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Analytical and Bioanalytical Chemistry (v.403, #6)

Basic mathematics and physics for undergraduate chemistry students according to the Eurobachelor® curriculum by Jens E. T. Andersen; D. Thorburn Burns; P. Hu (pp. 1461-1464).

is Associate Professor of Analytical Chemistry at DTU Chemistry in Copenhagen. He obtained his academic degrees at the University of Copenhagen and served his post doc at the University of Cambridge, UK before taking up his appointment at the Technical University of Denmark (DTU). His main scientific interest include studies of osteoporosis and development of active pharmaceutical ingredients (API’s) for treatment of osteoporosis and skin conditions, medical imaging, development and implementation of quality assurance for science, theory of chromatography and theory of electrochemistry. He served as President of the Danish Society of Analytical Chemistry and is National Delegate to the Division of Analytical Chemistry (DAC) of EuCheMS. He is the current Secretary to the EuCheMS DAC, where he is Head of the Study Group Quality Assurance. He participated in developing the Eurobachelor™ label introduced in 2006 by DAC Study Group Education headed by Reiner Salzer. MRIA FRSE notionally retired and became the Emeritus Professor of Analytical Chemistry in the Queen’s University of Belfast in 1999. However, he remains professionally and academically active as a Senior Visiting Fellow in the School of Chemistry and Chemical Engineering. He has on-going professional practice interests in the interpretation and validity of analytical data for legal/forensic purposes which ties in symbiotically with his current involvement in research and associated publication activities carried out at LGC in the programme carried out to support of the Government Chemist’s role as the official referee analyst. He is Head of the Study Group History in the Division of Analytical Chemistry of EuCheMS and has published 12 books and 420 papers. is a Professor at the School of Chemistry and Chemical Engineering, The Queen’s University of Belfast. He obtained a BSc in China before taking a Ph.D. at the University of Cambridge. In 1995, he joined the School of Chemistry at Queen’s University of Belfast as a lecturer, promoted to a Readership in 2001 and a professorship in 2004. In 2009, he was elected a Member of Royal Irish Academy. His research concerns physical and theoretical chemistry. His main interest is in chemical reactions and particularly catalytic reactions making contributions to the understanding of the fundamental principles of catalysis and the mechanisms of catalytic reactions.

Elias A.G. Zagatto, Claudio C. Oliveira, Alan Townshend and Paul J. Worsfold: Flow analysis with spectrophotometric and luminometric detection by Víctor Cerdà (pp. 1465-1466).

Integrated microring resonator sensor arrays for labs-on-chips by Carlos Angulo Barrios (pp. 1467-1475).

Planar waveguide optical ring resonators have shown great potential as compact and sensitive biochemical sensors. Advances in integrated optics based on Si technologies have allowed researchers to integrate multiple micron-sized ring resonator sensors with other optical and fluidic functions on Si chips using mass production techniques. Recent demonstrations of clinically relevant analyte detection by MRR sensor arrays have moved this technology closer to commercialization. Here, a survey of the development of microring sensor arrays for lab-on-a-chip applications is presented and illustrated with state-of-the-art examples.

Keywords: Optical ring resonator; Waveguide; Evanescent field; Integrated optics; Biochemical sensor; Multianalyte; Multiplex; Lab-on-a-chip

Nanostructured substrates for portable and miniature SPR biosensors by Julien Breault-Turcot; Jean-Francois Masson (pp. 1477-1484).

Surface plasmon resonance (SPR) biosensing has matured into a valuable analytical technique for measurements related to biomolecules, environmental contaminants, and the food industry. Contemporary SPR instruments are mainly suitable for laboratory-based measurements. However, several point-of-measurement applications would benefit from simple, small, portable and inexpensive sensors to assess the health condition of a patient, potential environmental contamination, or food safety issues. This Trend article explores nanostructured substrates for improving the sensitivity of classical SPR instruments and nanoparticle (NP)-based colorimetric substrates that may provide a solution to the development of point-of-measurement SPR techniques. Novel nanomaterials and methodology capable of enhancing the sensitivity of classical SPR sensors are destined to improve the limits of detection of miniature SPR instruments to the level required for most applications. In a different approach, paper or substrate-based SPR assays based on NPs, are a highly promising topic of research that may facilitate the widespread use of a novel class of miniature and portable SPR instruments. Figure Miniature plasmonic sensors based on nano structured substrates

Keywords: Kretschmann SPR; Au nanoparticles; Portable SPR; Grating substrates; Enhanced sensitivity; Paper-based SPR sensors

Combining a hydrogel and an electrochemical biosensor to determine the extent of degradation of paper artworks by Laura Micheli; Claudia Mazzuca; Antonio Palleschi; Giuseppe Palleschi (pp. 1485-1489).

Paper-based artworks are among the most valuable assets for transmission of knowledge. Historical paper is composed of different polysaccharides (e.g. cellulose), binders, and glues. During aging all of these components undergo several degradation processes, as a result of external and intrinsic causes, and these can compromise the state of conservation of the document. In this work, application of a new biotechnological strategy for paper artefact preservation is reported. By making use of innovative and non-invasive materials, for example appropriate hydrogels, in combination with selective electrochemical biosensors, it is possible to simultaneously verify the degradation condition of the paper artwork and then to efficiently clean it, while monitoring the process of removal of both pollution and degradation products. In this paper, we focus on specific examples in which such techniques have been applied to paper artworks and that illustrate the advantages and potential of this biotechnology compared with the traditional paper-cleaning methods currently in use. Figure Scheme of cleaning treatment of old paper and determination of the interested analyte using Flow Injection Analysis system (FIA) with integrated electrochemical biosensor

Keywords: Cultural heritage; Paper artworks; Rheoreversible gel; Electrochemical biosensor; Clean up; Degradation process

Recent advances in sulfotransferase enzyme activity assays by Priscilla Paul; Jiraporn Suwan; Jian Liu; Jonathan S. Dordick; Robert J. Linhardt (pp. 1491-1500).

