Analytical and Bioanalytical Chemistry (v.407, #22)

ABC Spotlight on emerging microRNA analysis methods by Adam T. Woolley (6579-6581).
is Professor and Associate Chair in the Department of Chemistry and Biochemistry at Brigham Young University in Provo, Utah, USA. He is the recipient of a Presidential Early Career Award for Scientists and Engineers (2006) and the ACS Division of Analytical Chemistry Award for Young Investigators in Separation Science (2007). His current research focuses on three general topics: the development of novel and sophisticated integrated microfluidic systems for preterm birth biomarker quantitation, the design of simple miniaturized biomolecular assays, and biotemplated fabrication of nanoelectronic systems.

Elise Dennis wins ABC Best Paper Award by Nicola Oberbeckmann-Winter (6583-6585).
born 1988, is currently a PhD candidate in analytical chemistry at Indiana University under the direction of Professor Gary M. Hieftje. She has authored/co-authored eight peer-reviewed publications, is an inventor on two patents, and has presented two invited lectures and eight additional talks/posters at various scientific conferences. Prior to graduate school, Elise received her BS with honors from the College of William and Mary, where she performed research in the laboratory of Professor John C. Poutsma, and presented a poster on her research at a regional meeting of the American Chemical Society. In addition to completion of an honors BS thesis, Elise has received several recognitions for her academic accomplishments, including the 2014 Barbara Stull Graduate Student Award from the Society of Applied Spectroscopy, a Robert and Marjorie Mann Chair Fellowship, an REU summer research fellowship sponsored by NSF, a Howard Hughes Medical Institute Freshman Research Fellowship, a Metacyt Fellowship, an Indiana University Women in Chemistry Travel Award, and an Indiana University Provost Travel Award. Apart from her research duties, Elise also serves as the current treasurer of the Indiana Section of the Society of Applied Spectroscopy.

Rubbery egg challenge by Hervé This (6587-6588).

Solution to highest melting point challenge by John Arblaster (6589-6590).

ANAKON 2015 in Graz: a warm welcome for analytical chemists in the heart of Styria by Daniel Sydes; Patricia Weber; Katrin Krieg; Julia Widmaier (6591-6592).

Recent advances in quantitative LA-ICP-MS analysis: challenges and solutions in the life sciences and environmental chemistry by Andreas Limbeck; Patrick Galler; Maximilian Bonta; Gerald Bauer; Winfried Nischkauer; Frank Vanhaecke (6593-6617).
Laser ablation–inductively coupled plasma–mass spectrometry (LA-ICP-MS) is a widely accepted method for direct sampling of solid materials for trace elemental analysis. The number of reported applications is high and the application range is broad; besides geochemistry, LA-ICP-MS is mostly used in environmental chemistry and the life sciences. This review focuses on the application of LA-ICP-MS for quantification of trace elements in environmental, biological, and medical samples. The fundamental problems of LA-ICP-MS, such as sample-dependent ablation behavior and elemental fractionation, can be even more pronounced in environmental and life science applications as a result of the large variety of sample types and conditions. Besides variations in composition, the range of available sample states is highly diverse, including powders (e.g., soil samples, fly ash), hard tissues (e.g., bones, teeth), soft tissues (e.g., plants, tissue thin-cuts), or liquid samples (e.g., whole blood). Within this article, quantification approaches that have been proposed in the past are critically discussed and compared regarding the results obtained in the applications described. Although a large variety of sample types is discussed within this article, the quantification approaches used are similar for many analytical questions and have only been adapted to the specific questions. Nevertheless, none of them has proven to be a universally applicable method.
Keywords: LA-ICP-MS; Quantitative analysis; Certified reference material; Matrix-matched standards; Internal standard correction; Liquid standards

Micro-distribution of uranium in bone after contamination: new insight into its mechanism of accumulation into bone tissue by Damien Bourgeois; Brigitte Burt-Pichat; Xavier Le Goff; Jan Garrevoet; Pieter Tack; Gerald Falkenberg; Luc Van Hoorebeke; Laszlo Vincze; Melissa A. Denecke; Daniel Meyer; Claude Vidaud; Georges Boivin (6619-6625).
After internal contamination, uranium rapidly distributes in the body; up to 20 % of the initial dose is retained in the skeleton, where it remains for years. Several studies suggest that uranium has a deleterious effect on the bone cell system, but little is known regarding the mechanisms leading to accumulation of uranium in bone tissue. We have performed synchrotron radiation-based micro-X-ray fluorescence (SR μ-XRF) studies to assess the initial distribution of uranium within cortical and trabecular bones in contaminated rats’ femurs at the micrometer scale. This sensitive technique with high spatial resolution is the only method available that can be successfully applied, given the small amount of uranium in bone tissue. Uranium was found preferentially located in calcifying zones in exposed rats and rapidly accumulates in the endosteal and periosteal area of femoral metaphyses, in calcifying cartilage and in recently formed bone tissue along trabecular bone. Furthermore, specific localized areas with high accumulation of uranium were observed in regions identified as micro-vessels and on bone trabeculae. These observations are of high importance in the study of the accumulation of uranium in bone tissue, as the generally proposed passive chemical sorption on the surface of the inorganic part (apatite) of bone tissue cannot account for these results. Our study opens original perspectives in the field of exogenous metal bio-mineralization.
Keywords: Uranium; Bone tissue; Micro-X-ray fluorescence analysis (μ-XRF); Elemental mapping; Bio-mineralization

recently obtained his Masters from North Carolina State University, where he worked on the development of LC–MS–MS methods for detecting environmental exposures in biological fluids. is an Assistant Professor in Biological Sciences at NC State University and Member of the Center for Human Health and Environment. His research is centered on the development and applications of new analytical technology to identify biomarkers of exposures in ultimate efforts to better understand how the environment affects human health. We present a novel strategy based on data-independent acquisition coupled to targeted data extraction for the detection and identification of site-specific modifications of targeted peptides in a completely unbiased manner. This method requires prior knowledge of the site of the modification along the peptide backbone from the protein of interest, but not the mass of the modification. The procedure, named multiplex adduct peptide profiling (MAPP), consists of three steps: 1) A fragment-ion tag is extracted from the data, consisting of the b-type and y-type ion series from the N and C-terminus, respectively, up to the amino-acid position that is believed to be modified; 2) MS1 features are matched to the fragment-ion tag in retention-time space, using the isolation window as a pre-filter to enable calculation of the mass of the modification; and 3) modified fragment ions are overlaid with the unmodified fragment ions to verify the mass calculated in step 2. We discuss the development, applications, and limitations of this new method for detection of unknown peptide modifications. We present an application of the method in profiling adducted peptides derived from abundant proteins in biological fluids with the ultimate objective of detecting biomarkers of exposure to reactive species.
Keywords: Protein adducts; Toxicoproteomics; Mass spectrometry; Data-independent acquisition; Exposome

