Analytical and Bioanalytical Chemistry (v.404, #2)
Progress on environmental analysis and bioanalysis in Spain by Elena Domínguez; Alfredo Sanz-Medel (287-288).
has been Professor of Analytical Chemistry in the Department of Analytical Chemistry at the University of Alcala in Alcalá de Henares (Spain) since 2002. Her professional activity has been conducted at the Chemical Center at Lund University (Sweden), in the European Commission (Directorate-General for Research and Innovation, Brussels, Belgium), and mainly at the University of Alcalá. There, she leads a group specializing in bioelectrochemistry for the production of (bio)sensing systems for diagnostics, food control, and environmental monitoring. Her other research interests include data mining for sample identification and, more recently, biomimetic nanoparticles. She has published 80 scientific articles in peer-reviewed journals. Presently, she is President of the Spanish Society for Analytical Chemistry (SEQA). has been Professor in the Department of Physical and Analytical Chemistry at Oviedo University (Spain) since 1982. He is the author or coauthor of approximately 500 scientific publications in international journals, several patents, and books. His current research interests include new atomic detectors and ion sources for ultratrace analysis using plasmas, new molecular optical sensors, particularly those based on the use of quantum dots, and hybrid techniques, coupling a separation unit and an atomic detector, for ultratrace and trace metal speciation to solve biological and environmental problems and speciation for proteomics. This is aimed at by integrating molecular (matrix-assisted laser desorption/ionization and electrospray mass spectrometry) and atomic (inductively coupled plasma mass spectrometry) mass spectrometry techniques, and the extensive use of inductively coupled plasma mass spectrometry techniques to perform heteroatom-tagged proteomics, both for qualitative and for quantitative purposes. He has been an editor of Analytical and Bioanalytical Chemistry since 2002. At Euroanalysis 2007, in Antwerp, he received the Robert Kellner Award. In 2011, he received the European Award for Plasma Spectrochemistry.
Focused ultrasound solid–liquid extraction and gas chromatography tandem mass spectrometry determination of brominated flame retardants in indoor dust by María Pilar Martínez-Moral; María Teresa Tena (289-295).
This study presents a focused ultrasound solid–liquid extraction (FUSLE) and gas chromatography tandem mass spectrometry method for the determination of brominated diphenyl ethers (BDEs), from mono- to hexa-congeners, in indoor dust. This approach provided a simple, fast, and economical method. After the solvent extraction selection, the FUSLE conditions were studied using a central composite design. Finally, the number of extraction cycles was studied. The selected conditions were 8 mL of 3:1 n-hexane-acetone mixture as extraction solvent, at a power of 65% for 20 s. The proposed method allowed accurate determination of BDEs, with recovery values around 100% and detection limits between 0.05 and 0.8 ng g−1. It also has advantages over other existing methods in terms of simplicity, analysis time, and solvent consumption. The analysis of several indoor dust samples showed high concentration values of BDEs 47, 99, 100, 153, and 154 in some of the samples, moreover, BDEs 47 and 99 were found in all samples.
Keywords: Endocrine disruptors; Flame retardants; BDE; Dust; FUSLE; GC/MS-MS
Determination of 5-nitroimidazoles and metabolites in environmental samples by micellar electrokinetic chromatography by Maykel Hernández-Mesa; Carmen Cruces-Blanco; Ana M. García-Campaña (297-305).
A method based on micellar electrokinetic chromatography (MEKC) with UV detection has been developed for the determination of nine 5-nitroimidazoles (5-NDZs), including metabolites in river water samples. Due to the relative insensitivity of UV detection in MEKC, a solid-phase extraction (SPE) method has been proposed that preconcentrates water samples fiftyfold and cleans them up off-line. An on-line preconcentration approach based on sweeping and the use of an extended light path fused-silica capillary (64.5 cm × 50 μm i.d., 56 cm effective length) was also found to improve the sensitivity of the method. Separation was carried out in <21 min using 20 mM phosphate buffer (pH 6.5) and 150 mM SDS as the background electrolyte (BGE). The temperature of the capillary was kept constant at 20°C, a voltage of 25 kV was applied (normal mode), and a detected wavelength of 320 nm was utilized. Hydrodynamic injection (50 mbar for 15 s) of the samples, which were dissolved in 20 mM phosphate (pH 6.5), was employed. The limits of detection were lower than 1.1 μg L−1. Recoveries of >80% from spiked river water samples were obtained for most of the analytes at three different concentration levels with acceptable precision. This method could provide an efficient and economical alternative to the use of chromatographic methods to monitor nitroimidazole residues, thus supplementing the relatively few methods available for the analysis of these compounds in environmental samples.
Keywords: Micellar electrokinetic chromatography; Nitroimidazoles; SPE; River water samples
Non-aqueous capillary electrophoresis separation of fullerenes and C60 fullerene derivatives by Alina Astefanei; Oscar Núñez; M. Teresa Galceran (307-313).
As the interest in the use of fullerene compounds in biomedical and cosmetic applications increases, so too does the need to develop methods for their determination and quantitation in such complex matrices. In this work, we studied the behavior of C60 and C70 fullerenes in non-aqueous capillary electrophoresis, as well as two C60 fullerene derivatives not previously reported by any electrophoretic method, N-methyl-fulleropyrrolidine and (1,2-methanofullerene C60)-61-carboxylic acid. The separation was performed using fused-silica capillaries with an I.D. of 50 μm and tetraalkylammonium salts, namely tetra-n-decylammonium bromide (200 mM) and tetraethylammonium bromide (40 mM), in a solvent mixture containing 6 % methanol and 10 % acetic acid in acetonitrile/chlorobenzene (1:1 v/v) as the background electrolyte. Detection limits, based on a signal-to-noise ratio of 3:1, were calculated, and values between 1 and 3.7 mg/L were obtained. Good run-to-run and day-to-day precisions on concentration were achieved with relative standard deviation lower than 15 %. For the first time, an electrophoretic technique has been applied for the analysis of C60 fullerene in a commercial cosmetic cream. A standard addition method was used for quantitation, and the result was compared with that obtained by analyzing the same cream by liquid chromatography coupled to mass spectrometry.
Keywords: Fullerenes; Tetraalkylammonium salts; Non-aqueous capillary electrophoresis; Commercial cosmetic cream
Differential protein expression of hepatic cells associated with MeHg exposure: deepening into the molecular mechanisms of toxicity by Susana Cuello; Sonia Ramos; Yolanda Madrid; Jose L. Luque-Garcia; Carmen Cámara (315-324).