Sulfotransferases are enzymes that catalyze the transfer of sulfo groups from a donor, for example 3′-phosphoadenosine 5′-phosphosulfate, to an acceptor, for example the amino or hydroxyl groups of a small molecule, xenobiotic, carbohydrate, or peptide. These enzymes are important targets in the design of novel therapeutics for treatment of a variety of diseases. This review examines assays used for this important class of enzyme, paying particular attention to sulfotransferases acting on carbohydrates and peptides and the major challenges associated with their analysis. Figure When a sulfotransferase transfers a sulfo group from PAPS to RXH (X= or NH) substrate the reaction can be assayed using radiometric, photometric, fluorometric or mass spectrometric methods

Keywords: Sulfotransferases; Sulfonation; Carbohydrates; Enzyme assays; Peptides

Hyphenated techniques as tools for speciation analysis of metal-based pharmaceuticals: developments and applications by Björn Meermann; Michael Sperling (pp. 1501-1522).

Method development and applications of hyphenated techniques as tools for speciation analysis of metal-based pharmaceuticals are summarized within this review. Advantages and limitations of the separation modes—high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), and gas chromatography (GC)—as well as the detection modes—inductively coupled plasma–mass spectrometry (ICP-MS) and electrospray ionization–mass spectrometry (ESI-MS)—are discussed. ICP-MS detection is found to be advantageous for the quantification of drugs containing metals and other heteroatoms. The species-independent sensitivity and multielement capabilities of ICP-MS allow it to be used for quantification even when species-specific standards are not available, as well as to determine the stoichiometry in metallodrug–biomolecule interactions. Molecular information that is totally destroyed when ICP is applied as ionization source and is therefore not obtainable via ICP-MS detection can be accessed by the complementary technique of ESI-MS. Speciation analysis combining both elemental and molecular information is therefore a powerful tool for the analysis of metal-based pharmaceuticals and their metabolites in body fluids and other relevant matrices. Figure Method development and applications of hyphenated techniques as tools for speciation analysis of metal-based pharmaceuticals are summarized within this review. Advantages and limitations of both the separation modules and the detection modules mainly such as inductively coupled plasma- (ICP-MS) or electrospray ionization-mass spectrometry (ESIMS) are discussed. ICP-MS detection is found to be advantageous for the quantification of drugs containing metals other heteroatoms while molecular information totally destroyed by the plasma and therefore not obtainable by means of ICP-MS detection is the domain of the complementary used ESI-MS.

Keywords: Speciation analysis; Hyphenated techniques; Metal-based pharmaceuticals; Metallodrugs; Complementary information; ICP-MS; ESI-MS

Current trends and challenges in sample preparation for global metabolomics using liquid chromatography–mass spectrometry by Dajana Vuckovic (pp. 1523-1548).

The choice of sample-preparation method is extremely important in metabolomic studies because it affects both the observed metabolite content and biological interpretation of the data. An ideal sample-preparation method for global metabolomics should (i) be as non-selective as possible to ensure adequate depth of metabolite coverage; (ii) be simple and fast to prevent metabolite loss and/or degradation during the preparation procedure and enable high-throughput; (iii) be reproducible; and (iv) incorporate a metabolism-quenching step to represent true metabolome composition at the time of sampling. Despite its importance, sample preparation is often an overlooked aspect of metabolomics, so the focus of this review is to explore the role, challenges, and trends in sample preparation specifically within the context of global metabolomics by liquid chromatography–mass spectrometry (LC–MS). This review will cover the most common methods including solvent precipitation and extraction, solid-phase extraction and ultrafiltration, and discuss how to improve analytical quality and metabolite coverage in metabolomic studies of biofluids, tissues, and mammalian cells. Recent developments in this field will also be critically examined, including in vivo methods, turbulent-flow chromatography, and dried blood spot sampling.

Keywords: Metabolomics; Untargeted metabolite profiling; Sample preparation; Metabolism quenching; Solvent extraction; Ultrafiltration; In vivo sampling; Dried blood spots; Method development; Liquid chromatography–mass spectrometry (LC–MS); Mammalian cells; Tissue

Current bioanalytical methods for detection of penicillins by Ruth Babington; Sonia Matas; M.-Pilar Marco; Roger Galve (pp. 1549-1566).

With the worldwide use of penicillin antibiotics comes the need for tighter controls. Bacterial resistance is a genuine problem and governmental and international bodies, for example the European Medicines Agency (EMA) and the World Health Organization (WHO), have designed strategies to overcome this unfortunate consequence of antibiotic use. Foodstuffs are monitored to ensure they contain very low quantities of antibiotics, so they are not prejudicial to health and the environment. Detection is based on chromatographic methods. However, screening can be performed by use of simpler, rapid methods of detection, e.g. microbial inhibition test, lateral flow assays, immunoassays, and use of biosensors, to reduce the final number of samples to be analyzed by chromatography. In this review, we have gathered information regarding all such screening methods for the penicillins and have critically assessed their capability and specificity for detection of penicillins.