Structure characterization of unexpected covalent O-sulfonation and ion-pairing on an extremely hydrophilic peptide with CE-MS and FT-ICR-MS by Martin Pattky; Simone Nicolardi; Beatrix Santiago-Schübel; Daniel Sydes; Yuri E. M. van der Burgt; Antonia N. Klein; Nan Jiang; Jeannine Mohrlüder; Karen Hänel; Janine Kutzsche; S. A. Funke; D. Willbold; S. Willbold; C. Huhn (6637-6655).
is a Postdoctoral Researcher at the Eberhard Karls Universität Tübingen, Germany. His work focuses on the development and optimization of CE-MS methods for bioanalytical questions in pharmacy, medicine, and environmental analysis. is Postdoctoral Researcher at the Center for Proteomics and Metabolomics of the Leiden University Medical Center in the Netherlands. His work focuses on the development of ultrahigh resolution mass spectrometry-based methods for the analysis of body fluid-derived peptides and proteins in a clinical setting. is a PhD student in analytical chemistry in the working group of Professor Carolin Huhn at the Eberhard Karls Universität Tübingen, Germany. He studied BSc “Water Science” in Duisburg/Essen, Germany, and graduated with the degree of MSc. in analytical chemistry, at the Leibniz Universität Hannover, Germany. He is working on instrumental and method development of multidimensional electrophoretic separation platforms, with a special regard to the coupling of 2D CIEF/CE-MS for protein analysis. obtained his PhD in bio-organic chemistry at Utrecht University in the Netherlands. He works as an Associate Professor at the Leiden University Medical Center and is responsible for multiple MS-platforms that are applied both for in-depth analyses and high-throughput clinical proteomics studies. is a Scientist at the Institute of Complex Systems, Structural Biochemistry (ICS-6) at the Research Center Jülich, Germany. She obtained a diploma in biosystems engineering in 2012 from the Otto-von-Guericke University Magdeburg. Her research focuses on the development and characterization of d-peptides for the treatment of Alzheimer’s disease. Currently, she is carrying out her PhD in biology at the University of Düsseldorf. is Research Assistant at the Institute of Complex Systems (ICS-6) in Forschungszentrum Jülich, Germany. He is working on preclinical pharmacokinetic, therapeutic, and diagnostic studies of Aβ-aggregation modulating D-peptides on mouse models for the treatment of Alzheimer’s disease. is a Scientist at the Research Centre in Jülich, Germany. Her current work focuses on the identification and characterization of protein–protein interactions. She majored in Biology at Gutenberg University of Mainz, Germany, at University of Montpellier II, Faculté des sciences, France, and at University of Cologne, Germany, where she passed her diploma thesis in biochemistry. She obtained a PhD from the Heinrich-Heine University of Düsseldorf, Germany. is Group Leader at the Institute of Complex Systems–Structural Biochemistry (ICS-6), Forschungszentrum Jülich, Germany. She is working on the development of Aβ-aggregation modulating d-peptides for the therapy of Alzheimer’s disease. is Professor for molecular biology at the University of Applied Sciences in Coburg, Germany. Her research focuses on the investigation of protein–protein interactions, especially in the fields of neurodegenerative diseases, diagnostics, and food technology. is Full Professor for physical biology at the Heinrich Heine University, Düsseldorf, and Director of the Institute of Complex Systems at the Research Centre Jülich. He uses state-of-the-art techniques to decipher the complex three-dimensional structures of proteins and their interactions. This provides the basis for an understanding of how these proteins work, thus contributing to the development of methods for early diagnosis and treatment of neurodegenerative diseases. is Professor for Effect-based Environmental Analysis at the Eberhard Karls Universität Tübingen and has been a speaker of the platform Environmental System Analysis since 2013. Her research interests cover all aspects of analytical chemistry: instrumental development, analytical basic research, method development, and application. Major focuses are electromigrative separation techniques and their application in forensics, pharmacy, clinics, and environmental bioanalysis. In this study, we characterized unexpected side-products in a commercially synthesized peptide with the sequence RPRTRLHTHRNR. This so-called peptide D3 was selected by mirror phage display against low molecular weight amyloid-β-peptide (Aβ) associated with Alzheimer’s disease. Capillary electrophoresis (CE) was the method of choice for structure analysis because the extreme hydrophilicity of the peptide did not allow reversed-phase liquid chromatography (RPLC) and hydrophilic interaction stationary phases (HILIC). CE-MS analysis, applying a strongly acidic background electrolyte and different statically adsorbed capillary coatings, provided fast and efficient analysis and revealed that D3 unexpectedly showed strong ion-pairing with sulfuric acid. Moreover, covalent O-sulfonation at one or two threonine residues was identified as a result of a side reaction during peptide synthesis, and deamidation was found at either the asparagine residue or at the C-terminus. In total, more than 10 different species with different m/z values were observed. Tandem-MS analysis with collision induced dissociation (CID) using a CE-quadrupole-time-of-flight (QTOF) setup predominantly resulted in sulfate losses and did not yield any further characteristic fragment ions at high collision energies. Therefore, direct infusion Fourier transform ion cyclotron resonance (FT-ICR) MS was employed to identify the covalent modification and discriminate O-sulfonation from possible O-phosphorylation by using an accurate mass analysis. Electron transfer dissociation (ETD) was used for the identification of the threonine O-sulfation sites. In this work, it is shown that the combination of CE-MS and FT-ICR-MS with ETD fragmentation was essential for the full characterization of this extremely basic peptide with labile modifications.
Keywords: Extremely hydrophilic peptide; Capillary electrophoresis; Coating; Electron transfer dissociation; Alzheimer’s disease