Understanding the molecular mechanisms underlying MeHg toxicity and the way in which this molecule interacts with living organisms is a critical point since MeHg represents a well-known risk to ecosystems and human health. We used a quantitative proteomic approach based on stable isotopic labeling by amino acids in cell culture in combination with SDS-PAGE and nanoflow LC-ESI-LTQ for analyzing the differential protein expression of hepatic cells associated to MeHg exposure. Seventy-eight proteins were found de-regulated by more than 1.5-fold. We identified a number of proteins involved in different essential biological processes including apoptosis, mitochondrial dysfunction, cellular trafficking and energy production. Among these proteins, we found several molecules whose de-regulation has been already related to MeHg exposure, thus confirming the usefulness of our discovery approach, and new ones that helped to gain a deeper insight into the biomolecular mechanisms related to MeHg-induced toxicity. Overexpression of several HSPs and the proteasome 26S subunit itself showed the proteasome system as a molecular target of toxic MeHg. As for the interaction networks, the top ranked was the nucleic acid metabolism, where many of the identified de-regulated proteins are involved.
Keywords: Bioanalytical methods; Proteomics; Toxicology; Heavy metals; Mass spectrometry
Identification of budesonide metabolites in human urine after oral administration by Xavier Matabosch; Oscar J. Pozo; Clara Pérez-Mañá; Magi Farré; Josep Marcos; Jordi Segura; Rosa Ventura (325-340).
Budesonide (BUD) is a glucocorticoid widely used for the treatment of asthma, rhinitis, and inflammatory bowel disease. Its use in sport competitions is prohibited when administered by oral, intravenous, intramuscular, or rectal routes. However, topical preparations are not prohibited. Strategies to discriminate between legal and forbidden administrations have to be developed by doping control laboratories. For this reason, metabolism of BUD has been re-evaluated using liquid chromatography–tandem mass spectrometry (LC-MS/MS) with different scan methods. Urine samples obtained after oral administration of 3 mg of BUD to two healthy volunteers have been analyzed for metabolite detection in free and glucuronide metabolic fractions. Structures of the metabolites have been studied by LC-MS/MS using collision induced dissociation and gas chromatography–mass spectrometry (GC/MS) in full scan mode with electron ionization. Combination of all structural information allowed the proposition of the most comprehensive picture for BUD metabolism in humans to this date. Overall, 16 metabolites including ten previously unreported compounds have been detected. The main metabolite is 16α-hydroxy-prednisolone resulting from the cleavage of the acetal group. Other metabolites without the acetal group have been identified such as those resulting from reduction of C20 carbonyl group, oxidation of the C11 hydroxyl group and reduction of the A ring. Metabolites maintaining the acetal group have also been identified, resulting from 6-hydroxylation (6α and 6β-hydroxy-budesonide), 23-hydroxylation, reduction of C6-C7, oxidation of the C11 hydroxyl group, and reduction of the C20 carbonyl group. Metabolites were mainly excreted in the free fraction. All of them were excreted in urine during the first 24 h after administration, and seven of them were still detected up to 48 h after administration for both volunteers.
Keywords: Budesonide; Metabolism; Mass spectrometry; Doping analysis; Urine
Gold-nanosphere formation using food sample endogenous polyphenols for in-vitro assessment of antioxidant capacity by Diana Vilela; María Cristina González; Alberto Escarpa (341-349).
In this work it was demonstrated that sample endogenous polyphenols are selectively driving the gold-nanoparticle (AuNPs)-formation process when representative food samples were used as natural sources of reducing compounds. The process of AuNPs formation was characterized by UV–visible spectroscopy and was described by a sigmoidal curve (R 2 ≥ 0.990) which gave information about the polyphenol concentration at which the localized surface plasmon resonance (LSPR) absorption reached its half-value, X c 50 , and about AuNPs production per polyphenol concentration unit, K AuNPs. The behavior of phenolic acids was different, with lower X c 50 and higher K AuNPs values than flavonoids. For the food samples tea, apple, pear, wine, and honey X c 50 values were 0.22, 7.3, 11.5, 20.4, 30.3, and 53.5 (mg mL−1) and K AuNPs values were 28.7, 0.70, 0.60, 0.20, 0.14, and 0.10 (mg−1 mL), respectively. Excellent correlation between K AuNPs and total phenolics (TP) was obtained (r = 0.98, p-value < 0.05), implying K AuNPs is a novel marker for evaluation of food sample antioxidant capacity in vitro. The K AuNPs values of samples indicated their antioxidant capacity was in the order: tea > apple > pear > wine > honey. The reproducibility of the AuNPs formation approach was excellent, not only for polyphenol standards (RSD < 6 % for X c 50 and RSD < 11 % for K AuNPs) but also for food samples (RSD < 9 % for X c 50 and RSD < 15 % for K AuNPs). Transmission electronic microscopy (TEM) enabled confirmation of the formation of stabilized Au-nanospheres from endogenous polyphenols with very well-defined sizes under 20 nm diameter for all the food samples investigated. Figure Gold-nanosphere formation using food polyphenols for antioxidant activity assessment
Keywords: Gold nanoparticle formation; Antioxidant capacity; Food sample endogenous polyphenols
Autoindicating optical properties of laccase as the base of an optical biosensor film for phenol determination by J. Sanz; S. de Marcos; J. Galbán (351-359).
In the context of sustainable analytical chemistry, phenol has been determined through its enzymatic reaction with laccase. The method has been studied and optimized through the autoindicating optical properties of laccase both by intrinsic molecular absorption and fluorescence. The method shows a linear range from 9.79·10−6 to 7.50·10−4 M with a relative standard deviation of 1.07 %. The molecular absorption methodology has been implemented in a polyacrylamide film for the design of an autoindicating optical sensor. In order to increase the lifetime of the sensor, the reversibility study of the enzymatic reaction has proposed, as a novelty, the regeneration of laccase with an oxidase-type enzyme (glucose oxidase). The lifetime of the sensor film has improved from 15 to 30 measurements. The reaction mechanism has also been studied and confirmed by fluorescence and molecular absorption. The method leads to the determination of phenol in environmental samples.
Keywords: Laccase; Phenol; Autoindicating; Protein; Sensor film; Enzymatic
Pressurized liquid extraction as a sample preparation method for the analysis of isoflavones in pulses by Ma. Milagros Delgado-Zamarreño; Lara Pérez-Martín; Myriam Bustamante-Rangel; Rita Carabias-Martínez (361-366).
In this work, we describe a rapid and simple analytical method that exploits pressurized liquid extraction (PLE) and liquid chromatography with diode array detection for the determination of isoflavones in samples of Spanish pulses. Confirmation of the analytes present was performed using ion-trap mass spectrometry. To optimize the PLE extraction, variables such as the dispersing agent, type of solvent and sample amount, and the experimental parameters, such as temperature and the number of extraction cycles, were studied. Separation was carried out using a reverse-phase C18 with polar endcapping as the stationary phase and acetonitrile/water with 0.2 % of formic acid, under a gradient regime, as the mobile phase. Optimal extraction of formononetin and biochanin-A from chickpeas with PLE was achieved using Hydromatrix as a dispersant agent, methanol/water (50:50), a temperature of 90 °C, and three cycles. The same optimal conditions—except methanol/water (75:25)—for solvent extraction were obtained for the extraction of daidzin, genistin, and formononetin from lentils. Recoveries ranged from 97 to 110 %, and standard deviations lower than 20 % were obtained. The contents obtained for daidzin in lentils using the proposed method were not significantly different from those obtained using another official method of analysis.
