Mass spectrometry as a tool for studying tautomerism

The review contains new data on tautomerism of organic compounds belonging to different classes, which were obtained by mass spectrometry and confirmed by quantum-chemical calculations. Published in Russian in Zhurnal Organicheskoi Khimii, 2008, Vol. 44, No. 12, pp. 1751–1763. The text was submitted by the authors in English.     

Mass spectrometry: a tool for the identification of proteins separated by gels

Mass spectrometry (MS) has become the technique of choice to identify proteins. This has been largely accomplished by the combination of high-resolution two-dimensional (2-D) gel separation with robotic sample preparation, automated MS measurement, data analysis, and database query. Developments during the last five years in MS associated with protein gel separation are reviewed.     

Mass spectrometry as a tool for the selective profiling of destruxins; their first identification in Lecanicillium longisporum

Mass spectrometry was applied to the identification of the destruxins (dtxs), cyclic peptides that are commonly produced by the fungal insect‐pathogen, Metarhizium anisopliae. The aim of the study was to optimise a methodology in order to firstly determine whether these compounds were present in other species and to determine the effect of differing growth conditions upon the dtx content detected. Matrix‐assisted laser desorption/ionisation time‐of‐flight mass spectrometry (MALDI‐ToF‐MS) was initially used to analyse the dtxs, but limitations were indicated. Nano‐scale high‐performance liquid chromatography/electrospray ionisation mass spectrometry (HPLC/ESI‐MS) and automated ‘data‐dependent’ tandem mass spectrometric (MS/MS) analysis were also applied, utilising characteristic neutral losses during fragmentation to confirm the presence of the dtxs. This latter approach distinguished the dtx E and B isoforms by retention time and diagnostic neutral losses during fragmentation allowing extraction of the destruxin data from a complex dataset. This process revealed the presence of a number of dtxs in the fungal species Lecanicillium longisporum, a species previously not known to produce dtxs, and dtx production in this species was shown to be significantly higher in aerated cultures compared with still cultures. Copyright © 2009 John Wiley & Sons, Ltd.     

Mass spectrometry as a diagnostic tool for 5-hydroxysteroids

The mass spectra of twelve 5-hydroxysteroids (1–12), indicate that a common feature is the formation of the ion a by loss of the four ring A C atoms 1, 2, 3, and 4, with the H atoms and other substituents attached.     

HPLC/APCI-FTICR-MS as a tool for identification of partial polar mutagenic compounds in effect-directed analysis

Identification of unknown compounds remains one of the biggest challenges for the assignment of adverse effects of sediment contamination and other complex environmental mixtures to responsible toxicants by effect-directed analysis (EDA). The identification depends on information gained from biotesting, chromatographic separation, and mass spectrometric detection. Thus, a methodology is provided for non-target identification of partial polar mutagenic polyaromatic compounds in sediment extracts by using polymeric reversed-phase HPLC column, high-resolution mass spectrometry and PubChem database. After visualization and processing the chromatogram constituents by using deconvolution software, the unambiguous elemental compositions generated were used as input in PubChem database to find a possible identity for the suspected species. The retrieved structures from the database search were refined by characterized chromatographic and mass spectrometric classifiers based on 55 model compounds comprising eight different classes representing mutagenic substructures. The applicability of the method was demonstrated by positive and tentative identification of constituents of mutagenic sediment fractions similar to selected model compounds.     

H-N HMBC as a valuable tool for the identification and characterization of nitrones

¹H-¹⁵N HMBC has been evaluated as an efficient and high-speed method to determine ¹⁵N chemical shifts for nitrones, which can be used to identify aromatic nitrones and to extract structural information by comparison with reference data. Substituent effects have been measured on C and N aryl groups separately, showing up the influence of electronic effects on C-aryl groups rather than N-aryl groups on the ¹⁵N chemical shifts. Steric effects are remarkable in the case of C-aryl-N-alkyl nitrones. Depending on N or C substitution, chemical shift changes in such an additive way that it is possible to predict chemical shifts for unknown nitrones.     