Electrochemiluminescence DNA sensor array for multiplex detection of biowarfare agents by Anna-Maria Spehar-Délèze; Rainer Gransee; Sergio Martinez-Montequin; Diego Bejarano-Nosas; Samuel Dulay; Sandra Julich; Herbert Tomaso; Ciara K. O’Sullivan (6657-6667).
obtained her PhD from Helsinki University of Technology (now Aalto University) in 2006. Her thesis focused on electrochemiluminescence applications on microfabricated devices and was mainly carried out at the Institute of Microtechnology, Neuchâtel, currently part of the Ecole Polytechnique Fédéral de Lausanne (EPFL), Switzerland. Since then she has worked on novel transduction methods for bioassays in the Biomedical Diagnostic Institute in Dublin, Ireland, developed implantable and wearable sensors for sports and health-care applications within the Elite Sports Performance Research in Training (ESPRIT) program at Queen Mary University of London, and developed portable, sensitive biosensors for biowarfare agents as part of the Multisense chip EU project at the University of Rovira i Virgili in Tarragona, Spain. Her main research interests are electrochemical biosensors for health care, sports, and environmental applications. Currently she is developing her own consulting business. studied mechanical engineering at the Technical University of Stuttgart and received his degree in 2005. He joined the Microfluidic Analysis Systems department at the Institute for Microtechnology in Mainz, focusing on the development of disposable microfluidic total analysis systems for bio-chemical analyses. These tasks include the design and realisation of polymer-based cartridges using rapid prototyping technology (for example milling, laser ablation, or hot embossing) and system integration into automated lab prototypes (“sample in answer out” systems). He has been project leader of several publicly funded research projects and industrial projects with the objective of commercialising microfluidic technology. He has been Leader of the Fluidics Group at Fraunhofer ICT-IMM since 2009. has, since completing his Bachelor Degree in Chemistry at the University of Oviedo (Spain), been involved in several research and development (R&D) projects and in technology transfers. His first experience of R&D was as a research fellow working with immunosensors with electrochemical detection. Later, he obtained his PhD in Electrochemistry at the University of Southampton with a thesis focused on performance improvement of reference electrodes in commercial glucose screen-printed sensors sponsored by MediSense (now Abbott Laboratories, UK). Later, he worked on practical aspects of technology transfer at Nanotecture (UK), developing and commercially exploiting new nanoporous electrodeposited materials for batteries and sensor applications. He has also worked in the food sector of the Research and Development and Innovation (R&D&i) Department of Central Lechera Asturiana (Spain), carrying out pilot plant and production scale trials of new products. He is currently Manufacturing Director at iMiCROQ, but still deals with R&D&i because he is in charge of the development of the screen-printed microsystems. is quality manager of Integrated Microsystems for Quality of Life S.L. in Tarragona, Spain. He has been working for several years as project technician in the INTERFIBIO Research Center (University Rovira i Virgili, Tarragona, Spain) involved in the scientific study of design and improvement of microfabrication techniques, manufacture of screen-printed microsystems, and development of electrochemiluminescent biosensors. is a postdoctoral research technician fellow at the Institute for Bioengineering of Catalonia (IBEC), Barcelona, Spain. He has been working on the development of electrochemical biosensors for the detection of biowarfare agents and/or pathogenic species, including improving surface chemistry for better biosensor performance. is a research associate at the Institute of Bacterial Infections and Zoonoses in Germany. Her work is focused on innovative technology for detection of highly pathogenic bacteria that can be transferred between animals and humans. In particular, she is involved in evaluation studies of microfluidic chip systems. is Deputy Head of the Institute for Bacterial Infections and Zoonoses and the head of the National Reference Laboratory for Tularemia in Germany. The focus of the institute is not only on standard farm animals and their diseases, but also on companion animals, animals kept in zoos, and wildlife. He has been working for several years on the development of assays for the identification and typing of highly pathogenic bacteria. received a BSc in Analytical Chemistry from Dublin City University in 1992 and a joint PhD in Biotechnology at Cranfield University (UK) and University of Ioannina (Greece) in 1996, and then went on to become Research Director of the Guilbault Laboratory at National University of Ireland, Cork (Ireland) from 1996–1999. She then took up a Marie Curie Fellowship at the Universitat Rovira i Virgili (1999–2001), and was then awarded a Ramón y Cajal Fellowship which she pursued for one year before taking up her current position as ICREA Research Professor and establishing the Nanobiotechnology and Bioanalysis Group at the Universitat Rovira i Virgili. Her research interests lie in the development of electrochemical and optical biosensors exploiting advances in tailored biocomponents. Her current work focuses on cost-effective practical applications of molecular diagnostics for screening and monitoring of disease, and on the development of aptamers for application in optical and electrochemical molecular-aptamer beacons. The approaches for molecular diagnostics being developed include parallelised real-time electrochemical next-generation sequencing, electrochemical-array-based primer extension, and quantitative paper diagnostics as companion tools for future pharmacogenomics and personalised medicine. Development of a fully automated electrochemiluminescence (ECL) DNA assay for multiplex detection of six biowarfare agents is described. Aminated-DNA capture probes were covalently immobilised on activated-carbon electrodes and subsequently hybridised to target strands. Detection was achieved via a sandwich-type assay after Ru(bpy)3 2+-labelled reporter probes were hybridised to the formed probe–target complexes. The assay was performed in an automated microsystem in a custom-designed ECL detection box with integrated fluidics, electronics, and movable photomultiplier detector. The obtained limits of detection were 0.6–1.2 nmol L−1 for six targets ranging from 50 to 122 base pairs in size, with linear range 1–15 nmol L−1. Non-specific adsorption and cross-reactivity were very low. Detection of six targets on a single chip was achieved with subnanomolar detection limits. Graphical Abstract A photo of the electrode array containing 3 × 14 carbon working electrodes (WE), a common carbon counter electrode (CE), and Ag/AgCl reference electrode (RE) (top). The detection scheme is indicated for one WE. First, probe sequences were covalently immobilised on the activated-carbon surface, then target strands were introduced, and finally Ru(bpy)3 2+-labelled reporter strands were introduced to complete the sandwich-type hybridisation assay. The 3D graph shows multiplex detection of six different pathogens on a single chip
Keywords: Electrochemiluminescence; DNA biosensor; Biowarfare agent; Screen-printed carbon array; Multiplex detection

An improved atmospheric pressure chemical ionization (APCI II) source for gas chromatography–high-resolution time-of-flight mass spectrometry (GC–HRTOFMS) was compared to its first-generation predecessor for the analysis of fatty acid methyl esters, methoxime-trimethylsilyl derivatives of metabolite standards, and cell culture supernatants. Reductions in gas turbulences and chemical background as well as optimized heating of the APCI II source resulted in narrower peaks and higher repeatability in particular for late-eluting compounds. Further, APCI II yielded a more than fourfold median decrease in lower limits of quantification to 0.002–3.91 μM along with an average 20 % increase in linear range to almost three orders of magnitude with R 2 values above 0.99 for all metabolite standards investigated. This renders the overall performance of GC–APCI–HRTOFMS comparable to that of comprehensive two-dimensional gas chromatography (GC × GC)–electron ionization (EI)–TOFMS. Finally, the number of peaks with signal-to-noise ratios greater than 20 that could be extracted from metabolite fingerprints of pancreatic cancer cell supernatants upon switching from the APCI I to the APCI II source was more than doubled. Concomitantly, the number of identified metabolites increased from 36 to 48. In conclusion, the improved APCI II source makes GC–APCI–HRTOFMS a great alternative to EI-based GC–MS techniques in metabolomics and other fields.
Keywords: GC–APCI–HRTOFMS; Ionization efficiency; Atmospheric pressure chemical ionization; GC–APCI II source; Metabolomics