Keywords: Isoflavones; Pressurized liquid extraction; Chickpeas; Lentils; Liquid chromatography
Collision energy alteration during mass spectrometric acquisition is essential to ensure unbiased metabolomic analysis by Ntakadzeni E. Madala; Paul A. Steenkamp; Lizelle A. Piater; Ian A. Dubery (367-372).
Metabolomics entails identification and quantification of all metabolites within a biological system with a given physiological status; as such, it should be unbiased. A variety of techniques are used to measure the metabolite content of living systems, and results differ with the mode of data acquisition and output generation. LC–MS is one of many techniques that has been used to study the metabolomes of different organisms but, although used extensively, it does not provide a complete metabolic picture. Recent developments in technology, for example the introduction of UPLC–ESI–MS, have, however, seen LC–MS become the preferred technique for metabolomics. Here, we show that when MS settings are varied in UPLC–ESI–MS, different metabolite profiles result from the same sample. During use of a Synapt UPLC–high definition MS instrument, the collision energy was continually altered (3, 10, 20, and 30 eV) during MS acquisition. PCA and OPLS-DA analysis of the generated UPLC–MS data of metabolites extracted from elicited tobacco cells revealed different clustering and different distribution patterns. As expected, ion abundance decreases with increasing collision energy, but, more importantly, results in unique multivariate data patterns from the same samples. Our findings suggest that different collision energy settings should be investigated during MS data acquisition because these can contribute to coverage of a wider range of the metabolome by UPLC–ESI–MS and prevent biased results.
Keywords: Metabolomics; Collision energy; 2-Isonitrosoacetophenone; UPLC–MS; PCA; OPLS-DA
Comprehensive characterization of the N-glycosylation status of CD44s by use of multiple mass spectrometry-based techniques by Huanhuan Han; Martha Stapels; Wantao Ying; Yingqing Yu; Li Tang; Wei Jia; Weibin Chen; Yangjun Zhang; Xiaohong Qian (373-388).
The CD44 family are type-1 transmembrane glycoproteins which are important in mediating the response of cells to their microenvironment, including regulation of growth, survival, differentiation, and motility. All these important functions have been reported to be regulated by N-glycosylation; however, little is known about this process. In the CD44 family, the most prolific isoform is CD44 standard type (CD44s). In this work, an integrated strategy combining stable isotope labeling, chemical derivatization, hydrophilic-interaction liquid chromatographic (HILIC) separation, and mass spectrometric (MS) identification was used to perform a comprehensive qualitative and quantitative survey of the N-glycosylation of recombinant CD44s. Specifically, the occupation ratios of the N-glycosites were first determined by MS with 18O labeling; the results revealed five glycosites with different occupation ratios. Next, N-glycans were profiled by chemical derivatization and exoglycosidase digestion, followed by MALDI–TOF-MS and HILIC–ESI–MS–MS analysis. Interestingly, the quantitative analysis showed that non-sialylated, fucosylated complex-type glycans dominated the N-glycans of CD44s. Furthermore, the site-specific N-glycan distributions profiled by LC–ESI–MSE indicated that most glycosites bore complex-type glycans, except for glycosite N100, which was occupied by high-mannose-type N-glycans. This is the first comprehensive report of the N-glycosylation of CD44s. Figure Strategies for characterization of the N-glycosylation status of CD44s
Keywords: CD44s; Liquid chromatography; Mass spectrometry; N-glycosylation; Site-specific
Improved spatial resolution in the imaging of biological tissue using desorption electrospray ionization by Dahlia I. Campbell; Christina R. Ferreira; Livia S. Eberlin; R. Graham Cooks (389-398).
Desorption electrospray ionization imaging allows biomarker discovery and disease diagnosis through chemical characterization of biological samples in their native environment. Optimization of experimental parameters including emitter capillary size, solvent composition, solvent flow rate, mass spectrometry scan-rate and step-size is shown here to improve the resolution available in the study of biological tissue from 180 μm to about 35 μm using an unmodified commercial mass spectrometer. Mouse brain tissue was used to optimize and measure resolution based on known morphological features and their known relationships to major phospholipid components. Features of approximately 35 μm were resolved and correlations drawn between features in grey matter (principally PS (18:0/22:6), m/z 834) and in white matter (principally ST (24:1), m/z 888). The improved spatial resolution allowed characterization of the temporal changes in lipid profiles occurring within mouse ovaries during the ovulatory cycle. An increase in the production of phosphatidylinositol (PI 38:4) m/z 885 and associated fatty acids such as arachidonic acid (FA 20:4) m/z 303 and adrenic acid (FA 22:4) m/z 331was seen with the postovulatory formation of the corpus luteum.
Keywords: Ambient ionization; Tissue imaging; Electrospray ionization; Imaging mass spectrometry; High-resolution imaging; Morphologically friendly solvents; Histology; Ovulation; Corpus luteum
Detection of prolactin inducible protein mRNA, a biomarker for breast cancer metastasis, using a molecular beacon-based assay by Erik D. Guetschow; Will Black; Carolyn M. Walsh; Jennifer R. W. Furchak (399-406).
Mortality due to breast cancer is increasingly linked to early, undetected metastasis, making methods for earlier detection acutely necessary. We describe the development of an assay based on molecular beacon (MB) chemistry with fluorescence detection to monitor a breast cancer biomarker for the analysis of breast cancer metastasis. The MB assay is based on the complementary base-pairing interactions of the MB nucleic acid with mRNA indicative of breast cancer metastasis. The presence of mRNA is characterized by an increase in the fluorescence intensity of the molecular beacon. The assay gives a linear, reproducible response to prolactin inducible protein mRNA, with a limit of detection in the high picomolar range. This method sensitively and specifically identifies a biomarker directly in serum samples in minimal time and with a straightforward procedure, dramatically reducing the total time for sample analysis over current methods from days to hours. The potential impact of this work in detection and understanding of breast cancer metastasis lies in improvements in simplicity, accuracy, and speed over current methods, which could allow for improved patient treatment and prognoses. Ultimately, additional sample throughput will result in better understanding of disease progression. Figure Hybridization of MB with PIP mRNA results in an increase in fluorescence intensity. Heat denaturing and slowing annealing the nucleic acids dramatically increases the fluorescence intensity as the MB and mRNA form a stable duplex. Used in an assay, the response is linear and reproducible, while the measurement is straightforward and fast
Keywords: Fluorescence/luminescence; Nucleic acids (DNA|RNA); Bioanalytical methods; Bioassays
A high-throughput sphingomyelinase assay using natural substrate by Miao Xu; Ke Liu; Noel Southall; Juan J. Marugan; Alan T. Remaley; Wei Zheng (407-414).