Interpolation of relative retentions as a tool for identification of perhalogenated compounds

Summary Separation of perhalogenated compounds resulting from thermolysis, photolysis or radiolysis of mixtures of halogens in carbon tetrachloride was carried out by programmed temperature gas chromatography. Plots of relative retentions (to CBr₄) vs the number of bromine atoms present for known compounds of the general formula C_xBr_yCl_z (x=1 and 2; y+z=4 or 6) gave three linear correlations (all with r>0.99). These correlations permitted projections of the identities of reaction products for which no standards were available. These projections were later confirmed by CG-MS. *** DIRECT SUPPORT *** A0732166 00010     

Mass spectrometry imaging as a tool for surgical decision‐making

Despite significant advances in image‐guided therapy, surgeons are still too often left with uncertainty when deciding to remove tissue. This binary decision between removing and leaving tissue during surgery implies that the surgeon should be able to distinguish tumor from healthy tissue. In neurosurgery, current image‐guidance approaches such as magnetic resonance imaging (MRI) combined with neuronavigation offer a map as to where the tumor should be, but the only definitive method to characterize the tissue at stake is histopathology. Although extremely valuable information is derived from this gold standard approach, it is limited to very few samples during surgery and is not practically used for the delineation of tumor margins. The development and implementation of faster, comprehensive, and complementary approaches for tissue characterization are required to support surgical decision‐making – an incremental and iterative process with tumor removed in multiple and often minute biopsies. The development of atmospheric pressure ionization sources makes it possible to analyze tissue specimens with little to no sample preparation. Here, we highlight the value of desorption electrospray ionization as one of many available approaches for the analysis of surgical tissue. Twelve surgical samples resected from a patient during surgery were analyzed and diagnosed as glioblastoma tumor or necrotic tissue by standard histopathology, and mass spectrometry results were further correlated to histopathology for critical validation of the approach. The use of a robust statistical approach reiterated results from the qualitative detection of potential biomarkers of these tissue types. The correlation of the mass spectrometry and histopathology results to MRI brings significant insight into tumor presentation that could not only serve to guide tumor resection, but that is also worthy of more detailed studies on our understanding of tumor presentation on MRI. Copyright © 2013 John Wiley & Sons, Ltd.     

Efficient knot group identification as a tool for studying entanglements of polymers

A technique is presented for the identification of the knot group of knots, links, and other embedded graphs as a tool in numerical studies of entanglements of polymers. With this technique, the knot group is simultaneously more discriminating and easier to calculate than the knot invariants that have been used in such studies in the past. It can be applied even in cases of very complex knot projections with hundreds of crossings. Starting from an arbitrary projection of an embedded graph, we generate a sequence of representations, any one of which is a full and complete representation of the knot group. Any two knot groups are isomorphic if they have identical representations. Therefore, we compare the sequence of representations of any given knot or link against a previously determined lookup table, and if the group of the knot or link is represented in this table we eventually find a match and identify the knot group.     

Raman microspectroscopic mapping as a tool for detection of gunshot residue on adhesive tape

Analytical and Bioanalytical Chemistry - Our research group previously reported a novel method for the detection of gunshot residue (GSR) via tape lifting combined with Raman microspectroscopic...     

SERS as tool for the analysis of DNA-chips in a microfluidic platform

A sequence-specific detection method of DNA is presented combining a solid chip surface for immobilisation of capture DNAs with a microfluidic platform and a readout of the chip based on SERS. The solid chip surface is used for immobilisation of different capture DNAs, where target strands can be hybridised and unbound surfactants can be washed away. For the detection via SERS, short-labelled oligonucleotides are hybridised to the target strands. This technique is combined with a microfluidic platform that enables a fast and automated preparation process. By applying a chip format, the problems of sequence-specific DNA detection in solution phase by means of SERS can be overcome. With this setup, we are able to distinguish between different complementary and non-complementary target sequences in one sample solution.     