LC-MS/MS-based analysis of coenzyme A and short-chain acyl-coenzyme A thioesters by Stefan Neubauer; Dinh Binh Chu; Hans Marx; Michael Sauer; Stephan Hann; Gunda Koellensperger (6681-6688).
Absolute quantification of intracellular coenzyme A (CoA), coenzyme A disulfide, and short-chain acyl-coenzyme A thioesters was addressed by developing a tailored metabolite profiling method based on liquid chromatography in combination with tandem mass spectrometric detection (LC-MS/MS). A reversed phase chromatographic separation was established which is capable of separating a broad spectrum of CoA, its corresponding derivatives, and their isomers despite the fact that no ion-pairing reagent was used (which was considered as a key advantage of the method). Excellent analytical figures of merit such as high sensitivity (LODs in the nM to sub-nM range) and high repeatability (routinely 4 %; N = 15) were obtained. Method validation comprised a study on standard purity, stability, and recoveries during sample preparation. Uniformly labeled U13C yeast cell extracts offered ideal internal standards for validation purposes and for a quantification exercise in the rumen bacterium Megasphaera elsdenii.
Keywords: Coenzyme A; Short-chain acyl-coenzyme A; LC-MS/MS; Metabolite profiling; Megasphaera elsdenii ; Stability

Time- and spatially resolved emission spectroscopy of the dielectric barrier discharge for soft ionization sustained by a quasi-sinusoidal high voltage by Vlasta Horvatic; Antje Michels; Norman Ahlmann; Günter Jestel; Damir Veza; Cedomil Vadla; Joachim Franzke (6689-6696).
A helium capillary dielectric barrier discharge was investigated by means of time-resolved optical emission spectroscopy with the aim of elucidating the process of the formation of the plasma jet. The helium emission line at 706 nm was utilized to monitor spatial and temporal propagation of the excitation of helium atoms. The discharge was sustained with quasi-sinusoidal high voltage, and the temporal evolution of the helium atomic emission was measured simultaneously with the discharge current. The spatial development of the plasma was investigated along the discharge axis in the whole region, which covers the positions in the capillary between the electrodes as well as the plasma jet outside the capillary. The high voltage electrode was placed 2 mm from the capillary orifice, and the distance between the ground and high voltage electrode was 10 mm. The complete spatiotemporal grid of the development of the helium excitation has shown that during the positive half-period of the applied voltage, two independent plasmas, separated in time, are formed. First, the early plasma that constitutes the plasma jet is formed, while the discharge in the capillary follows subsequently. In the early plasma, the helium atom excitation propagation starts in the vicinity of the high voltage electrode and departs from the capillary towards the ground electrode as well as several millimeters outside of the capillary in the form of the plasma jet. After relatively slow propagation of the early plasma in the capillary and the jet, the second plasma starts between the electrodes. During the negative voltage period, only the plasma in the capillary between the electrodes occurs. Graphical Abstract Spatiotemporal evolution of the helium excitation propagation in the He capillary DBD
Keywords: Dielectric-barrier-discharge ionization; Soft ionization; Time-resolved emission spectroscopy

25I-NBOMe, a new psychoactive substance, is a potent 5-HT2A receptor agonist with strong hallucinogenic potential. Recently, it was involved in several fatal and non-fatal intoxication cases. The aim of the present work was to study its phase I and II metabolism and its detectability in urine screening approaches. After application of 25I-NBOMe to male Wistar rats, urine was collected over 24 h. The phase I and II metabolites were identified by LC-HR-MS/MS in urine after suitable workup. For the detectability studies, standard urine screening approaches (SUSA) by GC-MS, LC-MSn, and LC-HR-MS/MS were applied to rat and also to authentic human urine samples submitted for toxicological analysis. Finally, an initial CYP activity screening was performed to identify CYP isoenzymes involved in the major metabolic steps. 25I-NBOMe was mainly metabolized by O-demethylation, O,O-bis-demethylation, hydroxylation, and combinations of these reactions as well as by glucuronidation and sulfation of the main phase I metabolites. All in all, 68 metabolites could be identified. Intake of 25I-NBOMe was detectable mainly via its metabolites by both LC-MS approaches, but not by the GC-MS SUSA. Initial CYP activity screening revealed the involvement of CYP1A2 and CYP3A4 in hydroxylation and CYP2C9 and CYP2C19 in O-demethylation. The presented study demonstrated that 25I-NBOMe was extensively metabolized and could be detected only by the LC-MS screening approaches. Since CYP2C9 and CYP3A4 are involved in initial metabolic steps, drug–drug interactions might occur in certain constellations.
Keywords: Designer drugs; 25I-NBOMe; Metabolism; Cytochrome-P450; LC-MS n ; LC-HR-MS/MS

Quantification and kinetic study in plasma and tissues of (E)-1,1,4,4-tetramethyl-2-tetrazene, a liquid propellant and a transformation product of 1,1-dimethyl hydrazine by Léa Payen; Mylène Honorat; Charlotte Bouard; Guy Jacob; Raphael Terreux; Henri Delalu; Emilie Labarthe; Jérôme Guitton (6721-6729).
(E)-1,1,4,4-tetramethyl-2-tetrazene (TMTZ) is formed from the oxidation of the unsymmetrical 1,1-dimethylhydrazine (UDMH) and is used as a storable liquid fuel which can be considered as a new potential propellant for space rocket propulsion. To better understand the toxicological behavior of the compound, an intraperitoneal administration of TMTZ was performed in mice to define its toxicokinetics and tissue distribution. A fully validated liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) assay was developed to determine TMTZ levels in biological samples. Determination of TMTZ was achieved using 50 μL of plasma or tissue solution. Precipitation with ammonium sulfate and acetonitrile was used for sample preparation. Liquid chromatography was performed on an Atlantis HILIC Silica column (Waters; 3 μm, 150 mm × 2.1 mm i.d.). Isocratic elution with a mixture of ammonium acetate buffer (pH 5, 100 mM)/water/acetonitrile (3:2:95, v/v/v) was used. The detection was conducted using an electrospray source in positive ion mode. TMTZ and 15N2-TMTZ (internal standard) were quantitated in selected reaction monitoring mode using the transition m/z 117→72 and 119→74, respectively. Standard curves exhibited excellent linearity in the range of 10–500 ng/mL for plasma and 50–2000 ng/mL for all tissues (heart, liver, brain, kidney, and lung) analyzed, and acceptable precision and accuracy (<10 %) were obtained. The elimination rate constant strongly suggests that TMTZ was very quickly eliminated from the body. The results of tissue distribution experiments indicated that TMTZ underwent a rapid distribution into limited organs such as the liver, kidney, and brain.
Keywords: TMTZ; Alkyltetrazene; Liquid propellant; Tissues distribution; Liquid chromatography–mass spectrometry