Keywords: Penicillin; β-Lactam; PBP; Bioassay; Immunoassay; Biosensor

Multiplexed paper test strip for quantitative bacterial detection by S. M. Zakir Hossain; Cory Ozimok; Clémence Sicard; Sergio D. Aguirre; M. Monsur Ali; Yingfu Li; John D. Brennan (pp. 1567-1576).

Rapid, sensitive, on-site detection of bacteria without a need for sophisticated equipment or skilled personnel is extremely important in clinical settings and rapid response scenarios, as well as in resource-limited settings. Here, we report a novel approach for selective and ultra-sensitive multiplexed detection of Escherichia coli (non-pathogenic or pathogenic) using a lab-on-paper test strip (bioactive paper) based on intracellular enzyme (β-galactosidase (B-GAL) or β-glucuronidase (GUS)) activity. The test strip is composed of a paper support (0.5 × 8 cm), onto which either 5-bromo-4-chloro-3-indolyl-β-d-glucuronide sodium salt (XG), chlorophenol red β-galactopyranoside (CPRG) or both and FeCl₃ were entrapped using sol–gel-derived silica inks in different zones via an ink-jet printing technique. The sample was lysed and assayed via lateral flow through the FeCl₃ zone to the substrate area to initiate rapid enzyme hydrolysis of the substrate, causing a change from colorless-to-blue (XG hydrolyzed by GUS, indication of nonpathogenic E. coli) and/or yellow to red-magenta (CPRG hydrolyzed by B-GAL, indication of total coliforms). Using immunomagnetic nanoparticles for selective preconcentration, the limit of detection was ~5 colony-forming units (cfu) per milliliter for E. coli O157:H7 and ~20 cfu/mL for E. coli BL21, within 30 min without cell culturing. Thus, these paper test strips could be suitable for detection of viable total coliforms and pathogens in bathing water samples. Moreover, inclusion of a culturing step allows detection of less than 1 cfu in 100 mL within 8 h, making the paper tests strips relevant for detection of multiple pathogens and total coliform bacteria in beverage and food samples. Figure Pathogen Sensing Paper: Paper strips with ink-jet printed sensing zones can detect low levels of pathogenic or non-pathogenic bacteria. Incorporation of an immunomagnetic separation step results in selective detection of ~25 cfu of H7:O157 bacteria in under 1 h.

Keywords: Bacteria detection; Bioactive paper sensor; Colorimetric

Substrate-dependent kinetics in tyrosinase-based biosensing: amperometry vs. spectrophotometry by Liza Rassaei; Jin Cui; Edgar D. Goluch; Serge G. Lemay (pp. 1577-1584).

Despite the broad use of enzymes in electroanalytical biosensors, the influence of enzyme kinetics on the function of prototype sensors is often overlooked or neglected. In the present study, we employ amperometry as an alternative or complementary method to study the kinetics of tyrosinase, whose catalytic activity results in o-quinone products. We further compare our results for four monophenolic substrates with those obtained from ultraviolet–visible spectrophotometry and show that the results from both assays are in good agreement. We also observe large variations in the enzyme kinetics for different monophenolic substrates depending on the R-group at the para position. To further study this effect, we investigate the stability of quinone products in the enzymatic assay. This information can in principle be utilized to discriminate between different phenolic species by monitoring the reaction rate.

Keywords: Voltammetry; Biosensor; Amperometry; Electroanalysis; Tyrosinase; Phenols; Quinones stability; Diffusion; Michaelis–Menten kinetics; Turnover rate; Microelectrode

An electrochemical immunosensor for ochratoxin A determination in wines based on a monoclonal antibody and paramagnetic microbeads by Juan C. Vidal; Laura Bonel; Alba Ezquerra; Patricia Duato; Juan R. Castillo (pp. 1585-1593).

We report a direct competitive immunosensor for the rapid determination of ochratoxin A (OTA) in wine samples. Magnetic beads (1 ± 0.5 μm diameter) covered with streptavidin were functionalized with a monoclonal antibody against OTA, and then left to incubate in a solution of tracer (ochratoxin conjugated to the enzyme peroxidase) and a range of OTA concentrations (10⁻⁴ to 1,000 ng mL⁻¹). After washing and separation steps helped with a magnetic field, a volume of the dispersion was put on screen-printed electrodes under a magnet, and after adding the substrate the p-benzoquinone generated enzymatically was detected by differential-pulse voltammetry. Wine samples (2 mL) were easily prepared simply by adjusting to pH = 7.5 with diluted NaOH and by adding polyvinylpyrrolidone for complexing polyphenols, without any other clean-up or preconcentration steps. The limit of detection for detecting OTA in wines was of 0.11 ± 0.01 ng L⁻¹, well below the permitted content of the mycotoxin by the European Union (<2 ng mL⁻¹). Spiked wines were subjected to immunosensor calibrations to study the matrix effects. OTA concentrations measured with the immunosensor were compared with those obtained by high-performance liquid chromatography coupled to fluorescence detection (AOAC official method 2001.01). The OTA levels from two red wines of “Campo de Borja”, Spain, ranged from about 0.027 to 0.033 ng mL⁻¹ of OTA.

Keywords: Ochratoxin A; Electrochemical immunosensor; Wine samples; Magnetic beads

Factors influencing the detection limit of the lateral-flow sandwich immunoassay: a case study with potato virus X by Irina Safenkova; Anatoly Zherdev; Boris Dzantiev (pp. 1595-1605).