Sphingomyelinases are a group of hydrolases that cleave sphingomyelin, a common component of plasma membranes, to form ceramide and phosphocholine. Ceramide is a second messenger that is present in virtually all cell types and regulates a variety of cellular functions such as proliferation, differentiation, apoptosis, and inflammation response. Inhibition of sphingomyelinase activity to reduce ceramide concentrations has recently emerged as a potential therapeutic approach for several diseases including atherosclerosis, pathogen infections, inflammation, diabetes, and obesity. To effectively screen compound collections for the identification of new sphingomyelinase inhibitors, we have developed a high-throughput assay utilizing the natural substrate sphingomyelin in 1,536-well plate format. The assay has a signal-to-basal ratio of 6.1-fold in pH 5.0 buffer and 4.3-fold in pH 6.5 buffer, indicating a robust assay for compound library screening. A screen of ~300,000 compounds using this assay led to the identification of eight compounds as sphingomyelinase inhibitors (IC50s = 1.7 to 38.2 μM) that exhibited different activities between the natural substrate assay and profluorescence substrate assay. The results demonstrate the robustness and effectiveness of the natural substrate sphingomyelinase assay for screening sphingomyelinase inhibitors.
Keywords: Acid sphingomyelinase; ASM inhibitors; High-throughput screening; Natural enzyme substrate
Enzyme-free detection and quantification of double-stranded nucleic acids by Cécile Feuillie; Maxime Mohamad Merheb; Benjamin Gillet; Gilles Montagnac; Catherine Hänni; Isabelle Daniel (415-422).
We have developed a fully enzyme-free SERRS hybridization assay for specific detection of double-stranded DNA sequences. Although all DNA detection methods ranging from PCR to high-throughput sequencing rely on enzymes, this method is unique for being totally non-enzymatic. The efficiency of enzymatic processes is affected by alterations, modifications, and/or quality of DNA. For instance, a limitation of most DNA polymerases is their inability to process DNA damaged by blocking lesions. As a result, enzymatic amplification and sequencing of degraded DNA often fail. In this study we succeeded in detecting and quantifying, within a mixture, relative amounts of closely related double-stranded DNA sequences from Rupicapra rupicapra (chamois) and Capra hircus (goat). The non-enzymatic SERRS assay presented here is the corner stone of a promising approach to overcome the failure of DNA polymerase when DNA is too degraded or when the concentration of polymerase inhibitors is too high. It is the first time double-stranded DNA has been detected with a truly non-enzymatic SERRS-based method. This non-enzymatic, inexpensive, rapid assay is therefore a breakthrough in nucleic acid detection.
Keywords: SERRS; Double-stranded DNA; Non-enzymatic
Surface plasmon resonance imaging in arrays coupled with mass spectrometry (SUPRA–MS): proof of concept of on-chip characterization of a potential breast cancer marker in human plasma by F. Remy-Martin; M. El Osta; G. Lucchi; R. Zeggari; T. Leblois; S. Bellon; P. Ducoroy; W. Boireau (423-432).
Protein biomarker discovery and validation are crucial for diagnosis, prognosis, and theranostics of human pathologies; “omics” approaches bring new insights in this field. In particular, the combination of immuno-sensors in array format with mass spectrometry efficiently extends the classical immunoassay format and includes molecular characterization. Here, we coupled surface plasmon resonance imaging (SPRi) with MALDI-TOF mass spectrometry in a hyphenated technique which enables multiplexed quantification of binding by SPRi and molecular characterization of interacting partners by subsequent MS analysis. This adds specificity, because MS enables differentiation of molecules that are difficult to distinguish by use of antibodies, for example truncation variants or protein isoforms. Proof of concept was established for detection, identification, and characterization of a potential breast cancer marker, the LAG3 protein, at ~1 μg mL−1, added to human plasma. The analytical performance of this new method, dubbed “SUPRA-MS”, was established, particularly its specificity (S/N > 10) and reliability (100 % LAG3 identification with high significant mascot score >87.9). The adjusted format for rapid, collective, and automated on-chip MALDI-MS analysis is robust at the femtomole level and has numerous potential applications in proteomics.
Keywords: Surface plasmon resonance; Mass spectrometry; Immuno sensor; Biomarker; Proteomics
Rapid narrow band elution for on-line SPE using a novel solvent plug injection technique by David Gode; Markus M. Martin; Frank Steiner; Christian G. Huber; Dietrich A. Volmer (433-445).
Determination of trace constituents in biological and environmental samples usually requires a pre-concentration step. While solid-phase extraction (SPE) has been widely used, it is slow, labor intensive and adversely affected by analytical errors from handling. On-line SPE eliminates some of the flaws but often suffers from solvent compatibility problems with the subsequent chromatography separation. In this study, we are presenting a technical solution for overcoming some of these compatibility issues, by utilizing a fully automated, focused SPE sample transfer technique utilizing narrow-band solvent plugs, for seamless hyphenation with high-performance liquid chromatography (HPLC) or flow injection mass spectrometry (MS). A wide range of pharmaceutical compounds was studied in different sample matrices. Short plugs of high elution strength solvent were generated by means of an electrically actuated sample loop and enrichment and transfer steps monitored using on-line SPE-MS. The impact of the solvent plugs on chromatographic separation was studied using hyphenated SPE-LC-MS. By carefully examining elution profiles of solvent plugs of different compositions, optimum conditions for quantitative elution within well-defined volumes were found for all substances. In addition, the highly focused elution bands resulted in excellent retention time and peak area reproducibilities when injected on-line onto HPLC columns. Finally, to demonstrate proof-of-principle, the fully integrated on-line SPE-LC-MS system was applied to the analysis of spiked urine and river water samples. Figure Rapid narrow bad elution technique for on-line SPE
Keywords: HPLC; Mass spectrometry; On-line SPE; Sample preparation; Solid-phase extraction; Solvent plug
Comparison of dialysis and solid-phase extraction for isolation and concentration of dissolved organic matter prior to Fourier transform ion cyclotron resonance mass spectrometry by Malak M. Tfaily; Suzanne Hodgkins; David C. Podgorski; Jeffrey P. Chanton; William T. Cooper (447-457).
We compare two methods, solid-phase extraction (SPE) and dialysis, commonly used for extraction and concentration of dissolved organic matter (DOM) prior to molecular characterization by electrospray ionization (ESI) and ultrahigh-resolution Fourier transform ion cyclotron resonance mass spectrometry. Spectra of DOM samples from Minnesota and Sweden peatlands that were extracted with styrene divinyl benzene polymer SPE sorbents included ions with formulas that had higher oxygen to carbon (O/C) ratios than spectra of DOM from the same samples after de-salting by dialysis. The SPE method was not very effective in extracting several major classes of DOM compounds that had high ESI efficiencies, including carboxylic acids and organo-sulfur compounds, and that out-competed other less-functionalized compounds (e.g., carbohydrates) for charge in the ESI source. The large abundance of carboxylic acids in the dialysisextracted DOM, likely the result of in situ microbial production, makes it difficult to see other (mainly hydrophilic) compounds with high O/C ratios. Our results indicate that, while dialysis is generally preferable for the isolation of DOM, for samples with high microbial inputs, the use of both isolation methods is recommended for a more accurate molecular representation. Figure van Krevelen diagrams depicting elemental O/C and H/C ratios of sulfur-containing compounds unique to dialysis- and SPE-extracted DOM. (a) Minnesota bog, (b) Swedish bog, and (c) Minnesota fen.