In 2009, the genetically modified (GM) rice event TT51-1 with an engineered insect resistance trait became the first GM rice event to be granted certification for safe production in China, and its derivative lines Bt 63 and Huahui No.1 are expected to be commercialized soon. The development of certified reference material (CRM) for TT51-1 is necessary to monitor and inspect the TT51-1 event and its derivates. In this work, we developed four matrix-based TT51-1 rice CRMs (TT51-1a, TT51-1b, TT51-1c, and TT51-1d) with different TT51-1 mass fraction ratios by blending seed powders of homozygous TT51-1 and its recipient cultivar Minghui 63. The between-bottle homogeneity and the within-bottle homogeneity were tested, and good results were obtained. The potential degradation during transportation and shelf life were evaluated, and demonstrated an expiration period of at least 36 months. The characterization values of the four TT51-1 CRMs based on the mass fraction ratio were 1000.000 ± 51.430 g/kg, 49.940 ± 4.620 g/kg, 9.990 ± 1.110 g/kg, and 4.990 ± 0.620 g/kg, respectively. The characterization values based on the copy number ratio were certified by digital PCR analysis as 97.442 ± 5.253 %, 4.851 ± 0.486 %, 1.042 ± 0.135 %, and 0.556 ± 0.073 %, respectively. These results suggested that the TT51-1 matrix-based CRMs developed are of high quality and can be used as potential calibrators for TT51-1 GM rice inspection and monitoring. Graphical Abstract The general procedure for the development of TT51-1 rice certified reference materials
Keywords: Genetically modified rice; TT51-1; Certified reference materials; Characterization

A label-free and enzyme-free demultiplexer system for the fabrication of 1:2 molecular demultiplexer with luminol functionalized gold nanoparticles (Lum-AuNPs) as signal transducers was developed for the first time. The Lum-AuNPs had both chemiluminescence (CL) activity and surface plasmon resonance property. It was found that organothiols (RSH) could easily induce the aggregation of AuNPs via strong Au–S covalent interactions in the absence of hydrogen peroxide (H2O2), generating a red shift in the absorption band of AuNPs. However, the presence of H2O2 would readily oxidize RSH to disulfide (RS-SR), and the aggregation of Lum-AuNPs did not occur due to lack of the sulfhydryl group. Meanwhile, H2O2 could react with Lum-AuNPs, producing a strong CL emission owing to the enhancement effect of RSH on AuNPs-luminol-H2O2 CL system. Thus, RSH, H2O2, absorbance ratio, and CL intensity served as the signal input, address input, and two different signal outputs of the 1:2 molecular demultiplexer, respectively.
Keywords: Chemiluminescence; Gold nanoparticles; Molecular demultiplexer

Molecular imprinting is an emerging technique to create imprinted polymers that can be applied in affinity-based separation, in particular, biomimetic sensors. In this study, the matrix of siloxane bonds prepared from the polycondensation of hydrolyzed tetraethoxysilane (TEOS) was employed as the inorganic monomer for the formation of a creatinine (Cre)-based molecularly imprinted polymer (MIP). Doped aluminium ion (Al3+) was used as the functional cross-linker that generated Lewis acid sites in the confined silica matrix to interact with Cre via sharing of lone pair electrons. Surface morphologies and pore characteristics of the synthesized MIP were determined by field emission scanning electron microscopy (FESEM) and Brunauer-Emmet-Teller (BET) analyses, respectively. The imprinting efficiency of MIPs was then evaluated through the adsorption of Cre with regard to molar ratios of Al3+. A Cre adsorption capacity of up to 17.40 mg Cre g–1 MIP was obtained and adsorption selectivity of Cre to its analogues creatine (Cr) and N-hydroxysuccinimide (N-hyd) were found to be 3.90 ± 0.61 and 4.17 ± 3.09, respectively. Of all the studied MIP systems, chemisorption was predicted as the rate-limiting step in the binding of Cre. The pseudo-second-order chemical reaction kinetic provides the best correlation of the experimental data. Furthermore, the equilibrium adsorption capacity of MIP fit well with a Freundlich isotherm (R 2 = 0.98) in which the heterogeneous surface was defined. Graphical Abstract Affinity binding of Cre to specific recognition sites based on shape factor
Keywords: Molecularly imprinted polymer; Creatinine; Sol-gel; Isotherm; Shape recognition; Chemisorption

Interlaboratory study of novel halogenated flame retardants: INTERFLAB by Lisa Melymuk; Emma Goosey; Nicole Riddell; Miriam L. Diamond (6759-6769).
Flame retardants (FRs) have come under considerable scientific and public scrutiny over the past decade. A lack of reference materials and standardized analytical methods has resulted in questions regarding the variation of measurements from different studies. We evaluated this variation by performing an international interlaboratory study assessing analytical capabilities as well as the accuracy and precision of results for a range of flame retardants (International Flame Retardant Laboratory Study, INTERFLAB). Thirteen international research laboratories participated in a blind interlaboratory comparison of 24 FRs. Results demonstrate good precision within replicates of test mixtures from individual laboratories, but problematic accuracy for several FRs and laboratories. Large ranges in the values reported for decabromodiphenylethane (DBDPE), tris(1,3-dichloropropyl)phosphate (TDCIPP), tetrabromobisphenol-A (TBBPA), and hexabromocyclododecane (HBCD) (>50 % relative standard deviations among measured values) and large deviations from the reference values (>25 % bias in accuracy) suggest potential problems for comparability of results. DBDPE, HBCD, and TBBPA had significantly poorer accuracy and precision, suggesting that current analytical methods are not providing reliable results for these FRs.
Keywords: Interlaboratory comparison; Brominated flame retardants; Novel flame retardants; GC-MS; LC-MS

Magnetic resonance spectroscopy and imaging on fresh human brain tumor biopsies at microscopic resolution by M. Carmen Martínez-Bisbal; Beatriz Martínez-Granados; Vicente Rovira; Bernardo Celda; Vicent Esteve (6771-6780).
The metabolic composition and concentration knowledge provided by magnetic resonance spectroscopy (MRS) liquid and high-resolution magic angle spinning spectroscopy (HR-MAS) has a relevant impact in clinical practice during magnetic resonance imaging (MRI) monitoring of human tumors. In addition, the combination of morphological and chemical information by MRI and MRS has been particularly useful for diagnosis and prognosis of tumor evolution. MRI spatial resolution reachable in human beings is limited for safety reasons and the demanding necessary conditions are only applicable on experimental model animals. Nevertheless, MRS and MRI can be performed on human biopsies at high spatial resolution, enough to allow a direct correlation between the chemical information and the histological features observed in such biopsies. Although HR-MAS is nowadays a well-established technique for spectroscopic analysis of tumor biopsies, with this approach just a mean metabolic profile of the whole sample can be obtained and thus the high histological heterogeneity of some important tumors is mostly neglected. The value of metabolic HR-MAS data strongly depends on a wide statistical analysis and usually the microanatomical rationale for the correlation between histology and spectroscopy is lost. We present here a different approach for the combined use of MRI and MRS on fresh human brain tumor biopsies with native contrast. This approach has been designed to achieve high spatial (18 × 18 × 50 μm) and spectral (0.031 μL) resolution in order to obtain as much spatially detailed morphological and metabolical information as possible without any previous treatment that can alter the sample. The preservation of native tissue conditions can provide information that can be translated to in vivo studies and additionally opens the possibility of performing other techniques to obtain complementary information from the same sample.
Keywords: Magnetic resonance imaging; Magnetic resonance spectroscopy; Magnetic resonance microscopy; High-resolution magic angle spinning spectroscopy (HR-MAS); Biopsy; Human brain tumor; Fresh tissue; Glioblastoma

