UV stabilizers in water samples by Inmaculada Carpinteiro; Brais Abuín; Isaac Rodríguez; Rafael Cela; Maria Ramil (pp. 829-839).

A sensitive procedure for the determination of five ultraviolet (UV) absorbers, belonging to the benzotriazole class, in environmental water samples is proposed. Analytes were first extracted and concentrated from the matrix and then selectively determined by gas chromatography in combination with tandem mass spectrometry detection. The high lipophilic character of some of the investigated species resulted in a strong trend to remain sorbed on solid surfaces, even after addition of considerable percentages of methanol (up to 30%) to water. Thus, minimizing sample handling during the enrichment step is mandatory in order to obtain acceptable accuracy and precision. Solid-phase microextraction (SPME), as sample preparation approach, fulfilled the above requirement and provided acceptable figures of merit for the determination of target species in environmental water samples, including raw wastewater. Optimization of SPME conditions showed that the combination of headspace extraction, with a sample temperature of 100 °C and addition of 15 mg of NaCl per milliliter of sample rendered the best compromise in terms of extraction efficiency for all species. Considering a sampling time of 30 min with a poly(dimethylsiloxane)–divinylbenzene-coated SPME fiber, limits of quantification below 2 ng l⁻¹ and relative standard deviations between 5% and 12% were achieved. Three of the five species included in this research were determined in raw wastewater with a maximum concentration of 57 ng l⁻¹ for the Tinuvin 326 UV absorber. Figure Proposed structures for product ions observed in the MS/MS spectra of Tinuvin 326

Keywords: UV stabilizers; Benzotriazoles; Water samples; Solid-phase microextraction; Gas chromatography; Mass spectrometry

Metal oxalates in paints: a Raman investigation on the relative reactivities of different pigments to oxalic acid solutions by A. Zoppi; C. Lofrumento; N. F. C. Mendes; E. M. Castellucci (pp. 841-849).

One degradation phenomenon that occurs in artworks is the formation of metal oxalates on their surfaces. In order to gain insight into the inclination of pigments to produce oxalates, nine pigments including Na, Ca, Fe, Pb and Cu cations were selected to react with oxalic acid solutions at different concentrations (1 M, 0.1 M, 0.01 M and 0.005 M). Micro-Raman spectroscopy was used to detect the different reaction products. Pigments containing calcium (calcite, gypsum and Volterra gypsum) showed a high tendency to form weddellite as well as whewellite, especially at high acidic concentrations; among copper-based pigments (malachite, azurite, verdigris), the formation of moolooite was observed for high concentrations of acid and down to the lowest concentration (0.005 M) in the case of verdigris. Lead oxalate was detected on lead white. No iron oxalates were observed for hematite; the formation of calcium oxalate crystals was observed instead. Ultramarine blue reacted to produce elemental sulfur. According to the results obtained, calcite and verdigris showed the highest reactivity in oxalic acid environments, resulting in a high tendency to form calcium and copper oxalates, even at very low acidic concentrations; this behavior seems to arise from the high solubilities of these pigments in acidic environments.

Keywords: Moolooite; Whewellite; Weddellite; Oxalates; Oxalic acid; Micro-Raman

Approach combining on-line metal exchange and tangential-flow ultrafiltration for in-situ characterization of metal species in humic hydrocolloids by Danielle Goveia; Fabiana Aparecida Lobo; Peter Burba; Leonardo Fernandes Fraceto; Newton Luiz Dias Filho; André Henrique Rosa (pp. 851-860).

This paper deals with the development and optimization of an analytical procedure using ultrafiltration and a flow-injection system, and its application in in-situ experiments to characterize the lability and availability of metal species in humic-rich hydrocolloids. The on-line system consists of a tangential flow ultrafiltration device equipped with a 3-kDa filtration membrane. The concentration of free ions in the filtrate was determined by atomic-absorption spectrometry, assuming that metals not complexed by aquatic humic substances (AHS) were separated from the complexed species (M–AHS) retained by the membrane. For optimization, exchange experiments using Cu(II) solutions and AHS solutions doped with the metal ions Ni(II), Mn(II), Fe(III), Cd(II), and Zn(II) were carried out to characterize the stability of the metal–AHS complexes. The new procedure was then applied in-situ at a tributary of the Ribeira do Iguape river (Iguape, São Paulo State, Brazil) and evaluated using the ions Fe(III) and Mn(II), which are considered to be essential constituents of aquatic systems. From the exchange between metal–natural organic matter (M–NOM) and the Cu(II) ions it was concluded that Cu(II) concentrations >485 μg L⁻¹ were necessary to obtain maximum exchange of the complexes Mn–NOM and Fe–NOM, corresponding to 100% Mn and 8% Fe. Moreover, the new analytical procedure is simple and opens up new perspectives for understanding the complexation, transport, stability, and lability of metal species in humic-rich aquatic environments.