Key factors influencing the analyte detection limit of the sandwich immunochromatographic assay (ICA), namely, the size of gold nanoparticles, the antibody concentration, the conjugation pH, and characteristics of membranes, are discussed. The impacts of these factors were quantitatively characterized and compared for the first time using the same antigen (potato virus X). The antibody–colloidal gold conjugates synthesized at pH 9.0–9.5 (the pH was examined in the range from 7.5 to 10.0) and at an antibody concentration of 15 μg/mL (the concentration was tested from 10 to 100 μg/mL) demonstrated maximum binding with the analyte. The relationship between the size of gold nanoparticles and the ICA detection limit was determined. The detection limit decreases from 80 to 3 ng/mL (for antibodies with K _D = 1.0 × 10^-9 M, data were obtained using a BIAcore X instrument) for a series of particles with a diameter from 6.4 to 33.4 nm (electron microscopy and dynamic light scattering data). In the case of larger particles (52 nm in diameter), the detection limit increases and reaches 9 ng/mL. A 10 mM phosphate buffer, pH 8, and a 50 mM phosphate buffer, pH 7, were the conditions of choice for the deposition of reactants. Taking into account these facts, we developed a lateral-flow test system for the rapid (10 min) detection of potato virus X in plant leaves. The ICA provided a visual detection limit of 3 ng/mL. In the case of the instrumental processing, potato virus X can be determined in the concentration range from 3 to 300 ng/mL with a detection limit 2 ng/mL. Figure The detection limit of immunochromatographic test systems, corresponding average diameters of the gold nanoparticles in the conjugates and color intensities in the test zones

Keywords: Lateral-flow immunoassay; Gold nanoparticles; Antibody–colloidal gold conjugate; Potato virus X

Development of sensitive direct and indirect enzyme-linked immunosorbent assays (ELISAs) for monitoring bisphenol-A in canned foods and beverages by Yang Lu; Joshua Richard Peterson; John Justin Gooding; Nanju Alice Lee (pp. 1607-1618).

Enzyme-linked immunosorbent assays (ELISAs) are investigated in this work for the detection of bisphenol-A (BPA), a plastic monomer and a critical contaminant in food and environment. A series of polyclonal antibodies generated in vivo using BPA-butyrate-protein conjugate and BPA-valerate-protein conjugate were evaluated on direct and indirect competitive assay formats with five competing haptens (BPA-butyrate, BPA-valerate, BPA-crotonate, BPA-acetate, and BPA-2-valerate). Two indirect ELISAs and one direct ELISA exhibiting high sensitivity and specificity for BPA were developed. The 50 % inhibition of antibody binding (IC₅₀) values were 0.78 ± 0.01–1.20 ± 0.26 μg L⁻¹, and the limits of detection as measured by the IC₂₀ values were 0.10 ± 0.03–0.20 ± 0.04 μg L⁻¹. The assays were highly specific to BPA, only displaying low cross-reactivity (3–8 % for the indirect assays and 26 % for the direct assay) for 4-cumylphenol (4-CP), at pH 7.2. The degree of cross-reaction of 4-CP was influenced by the antibody/hapten conjugate combination, assay conditions, and the assay format. The assays were optimized for the analysis of BPA in canned vegetables, bottled water and carbonated drinks. The limits of quantification for these three evaluated sample types, based on the spike and recovery data, were 0.5, 2.5, and 100 μg L⁻¹, respectively.

Keywords: Bisphenol-A; Enzyme-linked immunosorbent assays; Canned corn; Bottled water; Carbonated drinks

Detection of recombinant human EPO administered to horses using MAIIA lateral flow isoform test by Maria Lönnberg; Ulf Bondesson; Florence Cormant; Patrice Garcia; Yves Bonnaire; Jan Carlsson; Marie-Agnes Popot; Niclas Rollborn; Kristina Råsbo; Ludovic Bailly-Chouriberry (pp. 1619-1628).

Doping of horses with recombinant human erythropoietin (rHuEPO) to illegally enhance their endurance capacity in horseracing has been reported during the last years. This leads to increased blood viscosity which can result in sudden death and is of concern for the horse welfare. Additionally, the horse can start production of rHuEPO antibodies, which cross-reacts with endogenous equine EPO and can lead to severe anaemia and even death. In this study, a novel micro-chromatographic method, EPO WGA MAIIA, has been tested for the capability in plasma and urine samples to detect administration of erythropoiesis-stimulating agents, like the rHuEPO glycoprotein varieties Eprex and Aranesp, to horses. After administration of 40 IU Eprex kg⁻¹ day⁻¹ to seven horses during 6 days, the presence of Eprex in horse plasma was detected up to 2–5 days after last injection. In urine samples collected from two horses, Eprex was detected up to 3 days. A single injection of Aranesp (0.39 μg/kg) was detected up to 9 days in plasma and up to 8 days, the last day of testing, in the urine sample. The LC-FAIMS-MS/MS system, with 1 day reporting time, confirmed the presence of Eprex up to 1 day after last injection for six out of seven horses and the presence of Aranesp up to 5 days after last injection in plasma samples. The MAIIA system showed to be a promising tool with high sensitivity and extremely short reporting time (1 h). Fig The interior of the MAIIA membrane. EPO protein molecules (blue balls) flow through the WGA lectin ligands immobilised in the network structure. Some types of EPO have oligosaccharide structures with higher interaction strength and will be delayed, while other low reacting types will rapidly reach the anti-EPO antibodies immobilised downstream, during the 5 min. reaction time.