Mass spectrometry and partial least-squares regression: a tool for identification of wheat variety and end-use quality

Rapid methods for the identification of wheat varieties and their end-use quality have been developed. The methods combine the analysis of wheat protein extracts by mass spectrometry with partial least-squares regression in order to predict the variety or end-use quality of unknown wheat samples. The whole process takes approximately 30 min. Extracts of alcohol-soluble storage proteins (gliadins) from wheat were analysed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Partial least-squares regression was subsequently applied using these mass spectra for making models that could predict the wheat variety or end-use quality. Previously, an artificial neural network was used to identify wheat varieties based on their protein mass spectra profiles. The present study showed that partial least-squares regression is at least as useful as neural networks for this identification. Furthermore, it was demonstrated that partial least-squares regression could be used to predict wheat end-use quality, which has not been possible using neural networks.     

A flexible data analysis tool for chemical genetic screens

High-throughput assays generate immense quantities of data that require sophisticated data analysis tools. We have created a freely available software tool, SLIMS (Small Laboratory Information Management System), for chemical genetics which facilitates the collection and analysis of large-scale chemical screening data. Compound structures, physical locations, and raw data can be loaded into SLIMS. Raw data from high-throughput assays are normalized using flexible analysis protocols, and systematic spatial errors are automatically identified and corrected. Various computational analyses are performed on tested compounds, and dilution-series data are processed using standard or user-defined algorithms. Finally, published literature associated with active compounds is automatically retrieved from Medline and processed to yield potential mechanisms of actions. SLIMS provides a framework for analyzing high-throughput assay data both as a laboratory information management system and as a platform for experimental analysis.     

Atomic force microscopy as a tool for binder identification in ancient photographic processes

Focusing in the photographic archive of Julián Carrillo (México), a microstructural analysis of surface photography was carried on to prove the feasibility of atomic force microscopy to identify the binder used in the manufactured ancient photography. It has been identified 2 structural groups in which known binders may be classified, one of them is associated with gelatin and the other one to albumen and collodion. The former group present a higher roughness values than the second one, being able to identify the binder by means of its morphology aspect related with the width of linear traces. From the obtained results, it has been observed that atomic force microscopy is a very good technique to properly identify the binder used in a photograph according to the surface morphology and roughness values.     

Polarography as a tool in organic analysis

Summary Rapid indirect polarographic and visual micromethods have been developed for the determination of the quinone and azoxy groups by reduction with titanium(III) at pH values of 3 and 5.5, respectively. The excess of reductant is first determined polarographically followed by visual back-titration of the same solution with iron(III) using thiocyanate as indicator a drop of neutral red is added in case of the quinone function. Although the results obtained by the two finishes are of approximately the same order of accuracy (±0.46%), yet the polarographic finish proved to be of more general applicability particularly for quinones that give rise, after reduction, to a variety of colours on adding the iron(III) solution.

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Anwendung der Polarographie in der organischen AnalyseIV. Bestimmung von Chinon- und Azoxygruppen durch Reduktion mit Titan(III)

Zusammenfassung Zur schnellen Bestimmung von Chinon- und Azoxygruppen durch Reduktion mit Titan(III) bei pH 3 bzw. 5,5 werden indirekte polarographische und visuelle Mikromethoden beschrieben. Der Überschuß an Reduktionsmittel wird zunächst polarographisch und anschließend volumetrisch durch Rücktitration mit Eisen(III)-lösung bestimmt, wobei Thiocyanat (bei der Chinongruppe noch zusätzlich Neutralrot) als Indicator dient. Beide Ergebnisse weisen etwa dieselbe Genauigkeit auf (±0,46%); doch ist die polarographische Methode allgemeiner anwendbar, besonders bei Chinonen, die nach der Reduktion auf Zusatz Eisen(III) verschiedenartige Färbungen hervorrufen.

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Part III: Mikrochim. Acta 1969, 44.     

Procrustes rotation as a diagnostic tool for projection pursuit analysis

Projection pursuit (PP) is an effective exploratory data analysis tool because it optimizes the projection of high dimensional data using distributional characteristics rather than variance or distance metrics. The recent development of fast and simple PP algorithms based on minimization of kurtosis for clustering data has made this powerful tool more accessible, but under conditions where the sample-to-variable ratio is small, PP fails due to opportunistic overfitting of random correlations to limiting distributional targets. Therefore, some kind of variable compression or data regularization is required in these cases. However, this introduces an additional parameter whose optimization is manually time consuming and subject to bias. The present work describes the use of Procrustes analysis as diagnostic tool that can be used to evaluate the results of PP analysis in an efficient manner. Through Procrustes rotation, the similarity of different PP projections can be examined in an automated fashion with “Procrustes maps” to establish regions of stable projections as a function of the parameter to be optimized. The application of this diagnostic is demonstrated using principal components analysis to compress FTIR spectra from ink samples of ten different brands of pen, and also in conjunction with regularized PP for soybean disease classification.     