Keywords: Lability/availability; Metal; Aquatic humic substances; Ultrafiltration

Direct determination of polymerized triglycerides in deep-frying olive oil by attenuated total reflectance–Fourier transform infrared spectroscopy using partial least squares regression by Julia Kuligowski; Guillermo Quintás; Salvador Garrigues; Miguel de la Guardia (pp. 861-869).

A partial least squares (PLS) regression model based on attenuated total reflectance–Fourier transform infrared spectra of heated olive oil samples has been developed for the determination of polymerized triacylglycerides (PTGs) generated during thermal treatment of oil. Three different approaches for selection of the spectral regions used to build the PLS model were tested and compared: (1) variable selection based on expert knowledge, (2) uninformative variable elimination PLS, and (3) interval PLS. Each of the three variable selection methods provided PLS models from heated olive oil samples with excellent performance for the prediction of PTGs in fried olive oils with comparable model statistics. However, besides a high coefficient of determination (R ² of 0.991) and low calibration, validation, and prediction errors of 1.14%, 1.21%, and 1.40% w/w, respectively, variable selection based on expert knowledge gave additionally almost identical low calibration (−0.0017% w/w) and prediction (−0.0023% w/w) bias. Furthermore, it was verified that the determination of PTGs was not influenced by the type of foodstuff fried in the olive oil.

Keywords: Polymerized triacylglycerides; Fried olive oil; Attenuated total reflectance–Fourier transform infrared spectroscopy (ATR-FTIR); Partial least squares (PLS)

Automated determination of uranium(VI) at ultra trace levels exploiting flow techniques and spectrophotometric detection using a liquid waveguide capillary cell by Jessica Avivar; Laura Ferrer; Montserrat Casas; Víctor Cerdà (pp. 871-878).

Rapid and fully automated multisyringe flow-injection analysis (MSFIA) with a multi-pumping flow system (MPFS) coupled to a long path-length liquid waveguide capillary cell (LWCC) is proposed for the determination of uranium(VI) at ultra trace levels. On-line separation and pre-concentration of uranium is carried out by means of a TRU resin. After elution, uranium(VI) is spectrophotometrically detected after reaction with arsenazo-III. Combination of the MSFIA and MPFS techniques with the TRU-resin enables the analysis to be performed in a short time, using large sample volumes and achieving high selectivity and sensitivity levels. A detection limit of 12.6 ng L⁻¹ (ppt) is reached for a 100-mL sample volume. The versatility of the proposed method also enables pre-concentration of variable sample volumes, enabling application of the analysis to a wide concentration range. Reproducibility of better than 5% and a resin durability of 40 injections should be emphasized. The developed method was successfully applied to different types of environmental sample matrices with recoveries between 95 and 108%.

Keywords: Uranium; Flow techniques; Spectrophotometric detection; TRU resin; Pre-concentration

Quantification of nitrotyrosine in nitrated proteins by Hong Yang; Yingyi Zhang; Ulrich Pöschl (pp. 879-886).

For kinetic studies of protein nitration reactions, we have developed a method for the quantification of nitrotyrosine residues in protein molecules by liquid chromatography coupled to a diode array detector of ultraviolet-visible absorption. Nitrated bovine serum albumin (BSA) and nitrated ovalbumin (OVA) were synthesized and used as standards for the determination of the protein nitration degree (ND), which is defined as the average number of nitrotyrosine residues divided by the total number of tyrosine residues in a protein molecule. The obtained calibration curves of the ratio of chromatographic peak areas of absorbance at 357 and at 280 nm vs. nitration degree are nearly the same for BSA and OVA (relative deviations <5%). They are near-linear at low ND (< 0.1) and can be described by a second-order polynomial fit up to $$ {hbox{ND}} = 0.5left( {{R^2} > 0.99} ight) $$ . A change of chromatographic column led to changes in absolute peak areas but not in the peak area ratios and related calibration functions, which confirms the robustness of the analytical method. First results of laboratory experiments confirm that the method is applicable for the investigation of the reaction kinetics of protein nitration. The main advantage over alternative methods is that nitration degrees can be efficiently determined without hydrolysis or digestion of the investigated protein molecules.

Keywords: Nitrotyrosine; HPLC–DAD; BSA; OVA; Nitration degree

Erratum to: A new ozone denuder for aerosol sampling based on an ionic liquid coating by Alexandre Albinet; Nicolas Papaiconomou; Julien Estager; Joël Suptil; Micheline Draye; Jean-Luc Besombes (pp. 887-887).

Sections

Personal tools

Analytical and Bioanalytical Chemistry (v.397, #2)

Sections

Personal tools

Local resources

Analytical and Bioanalytical Chemistry (v.397, #2)