Keywords: Aranesp; EPO doping control; Eprex; Equine EPO; Micro-chromatography; WGA

Quantification of steroids and endocrine disrupting chemicals in rat ovaries by LC-MS/MS for reproductive toxicology assessment by Nadia Quignot; Mikaël Tournier; Charlène Pouech; Cécile Cren-Olivé; Robert Barouki; Emmanuel Lemazurier (pp. 1629-1640).

Reproductive function is controlled by a finely tuned balance of androgens and estrogens. Environmental toxicants, notably endocrine disrupting chemicals (EDCs), appear to be involved in the disruption of hormonal balance in several studies. To further describe the effects of selected EDCs on steroid secretion in female rats, we aim to simultaneously investigate the EDC concentration and the sex hormone balance in the ovaries. Therefore, an effective method has been developed for the quantification of the sex steroid hormones (testosterone, androstenedione, estradiol, and estrone) and four endocrine disrupting chemicals (bisphenol A, atrazine, and the active metabolites of methoxychlor and vinclozolin) in rat ovaries. The sample preparation procedure is based on the so-called “quick, easy, cheap, effective, rugged, and safe” approach, and an analytical method was developed to quantify these compounds with low detection limits by liquid chromatography coupled with a tandem mass spectrometer. This analytical method, applied to rat ovary samples following subacute EDC exposure, revealed some new findings for toxicological evaluation. In particular, we showed that EDCs with the same described in vitro mechanisms of action have different effects on the gonadal steroid balance. These results highlight the need to develop an integrative evaluation with the simultaneous measurement of EDCs and numerous steroids for good risk assessment. Fig Ovaries from rats treated or not with model endocrine disrupting chemicals were subjected to LC-MS/MS analysis for the simultaneous quantification of chemicals and sex hormones.

Keywords: Endocrine disrupting chemicals; Hormonal balance; LC-MS/MS; Steroids; Toxicity

Elemental imaging of MRI contrast agents: benchmarking of LA-ICP-MS to MRI by J. A. T. Pugh; A. G. Cox; C. W. McLeod; J. Bunch; M. J. Writer; S. L. Hart; A. Bienemann; E. White; J. Bell (pp. 1641-1649).

Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has been used to map the spatial distribution of magnetic resonance imaging (MRI) contrast agents (Gd-based) in histological sections in order to explore synergies with in vivo MRI. Images from respective techniques are presented for two separate studies namely (1) convection enhanced delivery of a Gd nanocomplex (developmental therapeutic) into rat brain and (2) convection enhanced delivery, with co-infusion of Magnevist (commercial Gd contrast agent) and Carboplatin (chemotherapy drug), into pig brain. The LA technique was shown to be a powerful compliment to MRI not only in offering improved sensitivity, spatial resolution and signal quantitation but also in giving added value regarding the fate of administered agents (Gd and Pt agents). Furthermore simultaneous measurement of Fe enabled assignment of an anomalous contrast enhancement region in rat brain to haemorrhage at the infusion site. Figure Gd contrast-enhanced MRI image (left) and LA-ICP-MS ¹⁵⁷Gd elemental distribution (right) for pig brain hemisphere dosed via convection enhanced delivery.

Keywords: MRI, laser ablation; ICP-MS; Elemental imaging; Fe, Gd, Pt; Biological thin sections

Metabolomic study of plasma of patients with abdominal aortic aneurysm by Francisco J. Rupérez; Priscila Ramos-Mozo; Joanna Teul; Roxana Martinez-Pinna; Antonia Garcia; Myriam Malet-Martino; Emilio Camafeita; Juan Antonio Lopez; Carlos Pastor-Vargas; Jesús Egido; Stéphane Balayssac; Véronique Gilard; Coral Barbas; Jose Luis Martin-Ventura (pp. 1651-1660).

Abdominal aortic aneurysm (AAA) is an important health problem, both because of AAA rupture and death and because of increased cardiovascular mortality. Identification of new biomarkers of AAA may suggest novel pathological mechanisms and targets for new medical treatments to slow AAA progression. Metabolic changes in AAA patients were mainly related to carbohydrate and lipid metabolism and many of these changes can be associated with a situation of insulin resistance (which can be related to metabolic syndrome) together with altered amino acid metabolism. For the first time, metabolites that can be associated with differential metabolism by the gut microflora of AAA patients have also been found. Moreover, aminomalonic acid in plasma has been shown to be the metabolite with the biggest difference between patients suffering from large aneurysm (>5 cm) and controls. Figure In the quest for adequate biomarkers, suitable for diagnosis and/or monitoring the presence and size of abdominal aortic aneurysm, metabolomics opens new possibilities thanks to the non-targetted approach. That permits to evaluate differences without previous biasing hypotheses. By coupling signals from 1H NMR and GC-MS, it has been possible to find metabolites, quantitatively different in aneurysm patients when compared with controls.

Keywords: Metabonomics; Aortic abdominal aneurysm; Metabolic pathways

Cell surface glycoprotein profiling of cancer cells based on bioorthogonal chemistry by Peng-wei Pan; Qi Zhang; Jie Hou; Ze Liu; Fang Bai; Mei-rong Cao; Ting Sun; Gang Bai (pp. 1661-1670).