Biodegradable packagings are made by combination of several materials creating a multilayer with the properties needed. Each material, including the adhesive, could contain substances that could migrate to the food. In this work, gas chromatography coupled with mass spectrometry and ultra-high-pressure liquid chromatography coupled with quadrupole time-of-flight mass spectrometry were used to identify the biodegradable adhesive compounds. Five of the 13 compounds identified were nonintentionally added substances; they were neoformed compounds created by the reaction of added compounds in the adhesive. Moreover, the migration of the compounds through four different biodegradable materials—paper, polylactic acid, ecovio®, and polyvinyl alcohol—was studied for the first time. Three of the 13 compounds identified in the adhesive migrated from the adhesive to Tenax®, which was used as a solid food simulant. One of them, 2,4,7,9-tetramethyl-5-decyne-4,7-diol, was an intentionally added substance, and the other two were 1,6-dioxacyclododecane-7,12-dione and 1,6,13,18-tetraoxacyclotetracosane-7,12,19,24-tetraone, which were nonintentionally added substances identified in this work. Higher migration values (ranging from 0.81 to 2.07 mg/kg) were observed for migration through ecovio® than through the multilayer made by combination of ecovio® and polyvinyl alcohol (0.07–0.39 mg/kg) owing to the barrier effect provided by polyvinyl alcohol. The migration values for migration through paper and polylactic acid were below the limits of detection.
Keywords: Ultra-high-pressure liquid chromatography coupled with quadrupole time-of-flight mass spectrometry; Biodegradable packaging; Nonintentionally added substances; Migration

Identification of microplastics by FTIR and Raman microscopy: a novel silicon filter substrate opens the important spectral range below 1300 cm−1 for FTIR transmission measurements by Andrea Käppler; Frank Windrich; Martin G. J. Löder; Mikhail Malanin; Dieter Fischer; Matthias Labrenz; Klaus-Jochen Eichhorn; Brigitte Voit (6791-6801).
The presence of microplastics in aquatic ecosystems is a topical problem and leads to the need of appropriate and reliable analytical methods to distinctly identify and to quantify these particles in environmental samples. As an example transmission, Fourier transform infrared (FTIR) imaging can be used to analyze samples directly on filters without any visual presorting, when the environmental sample was afore extracted, purified, and filtered. However, this analytical approach is strongly restricted by the limited IR transparency of conventional filter materials. Within this study, we describe a novel silicon (Si) filter substrate produced by photolithographic microstructuring, which guarantees sufficient transparency for the broad mid-infrared region of 4000–600 cm-1. This filter type features holes with a diameter of 10 μm and exhibits adequate mechanical stability. Furthermore, it will be shown that our Si filter substrate allows a distinct identification of the most common microplastics, polyethylene (PE), and polypropylene (PP), in the characteristic fingerprint region (1400–600 cm-1). Moreover, using the Si filter substrate, a differentiation of microparticles of polyesters having quite similar chemical structure, like polyethylene terephthalate (PET) and polybutylene terephthalate (PBT), is now possible, which facilitates a visualization of their distribution within a microplastic sample by FTIR imaging. Finally, this Si filter can also be used as substrate for Raman microscopy—a second complementary spectroscopic technique—to identify microplastic samples. Graphical Abstract Optical and FTIR images of a microplastic model sample of PET and PBT on the novel Si filter substrate. The distribution of this quite similar polymers within a microplastic sample is visible by choosing a band region of 1060–1033 cm-1 for PET and of 955–925 cm-1 for PBT
Keywords: Filter substrate; Microplastic identification; FTIR imaging; Raman; Silicon filter

Raman spectroscopic differentiation of planktonic bacteria and biofilms by Dragana Kusić; Bernd Kampe; Anuradha Ramoji; Ute Neugebauer; Petra Rösch; Jürgen Popp (6803-6813).
Both biofilm formations as well as planktonic cells of water bacteria such as diverse species of the Legionella genus as well as Pseudomonas aeruginosa, Klebsiella pneumoniae, and Escherichia coli were examined in detail by Raman microspectroscopy. Production of various molecules involved in biofilm formation of tested species in nutrient-deficient media such as tap water was observed and was particularly evident in the biofilms formed by six Legionella species. Biofilms of selected species of the Legionella genus differ significantly from the planktonic cells of the same organisms in their lipid amount. Also, all Legionella species have formed biofilms that differ significantly from the biofilms of the other tested genera in the amount of lipids they produced. We believe that the significant increase in the synthesis of this molecular species may be associated with the ability of Legionella species to form biofilms. In addition, a combination of Raman microspectroscopy with chemometric approaches can distinguish between both planktonic form and biofilms of diverse bacteria and could be used to identify samples which were unknown to the identification model. Our results provide valuable data for the development of fast and reliable analytic methods based on Raman microspectroscopy, which can be applied to the analysis of tap water-adapted microorganisms without any cultivation step. Graphical abstract Biofilm and planktonic forms of L. pneumophila ssp. pneumophila exhibit different Raman spectra. L. pneumophila ssp. pneumophila in biofilms display a significant increase in the synthesis of lipids compared to the planktonic state
Keywords: Raman spectroscopy; Biofilm; Legionella species; Tap water