Keywords: Solid-phase extraction; Dialysis; Peatlands; Dissolved organic matter; Ultrahigh-resolution mass spectrometry of DOM
Clean-up method for determination of established and emerging brominated flame retardants in dust by Leena Sahlström; Ulla Sellström; Cynthia A. de Wit (459-466).
A clean-up method was developed to enable the determination of tri-decabrominated diphenyl ethers, isomer-specific hexabromocyclododecanes (HBCDs), 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE), decabromodiphenyl ethane (DBDPE), (2-ethylhexyl)tetrabromobenzoate (TBB), and bis(2-ethylhexyl)tetrabromophthalate (TBPH) in the same dust sample extract using reasonable amounts of solvents and without dividing the sample. After extraction, the sample was separated on a silica column into three fractions that were subsequently cleaned up individually. The polybrominated diphenyl ethers (PBDEs) and DBDPE were eluted in Fraction I, TBB, TBPH, and BTBPE in Fraction II, and HBCDs in Fraction III. Fractions I and II were analyzed using gas chromatography/mass spectrometry and Fraction III using liquid chromatography/mass spectrometry. The method gave good recoveries (60–120%), precise results using 13 C-labelled internal standards and was accurate when comparing results to certified values (PBDEs in NIST SRM 2585). The method was applied to dust samples from the Stockholm (Sweden) area. All the emerging brominated flame retardants (BFRs) studied, except BTBPE, were present in all the samples in quantifiable concentrations, often higher than the PBDEs. BTBPE was quantified in only one sample. It is evident that emerging BFRs are present in Swedish homes, and these compounds should be included in the BFR analyses of indoor environments. Figure Clean-up method for determination of BFRs in dust
Keywords: Emerging BFRs; PBDE; HBCD; Analytical method; Dust
Determination of synthetic musk compounds in indoor house dust by gas chromatography–ion trap mass spectrometry by Cariton Kubwabo; Xinghua Fan; Pat E. Rasmussen; Fang Wu (467-477).
A new method for the simultaneous determination of 11 synthetic musks and one fragrance compound in house dust was developed. The nitro musks included musk ketone (MK, 4-tert-butyl-3,5-dinitro-2,6-dimethylacetophenone), musk xylene (MX, 1-tert-butyl-3,5-dimethyl-2,4,6-trinitrobenzene), musk ambrette (1-tert-butyl-2-methoxy-4-methyl-3,5-dinitrobenzene) and musk moskene (1,1,3,3,5-pentamethyl-4,6-dinitroindane). The polycyclic musk compounds were 1,3,4,6,7,8-hexahydro-4,6,6,7,8,8-hexamethylcyclopenta-(γ)-2-benzopyran (HHCB), 7-acetyl-1,1,3,4,4,6-hexamethyl-1,2,3,4-tetrahydronaphthalene (AHTN), 4-acetyl-1,1-dimethyl-6-tert-butylindane, 6-acetyl-1,1,2,3,3,5-hexamethylindane, 5-acetyl-1,1,2,6-tetramethyl-3-isopropylindane, 6,7-dihydro-1,1,2,3,3-pentamethyl-4(5H)-indanon. The one macrocyclic musk was 1,4-dioxacycloheptadecane-5,17-dione. The bicyclic hydrocarbon fragrance compound (1,2,3,4,5,6,7,8-octahydro-2,3,8,8-tetramethylnaphthalen-2yl)ethan-1-one (OTNE) and HHCB-lactone (4,6,6,7,8,8-hexamethyl-1H,3H,4H,6H,7H, 8H-indeno[5,6-c]pyran-1-one), a degradation product of HHCB, were also analysed. NIST SRM 2781 (domestic sludge) and SRM 2585 (organic contaminants in house dust) were analysed for these target compounds. The method was applied for the analysis of 49 paired samples collected using two vacuum sampling methods: a sample of fresh or “active” dust (FD) collected using a Pullman–Holt vacuum sampler, and a household dust (HD) sample taken from the participants’ vacuum cleaners. Method detection limits and recoveries ranged from 12 to 48 ng/g and 54 to 117 %, respectively. AHTN, HHCB, OTNE and HHCB-lactone were detected in all samples, with median concentrations of 552, 676, 252 and 453 ng/g for FD samples, respectively; and 405, 992, 212 and 492 ng/g for HD samples, respectively. MX and MK were detected with high frequencies but with much lower concentrations. The two sampling methods produced comparable results for the target analytes. Widely scattered concentration levels were observed for target analytes from this set of 49 house dust samples, suggesting a wide variability in Canadian household exposure to synthetic musks.
Keywords: GC/ITMS; Indoor house dust; Sampling techniques; Nitro musks; Polycyclic musks; Macrocyclic musks
Chemical interactions in the layered system BC x N y /Ni(Cu)/Si, produced by CVD at high temperature by P. S. Hoffmann; M. I. Kosinova; S. Flege; O. Baake; B. Pollakowski; V. A. Trunova; A. Klein; B. Beckhoff; F. A. Kuznetsov; W. Ensinger (479-487).
Layered samples Si(100)/C/Ni/BC x N y and Si(100)/C/Cu/BC x N y were produced by physical vapor deposition of a metal (Ni, Cu, resp.) and low-pressure chemical vapor deposition of the boron carbonitride on a Si(100) substrate. Between the Si and the Ni (Cu) and on the surface of the Ni (Cu) layer, thin carbon layers were deposited, as a diffusion barrier or as a protection against oxidation, respectively. Afterwards, the surface carbon layer was removed. As precursor, trimethylamine borane and, as an auxiliary gas, H2 and NH3 were used, respectively. The chemical compositions of the layers and of the interfaces in between were characterized by total-reflection X-ray fluorescence spectrometry combined with near-edge X-ray absorption fine-structure spectroscopy, X-ray photoelectron spectroscopy, and secondary ion mass spectrometry. The application of H2 yielded the BC x N y compound whereas the use of NH3 led to a mixture of h-BN and graphitic carbon. At the BC x N y /metal interface, metal borides could be identified. At the relatively high synthesis temperature of 700 °C, broad regions of Cu or Ni and Si were observed between the metal layer and the substrate Si.
Keywords: Boron carbonitride/metal/silicon-layered system; Interfaces; Near-edge X-ray absorption fine-structure spectroscopy; X-ray photoelectron spectroscopy; Secondary ion mass spectroscopy
Monolithic molecularly imprinted solid-phase extraction for the selective determination of trace cytokinins in plant samples with liquid chromatography–electrospray tandem mass spectrometry by Fuyou Du; Guihua Ruan; Shenghua Liang; Fuwei Xie; Huwei Liu (489-501).