Bioorthogonal chemistry refers to chemical reactions that can occur within a living system without altering native biochemical processes. Applications of this concept extend to studies on a group of biomolecules that includes glycans, proteins, and lipids. In this study, a strategy for isolating cell surface glycoproteins and based on bioorthogonal chemistry was employed to identify new cancer-related glycoproteins. A novel alkyne reagent containing one disulfide bond was synthesized for the enrichment of glycoproteins metabolized with peracetylated N-azidoacetylmannosamine, which was applied on three different cancer cell lines, and all isolated proteins were analyzed by high-performance liquid chromatography–tandem mass spectrometry. The strategy of purifying cell surface glycoproteins introduced in this article was shown to be reliable, and a total of 56 cell surface glycoproteins were identified. Neuronal cell adhesion molecule was found uniquely expressed in A549 lung adenocarcinoma, and its expression in non-small-cell lung carcinomas was detected by immunohistochemistry. Furthermore, a significant increase of neuronal cell adhesion molecule expression was identified in non-small-cell lung adenocarcinoma compared with adjacent noncancerous tissues, and could be a novel potential target and marker in cancer treatment and detection.

Keywords: Click chemistry; Sialic acid; Glycoprotein; Lung adenocarcinoma; Neuronal cell adhesion molecule

Analysis of urinary oligosaccharides in lysosomal storage disorders by capillary high-performance anion-exchange chromatography–mass spectrometry by Cees Bruggink; Ben J. H. M. Poorthuis; André M. Deelder; Manfred Wuhrer (pp. 1671-1683).

Many lysosomal storage diseases are characterized by an increased urinary excretion of glycoconjugates and oligosaccharides that are characteristic for the underlying enzymatic defect. Here, we have used capillary high-performance anion-exchange chromatography (HPAEC) hyphenated to mass spectrometry to analyze free oligosaccharides from urine samples of patients suffering from the lysosomal storage disorders fucosidosis, α-mannosidosis, G_M1-gangliosidosis, G_M2-gangliosidosis, and sialidosis. Glycan fingerprints were registered, and the patterns of accumulated oligosaccharides were found to reflect the specific blockages of the catabolic pathway. Our analytical approach allowed structural analysis of the excreted oligosaccharides and revealed several previously unpublished oligosaccharides. In conclusion, using online coupling of HPAEC with mass spectrometric detection, our study provides characteristic urinary oligosaccharide fingerprints with diagnostic potential for lysosomal storage disorders.

Keywords: HPAEC-IPAD; Catabolism; Metabolic disorder; Clinical glycomics; N-linked glycans; Glycolipids

An automated method for the measurement of a range of tyrosine kinase inhibitors in human plasma or serum using turbulent flow liquid chromatography–tandem mass spectrometry by L. Couchman; M. Birch; R. Ireland; A. Corrigan; S. Wickramasinghe; D. Josephs; J. Spicer; R. J. Flanagan (pp. 1685-1695).

Tyrosine kinase inhibitors (TKIs) are used to treat a number of cancers, including chronic myeloid leukaemia and hepatocellular carcinoma. Therapeutic drug monitoring (TDM) may be indicated to (1) monitor adherence, (2) guide dosage, and (3) minimise the risk of drug–drug interactions and dose-related toxicity. On-line, automated sample preparation provided by TurboFlow technology (ThermoFisher Scientific) in conjunction with the sensitivity and selectivity of tandem mass spectrometry (MS/MS) detection may be applied to the analysis of single drugs and metabolites. We report the use of TurboFlow LC–MS/MS for the analysis of nine TKIs and metabolites (imatinib, N-desmethylimatinib, dasatinib, nilotinib, erlotinib, gefitinib, lapatinib, sorafenib, sunitinib) in human plasma or serum for TDM purposes. An Aria Transcend TLX-II system coupled with a TSQ Vantage was used. Samples (50 μL) were vortex mixed with internal standard solution (150 μL imatinib-D₈, gefitinib-D₈, sunitinib-D₁₀, and nilotinib-¹³C ₂ ¹⁵ N₂ in acetonitrile) and, after centrifugation 100 μL supernatant were injected directly onto a 50 × 0.5-mm Cyclone TurboFlow column. Analytes were focussed onto a 50 × 2.1-mm (3 μm) Hypersil GOLD analytical column and eluted with an acetonitrile/water gradient. Analytes were monitored in selected reaction monitoring mode (positive APCI). Total analysis time was 7 min without multiplexing. Calibration was linear (R ² > 0.99) for all analytes. Inter- and intra-assay precision (in percent relative standard deviation, RSD) was <11 % and accuracy 89–117 % for all analytes. No matrix effects were observed. This method is suitable for high-throughput TDM in patients undergoing chronic therapy with TKIs and has been utilised in the analysis of clinical samples.

Keywords: TurboFlow MS/MS; TKIs; Imatinib; Dasatinib; Nilotinib; TDM; Automated sample preparation

Selective detection of alkaloids in MALDI-TOF: the introduction of a novel matrix molecule by Andreas Schinkovitz; Ghislain Tsague Kenfack; Denis Seraphin; Eric Levillain; Maryléne Dias; Pascal Richomme (pp. 1697-1705).

The current manuscript presents 3-[5′-(methylthio)-2,2′-bithiophen-5-ylthio]propanenitrile (MT3P), as a novel matrix molecule, which facilitates the selective ionization of alkaloids in matrix-assisted laser desorption/ionization mass spectrometry. Exhibiting strong ionizing properties at low levels of laser energy, MT3P was evaluated on 55 compounds belonging to various chemical families. The observed molecular ion yields induced by MT3P were compared with those obtained by commercially available matrices such as 1,8-dihydroxy-9,10-dihydroanthracen-9-one, α-cyano-4-hydroxycinnamic acid, 2,2′:5′,2″-terthiophene and 2,5-dihydroxybenzoic acid. In conclusion, MT3P displayed excellent ionization properties for 23 out of 25 investigated alkaloids, while showing little to no interaction with compounds from different chemical origin. Further, in comparison to other tested matrices, MT3P generally facilitated better ionization of alkaloids. Eventually, levels of laser energy were adjusted to obtain spectra with significantly reduced matrix noise.