Quantitative bile acid profiling by liquid chromatography quadrupole time-of-flight mass spectrometry: monitoring hepatitis B therapy by a novel Na+-taurocholate cotransporting polypeptide inhibitor by Mathias Haag; Ute Hofmann; Thomas E. Mürdter; Georg Heinkele; Patrick Leuthold; Antje Blank; Walter E. Haefeli; Alexander Alexandrov; Stephan Urban; Matthias Schwab (6815-6825).
A novel analytical approach for the targeted profiling of bile acids (BAs) in human serum/plasma based on liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) is presented. Reversed-phase chromatography enabled the baseline separation of 15 human BA species which could be readily detected by accurate mass analysis in negative ion mode. Blood proteins were removed by methanol precipitation in the presence of deuterium-labeled internal standards which allowed BA quantification in 50 μl plasma/serum. The assay was validated according to FDA guidance achieving quantification limits from 7.8 to 156 nM. Calibration curves prepared in charcoal-stripped serum/plasma showed excellent regression coefficients (R 2 > 0.997) and covered quantities from 7.8 to 10,000 nM depending on the analyzed species. Intra- and inter-day accuracy and precision were below 15 % for all analytes. Apparent extraction recoveries were above 97 %, and ion suppression rates were between 4 and 53 %. Mean BA level in serum/plasma from healthy volunteers ranged from 11 ± 4 nM (tauroursodeoxycholic acid) to 1321 ± 1442 nM (glycochenodeoxycholic acid). As a proof of concept, the assay was applied to plasma samples derived from a clinical phase I study of myrcludex B, a novel first-in-class virus entry inhibitor for the treatment of chronic hepatitis B and D. The results demonstrate that myrcludex-induced inhibition of the hepatic BA transporter Na+-taurocholate cotransporting polypeptide (NTCP) significantly affects plasma BA level. These observations provide novel insights into drug-induced metabolic responses and will be indispensable for the assessment of side effects and dose-finding processes during future clinical trials. Graphical Abstract Figure monitoring the metabolic response of myrcludex treatment in humans
Keywords: Bile acids; Liquid chromatography quadrupole time-of-flight mass spectrometry; Na+-taurocholate cotransporting polypeptide (NTCP); Hepatitis B; Hepatitis D; Myrcludex B

Optimization strategies of in-tube extraction (ITEX) methods by Jens Laaks; Maik A. Jochmann; Beat Schilling; Torsten C. Schmidt (6827-6838).
Microextraction techniques, especially dynamic techniques like in-tube extraction (ITEX), can require an extensive method optimization procedure. This work summarizes the experiences from several methods and gives recommendations for the setting of proper extraction conditions to minimize experimental effort. Therefore, the governing parameters of the extraction and injection stages are discussed. This includes the relative extraction efficiencies of 11 kinds of sorbent tubes, either commercially available or custom made, regarding 53 analytes from different classes of compounds. They cover aromatics, heterocyclic aromatics, halogenated hydrocarbons, fuel oxygenates, alcohols, esters, and aldehydes. The number of extraction strokes and the corresponding extraction flow, also in dependence of the expected analyte concentrations, are discussed as well as the interactions between sample and extraction phase temperature. The injection parameters cover two different injection methods. The first is intended for the analysis of highly volatile analytes and the second either for the analysis of lower volatile analytes or when the analytes can be re-focused by a cold trap. The desorption volume, the desorption temperature, and the desorption flow are compared, together with the suitability of both methods for analytes of varying volatilities. The results are summarized in a flow chart, which can be used to select favorable starting conditions for further method optimization.
Keywords: In-tube extraction; ITEX; ITEX DHS; Method development; Parameter optimization

High-resolution elemental mapping of human placental chorionic villi using synchrotron X-ray fluorescence spectroscopy by Tracy Punshon; Si Chen; Lydia Finney; Louisa Howard; Brian P. Jackson; Margaret R. Karagas; Kim Ornvold (6839-6850).
The placenta is the organ that mediates transport of nutrients and waste materials between mother and fetus. Synchrotron X-ray fluorescence (SXRF) microanalysis is a tool for imaging the distribution and quantity of elements in biological tissue, which can be used to study metal transport across biological membranes. Our aims were to pilot placental biopsy specimen preparation techniques that could be integrated into an ongoing epidemiology birth cohort study without harming rates of sample acquisition. We studied the effects of fixative (formalin or glutaraldehyde) and storage duration (30 days or immediate processing) on metal distribution and abundance and investigated a thaw-fixation protocol for archived specimens stored at −80 °C. We measured fixative elemental composition with and without a placental biopsy via inductively coupled plasma mass spectrometry (ICP-MS) to quantify fixative-induced elemental changes. Formalin-fixed specimens showed hemolysis of erythrocytes. The glutaraldehyde-paraformaldehyde solution in HEPES buffer (GTA-HEPES) had superior anatomical preservation, avoided hemolysis, and minimized elemental loss, although some cross-linking of exogenous Zn was evident. Elemental loss from tissue stored in fixative for 1 month showed variable losses (≈40 % with GTA-HEPES), suggesting storage duration be controlled for. Thawing of tissue held at −80 °C in a GTA-HEPES solution provided high-quality visual images and elemental images.
Keywords: Synchrotron X-ray fluorescence; Placenta; Sample preparation

In-depth LC-MS/MS analysis of the chicken ovarian cancer proteome reveals conserved and novel differentially regulated proteins in humans by Angelito I. Nepomuceno; Huanjie Shao; Kai Jing; Yibao Ma; James N. Petitte; Michael O. Idowu; David C. Muddiman; Xianjun Fang; Adam M. Hawkridge (6851-6863).
Ovarian cancer (OVC) remains the most lethal gynecological malignancy in the world due to the combined lack of early-stage diagnostics and effective therapeutic strategies. The development and application of advanced proteomics technology and new experimental models has created unique opportunities for translational studies. In this study, we investigated the ovarian cancer proteome of the chicken, an emerging experimental model of OVC that develops ovarian tumors spontaneously. Matched plasma, ovary, and oviduct tissue biospecimens derived from healthy, early-stage OVC, and late-stage OVC birds were quantitatively characterized by label-free proteomics. Over 2600 proteins were identified in this study, 348 of which were differentially expressed by more than twofold (p ≤ 0.05) in early- and late-stage ovarian tumor tissue specimens relative to healthy ovarian tissues. Several of the 348 proteins are known to be differentially regulated in human cancers including B2M, CLDN3, EPCAM, PIGR, S100A6, S100A9, S100A11, and TPD52. Of particular interest was ovostatin 2 (OVOS2), a novel 165-kDa protease inhibitor found to be strongly upregulated in chicken ovarian tumors (p = 0.0005) and matched plasma (p = 0.003). Indeed, RT-quantitative PCR and Western blot analysis demonstrated that OVOS2 mRNA and protein were also upregulated in multiple human OVC cell lines compared to normal ovarian epithelia (NOE) cells and immunohistochemical staining confirmed overexpression of OVOS2 in primary human ovarian cancers relative to non-cancerous tissues. Collectively, these data provide the first evidence for involvement of OVOS2 in the pathogenesis of both chicken and human ovarian cancer. Graphical Abstract Translational workflow for the LC-MS/MS identification of novel differentially expressed proteins such as ovostatin 2 (OVOS2) in the chicken followed by targeted analysis in humans
Keywords: Mass spectrometry; Proteomics; Ovarian Cancer; Chicken; Ovostatin 2; OVOS2