Polarography as a tool in organic analysis

Summary A polarographic micromethod has been developed for the analysis of mono-, di-, tri- and polynitro compounds, standard titanium(III) sulphate solution being used as a reducing agent. The cathodic part of the Kalousek cell has been modified to serve as a reaction vessel. The results agree favourably with those obtained by visual back-titration of excess of titanium(III) with iron(III), thiocyanate being used as indicator, and the relative error is ± 0.5%. This polarographic finish proved to be of general applicability particularly with nitro compounds that give rise to coloured reduction products.

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Zusammenfassung Eine polarographische Mikromethode zur Analyse von Mono-, Di-, Tri- und Polynitroverbindungen mit Titan(III)-sulfatlösung als Reduktionsmittel wurde ausgearbeitet. Der Kathodenteil der Kalousek-Zelle wurde als Reaktionsgefäß umgestaltet. Die Analysenresultate stimmen sehr gut mit jenen überein, die man durch visuelle Rücktitration des überschüssigen Titan(III) mit Eisen(III) gegen Thiocyanat als Indikator erhält. Der relative Fehler beträgt ± 0,5%. Dieses polarographische Verfahren dürfte allgemein für Nitroverbindungen anwendbar sein, aus denen sich gefärbte Reduktions-produkte bilden können.

     

The synergistic chaperoning operation in a Bi-chaperone system consisting of alpha-crystallin and beta-casein: bovine pancreatic insulin as the target protein

While chaperone activity of alpha-crystallin (α-Crs) is important in maintaining lens transparency that of beta-casein (β-CN) is vital to prevent the development of corpora amylacea (accumulation of amyloid deposits in mammary glands). These two chaperone proteins are amphiphilic, each contains distinct polar and non-polar regions in the structure. While polar domain of α-Crs is highly electropositive, the counterpart domain in β-CN is strongly electronegative. In this study a Bi-chaperone system consisting of α-Crs and β-CN with different molar ratios were used to prevent the chemical-induced insulin aggregation spectroscopically. As shown, α-Crs and β-CN in the Bi-chaperone system exhibit synergistic chaperoning operation which strongly depends to the specific ratio of the chaperone components. The results of both fluorescence study and native gel electrophoresis confirmed the non-covalent interactions between α-Crs and β-CN. Consequently the synergistic activity can be explained with the possible electrostatic interactions between their polar/charged domains which bring them in close proximity, allowing their synergistic chaperoning operation in the Bi-chaperone system.     

Validation of a new reversed-phase high-performance liquid chromatography method for separation and quantification of bovine milk protein genetic variants

A new RP-HPLC method for the separation and quantification of the most common genetic variants of bovine milk proteins is described. A reversed-phase analytical column C8 (Zorbax 300SB-C8 RP, 3.5 microm, 300A, 150 x 4.6 I.D.) was used. All the most common casein (CN) and whey protein genetic variants, including beta-CN(I) were detected and separated simultaneously in less then 40 min, with the exception of alpha(S1)-CN(B) and CN(C) variants. Purified protein genetic variants were employed in calibration and showed different absorbances at 214 nm. The procedure was developed using 40 raw individual milk samples of cows belonging to four different breeds and certified skim milk powder BCR-063R. Method validation consisted in testing linearity, repeatability, reproducibility and accuracy. A linear relationship (R(2)>0.99) between the concentrations of proteins and peak areas was observed over the concentration range, with low detection limits. Repeatability and reproducibility were satisfactory for both retention times and peak areas. The RSD of peak areas ranged from 0.92 to 4.32% within analytical day and from 0.85 to 9.52% across analytical days. The recoveries, calculated using mixtures of samples previously quantified, ranged from 98.1 to 103.7%.