Keywords: Mass spectrometry; MALDI; Aklaloids; Selective detection

Reduction of matrix effects and improvement of sensitivity during determination of two chloridazon degradation products in aqueous matrices by using UPLC-ESI-MS/MS by Sebastian Kowal; Peter Balsaa; Friedrich Werres; Torsten C. Schmidt (pp. 1707-1717).

The development and validation of a sensitive and reliable detection method for the determination of two polar degradation products, desphenyl-chloridazon (DPC) and methyl-desphenyl-chloridazon (MDPC) in surface water, ground water and drinking water is presented. The method is based on direct large volume injection ultra-performance liquid chromatography electrospray tandem mass spectrometry. This simple but powerful analytical method for polar substances in the aquatic environment is usually hampered by varying matrix effects, depending on the nature of different water bodies. For the two examined degradation products, the matrix effects are particularly strong compared with other polar degradation products of pesticides. Therefore, matrix effects were studied thoroughly with the aim of minimising them and improving sensitivity during determination by postcolumn addition of ammonia solution as a modifier. An internal standard was used in order to compensate for remaining matrix effects. The calibration curve shows very good coefficients of correlation (0.9994 for DPC and 0.9999 for MDPC). Intraday precision values were lower than 5 % for DPC, 3 % for MDPC and the limits of detection were 10 ng/L for both substances. The method was successfully used in a national round robin test with a deviation between 3 and 8 % from target values. Finally, about 1,000 samples from different water bodies have been examined with this method in the Rhine and Ruhr region of North-Rhine-Westphalia (Germany) and in the European Union. Approximately 76 % of analysed samples contained measurable amounts of DPC at concentrations up to 8 μg/L while 53 % of the samples showed MDPC concentrations up to 2.3 μg/L

Keywords: Pesticides; Polar degradation products; LC-ESI-MS/MS; Matrix effects; Ammonia as modifier; Postcolumn infusion; On-column focusing; Internal standard

Selective LC-MS/MS method for the identification of BMAA from its isomers in biological samples by Liying Jiang; Benoit Aigret; Wim M. De Borggraeve; Zdenek Spacil; Leopold L. Ilag (pp. 1719-1730).

Algal blooms are well-known sources of acute toxic agents that can be lethal to aquatic organisms. However, one such toxin, β-N-methylamino-l-alanine (BMAA) is also believed to cause amyotrophic lateral sclerosis, also known as Lou Gehrig’s disease. The detection and identification of BMAA in natural samples were challenging until the recent introduction of reliable methods. However, the issue of potential interference from unknown isomers of BMAA present in samples has not yet been thoroughly investigated. Based on a systematic database search, we generated a list of all theoretical BMAA structural isomers, which was subsequently narrowed down to seven possible interfering compounds for further consideration. The seven possible candidates satisfied the requirements of chemical stability and also shared important structural domains with BMAA. Two of the candidates, 2,4-diaminobutyric acid (DAB) and N-(2-aminoethyl) glycine (AEG) have recently been studied in the context of BMAA. A further isomer, β-amino-N-methyl-alanine (BAMA), has to be considered because it can potentially yield the fragment ion, which is diagnostic for BMAA. Here, we report the synthesis and analysis of BAMA, together with AEG, DAB, and other isomers that are of interest in the separation and detection of BMAA in biological samples by using either high-performance liquid chromatography or ultra-high-performance liquid chromatography coupled with tandem mass spectrometry. We detected for the first time BAMA in blue mussel and oyster samples. This work extends the previously developed liquid chromatography–tandem mass spectrometry platform Spacil et al. (Analyst 135:127, 2010) to allow BMAA isomers to be distinguished, improving the detection and identification of this important amino acid.

Keywords: ALS; Cyanobacteria; β-amino-N-methylalanine (BAMA); AEG; DAB

Determination of pesticide residues in wine by membrane-assisted solvent extraction and high-performance liquid chromatography–tandem mass spectrometry by M. Moeder; C. Bauer; P. Popp; M. van Pinxteren; T. Reemtsma (pp. 1731-1741).

The determination of pesticides in food products is an essential issue to guarantee food safety and minimise health risks of consumers. A protocol based on membrane-assisted solvent extraction and liquid chromatography–tandem mass spectrometry (HPLC-MS/MS) that allows the determination of 18 pesticides in red wine at minimum labour effort for sample preparation was developed and validated. Ten millilitres of wine were extracted using 100 μL of toluene filled in a non-porous polyethylene membrane bag which is immersed in the wine sample. After 150 min extraction under stirring, an aliquot of the extraction solution is analysed using HPLC-MS/MS. The limits of quantification ranged from 3 ng/L for Pirimicarb to 1.33 μg/L for Imidacloprid. Quantification by matrix-matched calibration provided relative standard deviations ≤16 % for most of the target pesticides. The linearity of calibration was given over three to four orders of magnitude, which enables the reliable measurement of a broad range of pesticide concentrations, and for each target pesticide, the sensitivity of the protocol meets the maximum residue levels set by legislations at least for wine grapes. Good agreement of results was found when the new method was compared with a standard liquid–liquid extraction protocol. In five wine samples analysed, Carbendazim and Metalaxyl were determined at micrograms per litre concentrations, even in some of the organic wines. Tebuconazol and Cyprodinitril were determined at lower abundance and concentration, followed by Spiroxamin and Diuron.