Detection of bioavailable cadmium, lead, and arsenic in polluted soil by tailored multiple Escherichia coli whole-cell sensor set by Qihui Hou; Anzhou Ma; Thanh Wang; Jianqiang Lin; Hailin Wang; Binghai Du; Xuliang Zhuang; Guoqiang Zhuang (6865-6871).
Microbial whole-cell sensor has been widely used to assess bioavailability and risk of toxic elements, but their environmental use is still limited due to the presence of other interfering pollutants and the nonspecific binding in cells, which leads to inaccurate results. Here, we proposed a strategy combining Escherichia coli sensor set with binary regression models for the specific detection of bioavailable cadmium (Cd), lead (Pb), and arsenic (As) in a co-polluted environment. Initial tests suggested that the sensor set respectively termed pcadCluc, pzntRluc, and parsRluc could be classified into two groups according to their specific response to Cd, Pb, and As: group 1 (pcadCluc and pzntRluc) induced by a Cd-Pb mix and group 2 (parsRluc) induced by a Cd-As mix. Based on the variance in responses of each sensor to mixtures of target elements, three binary linear equations for two sensor groups were set up to calculate the individual concentrations in the mixture solutions. This method was then used to quantify the bioavailable Cd, Pb, and As in soils from a co-polluted mining region and to compare the results with other methods. Results showed that the conventional single target sensor method overestimated the bioavailability of each element, while sensor set was credible for accurate bioavailable Cd, Pb, and As quantification and comparable with the results from inductively coupled plasma mass spectrometry (ICP-MS) analysis. Our method can potentially be extended to cover the specific detection of other bioavailable toxic elements in different environmental settings. Graphical Abstract Illustration of whole-cell sensor set for bioavailable cadmium, lead, and arsenic detection
Keywords: Heavy metals; Bioavailability; Specificity; Whole-cell sensor set; Cross-mixed induction; Binary linear regression

Comprehensive evaluation of methods to isolate, quantify, and characterize circulating cell-free DNA from small volumes of plasma by Florence Mauger; Cécile Dulary; Christian Daviaud; Jean-François Deleuze; Jorg Tost (6873-6878).
Circulating cell-free DNA (ccfDNA) has great potential for non-invasive diagnostics, and prediction and monitoring of treatment response, but its amount is usually limited. Therefore, the choice of methods to extract and characterize ccfDNA is crucial. In the current study, we performed the most comprehensive comparison of methods for ccfDNA extraction (11 methods), quantification (3 methods), and estimation of the integrity index (2 methods) from small quantities of different kinds of plasma. The QIAamp® Circulating Nucleic Acid Kit and the Norgen Plasma/Serum Circulating DNA Purification Mini Kit showed the best accuracy and reproducibility, but the Norgen kit allowed to extract a higher amount of ccfDNA. This workflow provides a reliable protocol for the multiple applications of ccfDNA in biomedicine. Graphical Abstract Workflow for the evaluation of methods to isolate, quantify and characterize circulating cell-free DNA from small volumes of plasma
Keywords: Circulating cell-free DNA; Plasma; Extraction; Quantification; Integrity index; Cancer; Non-invasive diagnostics and personalized treatment

The paper presents the construction and metrological characteristics of the home-made in situ passive flux sampler, an analytical tool representing small-scale emission chambers working in situ and passively sampling analytes from the gaseous phase. The sorption element was a cylindrical container made of stainless steel net, packed with a carbon sorbent bed-graphitized charcoal, Carbograph 4 (35/50 mesh). The recommended working/exposure time of the constructed passive device was determined by carrying out model tests in the laboratory. In addition, a preliminary study was conducted to determine the rate of the emission flux of selected monoterpenes released from the surface of wood-based indoor materials (laminated chipboard) used in residential areas.
Keywords: Passive flux sampler; Emission flux; Monoterpenes; Indoor materials; In situ measurements

Hemoglobin assay for validation and quality control of medical device reprocessing by Justin Frey; Allan Guan; Zhenyu Li; Steven Turtil; K. Scott Phillips (6885-6889).
Hemoglobin (Hb) is an important analyte in medicine, forensics, and research. One area of crucial need for real-world Hb quantitation is the validation and quality control (QC) of reprocessed medical device cleaning. Here, we show how a microplate reader and colorimetric blood test strips can be used to quantitate nanogram (ng) quantities of Hb in a 1-min assay. The assay had a linear range of 0–50 ng (0–370 ng on a log scale) for Hb, with a limit of detection (LOD) of 3.3 ng, which was ∼500-fold more sensitive than the micro-BCA reagent (LOD = 1.6 μg) and on the same order of magnitude as detection of labeled Hb with fluorescence (LOD = 1.9 ng). For validation of medical device cleaning, the assay was specific for Hb in the presence of artificial test soil and was unaffected by interferences from common cleaning reagents at 10 ppm. Lubricant and sodium dodecyl sulfate did not significantly affect the assay at 10 ppm but affected the assay at 1 % g/g. The method showed 100 % recovery of hemoglobin in extracted soils, with extraction from silicone having the greatest variability in recovery, while Teflon and stainless steel had <10 % RSD. The assay makes it possible for medical device companies and health-care providers to obtain crucially needed information on the cleanliness of reused devices.
Keywords: Hemoglobin; Medical device reprocessing; Reflectivity

Polyamines and their N-acetylated metabolites are potential biomarkers in the diagnosis and therapeutic evaluation of cancer. Thus, we present here an ultra high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for the simultaneous determination of 6 free, 3 monoacetylated, and 2 diacetylated polyamines without derivatization. The major improvement of this method is the use of 0.2 % perfluoroheptanoic acid methanol in the pretreatment step to achieve protein precipitation and 0.0125 % perfluoroheptanoic acid in the mobile phase to achieve analyte separation within 9 min. The established analytical method was validated with plasma, urine, and liver tissue and applied to determine plasma, urine, and liver tissue samples from healthy rats, hepatocellular carcinoma rats, and administrated rats successfully. Results indicated free polyamines such as putrescine mainly existed in liver tissue but more polar N-acetylated metabolites such as N 1,N 12-diacetylspermine seemed to exist in biological fluid. After carcinogenesis, the levels of polyamines were increased, but the elevated levels of polyamines and their metabolites tended to decrease when administrated with anticancer drug. The method provided a more versatile manner for clinical application in the diagnosis and therapeutic evaluation of hepatocellular carcinoma. Graphical Abstract The process of quantification of free polyamines and their metabolites in biofluids and liver tissue by UHPLC-MS/MSᅟ
Keywords: Polyamines; Biomarkers; Diagnosis; Hepatocellular carcinoma; UHPLC-MS/MS method