Cytokinins (CTKs) are a class of growth-regulating hormones involved in various physiological and developmental processes. More novel analytical methods for the accurate identification and quantitative determination of trace CTKs in plants have been desired to better elucidate the roles of CTKs. In this work, a novel method based on monolithic molecularly imprinted solid-phase extraction followed by liquid chromatography–electrospray tandem mass spectrometry (mMI-SPE-LC-MS/MS) was developed for accurate determination of four CTKs in plant samples. The molecularly imprinted polymer monolith was prepared by using kinetin as the template in syringes and exhibited specific recognition ability for the four CTKs in comparison with that of non-imprinted polymer monolith. Several factors affecting the extraction performance of mMI-SPE, including the pH of loading sample solution, the nature and volume of elution solvent, the flow rate of sample loading, and sample volume, were investigated, respectively. Under the optimized conditions, the proposed mMI-SPE-LC-MS/MS method was successfully applied in the selective extraction and determination of four CTKs in plant tissues, and it offers detection limits (S/N = 3) of 104, 113, 130, and 89 pg/mL and mean recoveries of 85.9%, 79.3%, 73.5%, and 70.1% for kinetin, kinetin glucoside, trans-zeatin, and meta-topolin (mT), respectively, with the corresponding RSDs less than 15%. Figure Trace determination of cytokinins in different plant samples by monolithic molecularly imprinted solid-phase extraction and liquid chromatography–mass spectrometry
Keywords: Cytokinins; Molecularly imprinted polymer; Solid-phase extraction; Liquid chromatography–electrospray tandem mass spectrometry
A novel test using dried blood spots for the chromatographic assay of methadone by Maria Addolorata Saracino; Chiara Marcheselli; Lorenzo Somaini; Maria Chiara Pieri; Gilberto Gerra; Anna Ferranti; Maria Augusta Raggi (503-511).
A novel test has been developed for the analysis of methadone in dried blood spot specimens from patients undergoing methadone maintenance treatment. An isocratic reversed-phase high-performance liquid chromatography method with coulometric detection has been optimized for the determination of methadone. The clean-up of dried blood spots was performed by means of an original microextraction by packed sorbent procedure after microwave-assisted extraction of the drug with a suitable solvent. Extraction yields were satisfactory, always being higher than 90.0 %. The calibration curve was linear over the 4–500 ng mL-1 concentration range. The method had satisfactory sensitivity (limit of quantitation of 4 ng mL-1), precision (relative standard deviation less than 5.8 %), selectivity and accuracy (recovery greater than 87.0 %). It was successfully applied to dried blood spot samples collected from heroin-addicted patients undergoing methadone maintenance therapy at dosages between 40 and 240 mg day-1. The statistical analysis (Bland–Altman plot) showed that the results were in good agreement with those found from the analysis of plasma samples obtained from the same patients. Thus, the method has proved to be suitable for the monitoring of methadone by means of dried blood spots. Figure Bland-Altman plot and methadone disposition in patients undergoing methadone maintenance treatment
Keywords: Methadone; Methadone maintenance treatment; Dried blood spots; Liquid chromatography; Microwave-assisted extraction; Microextraction by packed sorbent
An automated method for the simultaneous measurement of azole antifungal drugs in human plasma or serum using turbulent flow liquid chromatography-tandem mass spectrometry by L. Couchman; S. L. Buckner; P. E. Morgan; M. M. Ceesay; A. Pagliuca; R. J. Flanagan (513-523).
Azole antifungal drugs are important in the prophylaxis and treatment of invasive aspergillosis. Therapeutic drug monitoring may be indicated to (1) monitor adherence, (2) guide dosage and (3) minimise the risk of drug–drug interactions and dose-related toxicity. TurboFlowTM technology offers online, automated sample preparation. An Aria TranscendTM TLX-II coupled with a TSQ VantageTM MS was used. Centrifuged samples (25 μL) were mixed with internal standard solution (975 μL) and 30 μL injected directly onto a C18-P-XL TurboFlow column. Analytes were focussed onto a Phenomenex Gemini Phenyl analytical column and eluted using a methanol/water gradient (flow-rate, 0.8 mL/min). Analytes were monitored in selected reaction monitoring mode (two transitions per analyte, positive mode APCI). Calibration ranges were as follows: itraconazole, hydroxyitraconazole, and posaconazole 0.05–5.0 mg/L; voriconazole and fluconazole 0.1–10 mg/L. Total analysis time was 12 min. TurboFlow column recovery was >77% for all analytes. Calibration was linear (R 2 > 0.99) for all analytes. Inter- and intra-assay imprecision (% RSD) was <8% and accuracy (nominal internal quality control values) 90–105% for all analytes. The limit of detection was 0.01 mg/L for all analytes. No matrix effects were observed. This method is simple, robust and suitable for measuring these compounds at concentrations attained during therapy.
Keywords: TurboFlow MS/MS; Azole antifungals; Itraconazole; Posaconazole; Voriconazole; Automated sample preparation
Polymer-modified glassy carbon electrode for the electrochemical detection of quinine in human urine and pharmaceutical formulations by Alemnew Geto; Meareg Amare; Merid Tessema; Shimelis Admassie (525-530).
Glassy carbon electrode was modified by electropolymerization of 4-amino-3-hydroxynaphthalene sulfonic acid. Cyclic voltammetric study of quinine showed higher current response at the modified electrode compared to the bare and activated glassy carbon electrodes in pH 7.0 phosphate buffer solution. Under optimized conditions, a calibration curve was obtained by square wave voltammetry at the modified electrode. The linear relationship between the peak current and the concentration of quinine in the range of 1.0 × 10−7 to 1.0 × 10−5 M was I pa (in microamperes) = 6.26C (in micromolars) + 0.2997 (R 2 = 0.999). The detection limit calculated (S/N = 3) was 1.42 × 10−8 M, which is much lower than similar reports. The method was successfully applied for the determination of quinine in spiked human urine, and pharmaceutical formulations and recovery values >90 % were obtained.
Keywords: Glassy carbon electrode; Pharmaceutical; Modified electrode; Urine; Quinine; Voltammetry
Analysis of lysergic acid amide in human serum and urine after ingestion of Argyreia nervosa seeds by Alexander Paulke; Christian Kremer; Cora Wunder; Stefan W. Toennes (531-538).
The ergot alkaloid lysergic acid amide (LSA) is a secondary plant constituent in a number of plants, but it is mainly present in considerable amounts in Convolvulaceae, like Argyreia nervosa. Due to its close structural similarity to lysergic acid diethylamide, LSA is considered as psychedelic and therefore promoted as so-called “legal high” in various internet forums. During a human behavioral study with orally administered seeds of A. nervosa, blood and urine samples were obtained. The present study describes the validation of a sensitive and robust high performance liquid chromatography method with fluorescence detection, which was applied to the study samples. The limit of detection (LOD) and lower limit of quantification in human serum were 0.05 and 0.17 ng/mL, respectively, and in urine, the LOD was 0.15 ng/mL. Intra- and interday precision and accuracy were below 15 % relative standard deviation with a bias better than ±15 %. No conversion of LSA to its epimer iso-LSA was noted during analyses. The LSA concentrations in the authentic human serum samples were in the range of 0.66 to 3.15 ng/mL approximately 2 h after ingestion. In urine, LSA could be found 1–24 h after ingestion; after 48 h, no LSA could be detected. The LSA epimer iso-LSA was also detected in serum and urine in varying ratios. In conclusion, LSA serum levels in the low nanogram per milliliter range correlated with severe vegetative adverse effects (nausea, weakness, fatigue, tremor, blood pressure elevation) and a psychosis-like state, which led to study termination.