Keywords: Membrane-assisted solvent extraction; HPLC-MS/MS; Pesticides; Wine

Comparative study of hollow-fiber liquid-phase micro-extraction and an aqueous two-phase system for determination of phytohormones in soil by She Ying Dong; Zhen Yang; Penghui H. Zhang; Qing Hu; Ting Lin Huang (pp. 1743-1749).

Two methods, hollow-fiber liquid-phase micro-extraction (HF-LPME) and an aqueous two-phase system (ATPS), have been systematically optimized and compared for extraction and determination of phytohormones in soil by high-performance liquid chromatography (HPLC). The effects on extraction of conditions including solvent type and volume, extraction time, temperature, and amount of salt were evaluated. It was shown that ATPS was superior to HF-LPME for determination of paclobutrazol and uniconazole under the optimum conditions. The limits of detection (LODs) of ATPS were 0.002 μg g⁻¹ for uniconazole and 0.01 μg g⁻¹ for paclobutrazol, whereas LODs of HF-LPME were 0.005 μg g⁻¹ and 0.03 μg g⁻¹, respectively. Relative standard deviations (RSDs, n = 5) and recovery were in the range 1.7–5.3 % and 86–102 %, respectively, for ATPS and 6.7–7.9 % and 40–60 % for HF-LPME. In addition, the advantages of ATPS were shorter extraction time, suitable for simultaneous pretreatment of batches of samples, and higher extraction capacity. ATPS was therefore applied to the determination of paclobutrazol and uniconazole in real soil samples. Uniconazole was detected in all the samples analyzed whereas paclobutrazol was not found.

Keywords: Ionic liquid; hollow-fiber liquid-phase micro-extraction; aqueous two-phase systems; phytohormones; soil samples

Enantiomeric determination of azole antifungals in wastewater and sludge by liquid chromatography–tandem mass spectrometry by Qiuxin Huang; Kun Zhang; Zhifang Wang; Chunwei Wang; Xianzhi Peng (pp. 1751-1760).

A sensitive and reliable liquid chromatographic–tandem mass spectrometric method for enantiomeric determination of five chiral azole antifungals (econazole, ketoconazole, miconazole, tebuconazole, and propiconazole) in wastewater and sludge has been established and validated. An isotope-labeled internal standard was used for quantification. Recovery of the individual enantiomers was usually in the range of 77–102 % for wastewater and 71–95 % for sludge, with relative standard deviations within 20 %. No significant difference (p > 0.05) was observed between recovery of pairs of enantiomers of the chiral azole antifungals except for those of tebuconazole. Method quantification limits for individual enantiomers were 0.3–10 ng L⁻¹ and 3–29 ng g⁻¹ dry weight for wastewater and sludge, respectively. The method was used to investigate the enantiomeric composition of the azole pharmaceuticals in wastewater and sludge samples from a sewage treatment plant in China. Enantiomers of miconazole, ketoconazole, and econazole were widely detected. The results showed that the azole antifungals in wastewater and sludge were generally racemic or marginally non-racemic. The method is a useful tool for investigation of the enantiomeric occurrence, behavior, and fate of the chiral azole antifungals in the environment. FIGURE MRM chromatograms (black) and simulated chromatograms by PeakFit (red) for the chiral azole antifungals in standard solution and samples. MQL, method quantification limit

Keywords: Enantiomeric determination; Chiral azole antifungals; Liquid chromatography–tandem mass spectrometry; Wastewater; Sludge

Microwave-assisted headspace solid-phase microextraction to quantify polycyclic aromatic hydrocarbons in pine trees by Nuno Ratola; Paulo Herbert; Arminda Alves (pp. 1761-1769).

A methodology for the extraction and quantification of 16 polycyclic aromatic hydrocarbons (PAHs) based on microwave-assisted extraction coupled with headspace solid-phase microextraction followed by gas chromatography/mass spectroscopy was validated for needles and bark of two pine species (Pinus pinaster Ait. and Pinus pinea L.). The limits of detection were below 0.92 ng g⁻¹ (dry weight) for needles and below 0.43 ng g⁻¹ (dw) for bark. Recovery assays were performed with two sample masses spiked at three levels and the overall mean values were between 70 and 110 % for P. pinaster and 75 and 129 % for P. pinea. In the first species, the increase in sample mass lowered the recoveries slightly for most PAHs, whereas for the second, the recoveries were higher for the needles. Naturally contaminated samples from 4 sites were analysed, with higher levels for urban sites (1,320 and 942 ng g⁻¹ (dw) vs. 272 and 111 ng g⁻¹ (dw) for needles and 696 and 488 ng g⁻¹ (dw) vs. 270 and 103 ng g⁻¹ (dw) for bark) than for rural ones and also for P. pinaster samples over P. pinea. It is also shown that gas-phase PAHs are predominant in the needles (over 65 % of the total PAHs) and that the incidence for particulate material in bark, reaching 40 % as opposed to a maximum below 20 % for the needles. The method has proved to be fit and improved some of the existing approaches, on the assessment of particulate PAHs and bark levels.

Keywords: Solid-phase microextraction; Microwave-assisted extraction; Polycyclic aromatic hydrocarbons; Pine needles; Pine bark; GC-MS

Erratum to: A novel planar optical sensor for simultaneous monitoring of oxygen, carbon dioxide, pH and temperature by Sergey M. Borisov; Roman Seifner; Ingo Klimant (pp. 1771-1771).

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Analytical and Bioanalytical Chemistry (v.403, #6)

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Analytical and Bioanalytical Chemistry (v.403, #6)