Keywords: Lysergic acid amide/LSA; HPLC; Argyreia nervosa ; Blood/serum
Determination of S-containing drug metabolites from in vitro and in vivo metabolism studies by using LC-ICP/MS by Cristina Losada; Joan J. Albertí; Javier Saurina; Sonia Sentellas (539-551).
Recently, liquid chromatography coupled to inductively coupled plasma mass spectrometry (LC-ICP/MS) has been introduced to deal with some applications in the field of pharmaceutical, biomedical, and clinical analysis. In the case of drug research, the number of drugs and their metabolites containing detectable elements is quite limited. In this paper, LC-ICP/MS has been demonstrated to be suitable for the determination of S-containing drugs and their metabolites. In order to minimize the interference of polyatomic oxygen (m/z 32), the indirect detection of S, by means of the SO+ ion (m/z 48), was optimized. For quantification purposes, it has been encountered that the percentage of organic solvent in the mobile phase strongly affects the sensitivity. Here, corrective strategies based on calibration curves established at different solvent concentrations (solvent-zone quantification) and post-column gradient compensation have been proposed to circumvent sensitivity variations. Results obtained have shown that suitable calibration models have been built for any compound regardless of the solvent percentage at which it is eluted from the chromatographic column. To prove the applicability of this methodology, the metabolism of ethacrynic acid and tiotropium bromide has been studied in vitro and in vivo. In the first case, ethacrynic acid does not contain S in its structure, however, the major route of metabolism for this compound consists of the formation of glutathione adduct and its further degradation. In the second case, tiotropium bromide contains two S atoms in its structure. Figure Determination of S-containing drug metabolites from in vitro and in vivo metabolism studies by LC-ICP/MS. Quantification strategies (solvent-zone quantification and post-column gradient compensation) proposed in order to minimize the effect of organic solvent in the quantification.
Keywords: Sulfur; Drug metabolite; LC-ICP/MS; Glutathione; Ethacrynic acid; Tiotropium bromide; Speciation analysis
Global profiling of ultraviolet-induced metabolic disruption in Melissa officinalis by using gas chromatography-mass spectrometry by Sooah Kim; Eun Ju Yun; Md. Aktar Hossain; Hojoung Lee; Kyoung Heon Kim (553-562).
Melissa officinalis contains various secondary metabolites that have health benefits. Generally, irradiating plants with ultraviolet (UV)-B induces the accumulation of secondary metabolites in plants. To understand the effect of UV-B irradiation on the metabolism of M. officinalis, metabolomics based on gas chromatography-mass spectrometry (GC-MS) was used in this study. The GC-MS analysis revealed 37 identified metabolites from various chemical classes, including alcohols, amino acids, inorganic acids, organic acids, and sugars. The metabolite profiles of the groups of M. officinalis irradiated with UV-B were separated and differentiated according to their irradiation times (i.e., 0, 1, and 2 h), using principal component analysis (PCA) and hierarchical clustering analysis (HCA), respectively. The PCA score plots of PC1 and PC2 showed that the three groups with different irradiation times followed a certain trajectory with increasing UV-B irradiation. HCA revealed that metabolic patterns differed among the three groups, and the 1 h-irradiated group was more similar to the control group (0 h) than the 2 h-irradiated group. In particular, UV-B irradiation of plants led to a decrease in sugars such as fructose, galactose, sucrose, and trehalose and an increase in metabolites in the tricarboxylic acid cycle, the proline-linked pentose phosphate pathway, and the phenylpropanoid pathway. This study demonstrated that metabolite profiling with GC-MS is useful for gaining a holistic understanding of UV-induced changes in plant metabolism.
Keywords: Metabolomics; Metabolite profiling; Melissa officinalis ; Ultraviolet; Gas chromatography-mass spectrometry
Spectral signatures for the classification of microbial species using Raman spectra by Bobbie-Jo M. Webb-Robertson; Vanessa L. Bailey; Sarah J. Fansler; Michael J. Wilkins; Nancy J. Hess (563-572).
In general, classification-based methods based on confocal Raman microscopy are focused on targeted studies under which the spectral libraries are collected under controlled instrument parameters, which facilitate analyses via standard multivariate data analysis methods and cross-validation. We develop and compare approaches to transform spectra collected at different spectral ranges and varying levels of resolution into a single lower-dimension spectral signature library. This will result in a more robust analysis method able to accommodate spectra accumulated at different times and conditions. We demonstrate these approaches on a relevant test case; the identification of microbial species from a natural environment. The training data were based on samples prepared for three unique species collected at two time points and the test data consisted of blinded unknowns prepared and analyzed at a later date with different instrument parameters. The results indicate that using reduced dimension representations of the spectral signatures improves classification accuracy over basic alignment protocols. In particular, utilizing the microbial species partial least squares discriminant analysis classifier on the blinded samples based on alignment achieved ~78 % accuracy, while both binning and peak selection approaches yielded 100 % accuracy. Figure A probability heatmap associated with the identification of species di181 across 357 spectra collected from a single drop of a mixed microbial suspension, dry-mounted for Raman analysis
Keywords: Confocal Raman microscopy; Classification; Alignment; Binning; Peak selection
The effect of different gloss levels on in-line monitoring of the thickness of printed layers by NIR spectroscopy by Gabriele Mirschel; Olesya Savchuk; Tom Scherzer; Beatrix Genest (573-583).
Near-infrared (NIR) reflection spectroscopy was used for monitoring the thickness or rather the coating weight of thin printed layers of transparent oil-based offset printing varnishes in a range from 0.5 to 5 g m−2. Quantitative analysis of the spectral data was carried out with partial least squares regression. Surface properties such as the gloss were found to strongly affect the prediction of the coating weight. This influence was minimized by the development of calibration models, which contained spectra of layers with a broad range of gloss levels. The prediction error of these models was in the order of 0.12 to 0.16 g m−2. In-line measurements were carried out at a sheet-fed offset printing press in order to test the performance of the models under real process control conditions. Varnishes were applied to paper at printing speeds of 90 or 180 m min−1. A close correlation between the predictions from in-line NIR spectra and the reference data from gravimetry was observed regardless of the specific degree of gloss of the layers (errors between 0.15 and 0.17 g m−2). The results clearly prove the efficiency of NIR reflection spectroscopy for quantitative investigations on thin layers in fast processes such as printing and demonstrate its analytical potential for quality and process control. Figure In-line monitoring of the coating weight of printed layers of an oil-based varnish by NIR reflection spectroscopy.
Keywords: NIR reflection spectroscopy; Partial least squares (PLS) regression; In-line monitoring; Process control; Printing; Degree of gloss
Selective light-triggered chemiluminescence between fluorescent dyes and luminol, and its analytical application by Mingyang Ma; Fangning Diao; Xingwang Zheng; Zhihui Guo (585-592).
We report herein a novel chemiluminescence (CL) phenomenon triggered by light irradiation when a fluorescent dye, for example hematoporphyrin, fluorescein, eosin, or methylene blue is present in the luminol solution. A possible mechanism is proposed for the photoinduced chemiluminescence (PICL) reaction. Compared with reported methods for CL triggering, for example flow-injection, static reagent injection, and the electrochemical technique, the proposed in-situ PICL method presented has three advantages. First, the method is more selective, because the PICL signal of the target fluorescent dyes is initiated by excitation at a selective wavelength only. Second, the space and time resolution of the PICL method are better. Last, and most important, compared with injecting a reagent or inserting a electrode into the CL system to initiate the CL reaction, with the in-situ PICL method there is no physical interference with the target detecting system. All these advantages of the PICL method indicate it has many potential applications in the analytical sciences. The proposed method was applied to analysis of urine containing adrenaline. The linear range for adrenaline is 2.0 × 10−10–1.0 × 10−7 g mL−1 and the detection limit is 6.0 × 10−11 g mL−1. Figure Schematic PICL Mechanism for the CL reagents-fluorescence dyes systems
Keywords: Chemiluminescence; Photoinduced; Fluorescent dyes; ROSs; Adrenaline
NMR metabolomics for assessment of exercise effects with mouse biofluids by Laurence Le Moyec; Laurence Mille-Hamard; Mohamed N. Triba; Carole Breuneval; Hélène Petot; Véronique L. Billat (593-602).
Exercise modulates the metabolome in urine or blood as demonstrated previously for humans and animal models. Using nuclear magnetic resonance (NMR) metabolomics, the present study compares the metabolic consequences of an exhaustive exercise at peak velocity (Vp) and at critical velocity (Vc) on mice. Since small-volume samples (blood and urine) were collected, dilution was necessary to acquire NMR spectra. Consequently, specific processing methods were applied before statistical analysis. According to the type of exercise (control group, Vp group and Vc group), 26 male mice were divided into three groups. Mice were sacrificed 2 h after the end of exercise, and urine and blood samples were drawn from each mouse. Proton NMR spectra were acquired with urine and deproteinized blood. The NMR data were aligned with the icoshift method and normalised using the probabilistic quotient method. Finally, data were analysed with the orthogonal projection of latent-structure analysis. The spectra obtained with deproteinized blood can neither discriminate the control mice from exercised mice nor discriminate according to the duration of the exercise. With urine samples, a significant statistical model can be estimated when comparing the control mice to both groups, Vc and Vp. The best model is obtained according to the exercise duration with all mice. Taking into account the spectral regions having the highest correlations, the discriminant metabolites are allantoin, inosine and branched-chain amino acids. In conclusion, metabolomic profiles assessed with NMR are highly dependent on the exercise. These results show that urine samples are more informative than blood samples and that the duration of the exercise is a more important parameter to influence the metabolomic status than the exercise velocity.
Keywords: NMR; Mouse; Exercise; Urine; Blood; Exhaustion; Peak velocity; Critical velocity
Rapid determination of pH in solid-state fermentation of wheat straw by FT-NIR spectroscopy and efficient wavelengths selection by Hui Jiang; Guohai Liu; Congli Mei; Shuang Yu; Xiahong Xiao; Yuhan Ding (603-611).
In the work discussed in this paper we investigated the feasibility of determination of the pH of a fermented substrate in solid-state fermentation (SSF) of wheat straw. Fourier-transform near-infrared (FT-NIR) spectroscopy was combined with an appropriate multivariate method of analysis. A genetic algorithm and synergy interval partial least-squares (GA–siPLS) were used to select the efficient spectral subintervals and wavelengths by k-fold cross-validation during development of the model. The performance of the final model was evaluated by use of the root mean square error of cross-validation (RMSECV) and correlation coefficient (R c) for the calibration set, and verified by use of the root mean square error of prediction (RMSEP) and correlation coefficient (R p) for the validation set. The experimental results showed that the optimum GA-siPLS model was achieved by use of seven PLS factors, when four spectral subintervals were selected by siPLS and then 45 wavelength variables were chosen by use of the GA. The predicted precision of the best model obtained was: RMSECV = 0.0583, R c = 0.9878, RMSEP = 0.0779, and R p = 0.9779. Finally, the superior performance of the GA-siPLS model was demonstrated by comparison with four other PLS models. The overall results indicated that FT-NIR spectroscopy can be successfully used for measurement of pH in solid-state fermentation, and use of the GA-siPLS algorithm is the best means of calibration of the model. Figure The above figure shows the algorithm implementation process in this study. siPLS combined with a genetic algorithm (GA), called GA-siPLS, was used for model calibration. The specific objectives of the study were: to preprocess the raw spectra by use of the first derivative; to split the full-spectrum region into equidistant spectral subintervals and to select several efficient subintervals by use of siPLS; to select useful wavelength variables by use of a GA from the efficient subintervals selected; and to develop a PLS model based on the useful variables selected by GA-siPLS
Keywords: Fourier-transform near-infrared spectroscopy; Synergy interval partial least-squares; Genetic algorithm; Solid-state fermentation; pH
Letter to the editor regarding “Determination of glyphosate in groundwater samples using an ultrasensitive immunoassay and confirmation by on-line solid phase extraction followed by liquid chromatography coupled to tandem mass spectrometry” by Marie-Anne Reding (613-614).
Response to Letter to the Editor regarding “Determination of glyphosate in groundwater samples using an ultrasensitive immunoassay and confirmation by on-line solid phase extraction followed by liquid chromatography coupled to tandem mass spectrometry” by Josep Sanchís; Lina Kantiani; Marta Llorca; Fernando Rubio; Antoni Ginebreda; Josep Fraile; Teresa Garrido; Marinella Farré (615-616).
Erratum to: Determination of glyphosate in groundwater samples using an ultrasensitive immunoassay and confirmation by on-line solid-phase extraction followed by liquid chromatography coupled to tandem mass spectrometry by Josep Sanchís; Lina Kantiani; Marta Llorca; Fernando Rubio; Antoni Ginebreda; Josep Fraile; Teresa Garrido; Marinella Farré (617-617).
Erratum to: 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 (619-621).
Erratum to: Quantification of nanoparticles in aqueous food matrices using Particle-Induced X-ray Emission by Omar Lozano; Jorge Mejia; Tijani Tabarrant; Bernard Masereel; Jean-Michel Dogné; Olivier Toussaint; Stéphane Lucas (623-623).