Elena S. Chernetsova

Determination of drugs and drug-like compounds in different samples with direct analysis in real time mass spectrometry.

Direct analysis in real time (DART), a relatively new ionization source for mass spectrometry, ionizes small-molecule components from different kinds of samples without any sample preparation and chromatographic separation. The current paper reviews the published data available on the determination of drugs and drug-like compounds in different matrices with DART-MS, including identification and quantitation issues. Parameters that affect ionization efficiency and mass spectra composition are also discussed.

Combined multivariate data analysis of high-performance thin-layer chromatography fingerprints and direct analysis in real time mass spectra for profiling of natural products like propolis

Sophisticated statistical tools are required to extract the full analytical power from high-performance thin-layer chromatography (HPTLC). Especially, the combination of HPTLC fingerprints (image) with chemometrics is rarely used so far. Also, the newly developed, instantaneous direct analysis in real time mass spectrometry (DART-MS) method is perspective for sample characterization and differentiation by multivariate data analysis. This is a first novel study on the differentiation of natural products using a combination of fast fingerprint techniques, like HPTLC and DART-MS, for multivariate data analysis. The results obtained by the chemometric evaluation of HPTLC and DART-MS data provided complementary information. The complexity, expense, and analysis time were significantly reduced due to the use of statistical tools for evaluation of fingerprints. The approach allowed categorizing 91 propolis samples from Germany and other locations based on their phenolic compound profile. A high level of confidence was obtained when combining orthogonal approaches (HPTLC and DART-MS) for ultrafast sample characterization. HPTLC with selective post-chromatographic derivatization provided information on polarity, functional groups and spectral properties of marker compounds, while information on possible elemental formulae of principal components (phenolic markers) was obtained by DART-MS.

ID-CUBE direct analysis in real time high-resolution mass spectrometry and its capabilities in the identification of phenolic components from the green leaves of Bergenia crassifolia L.

RATIONALE Bergenia crassifolia is a plant widely used in herbal medicine. Its chemical composition has been little studied, and no studies using high-resolution mass spectrometry (HRMS) have been performed. Its phenolic components are of particular interest, due to the interest in such compounds in medicine and cosmetics. The ID-CUBE, a simplified Direct Analysis in Real Time (DART) ion source, suitable for the fast MS analysis of liquids without complex sample preparation, offers a new method of studying extracts of such plant. Coupling the ID-CUBE with a high-resolution mass spectrometer can provide identification of extract components. METHODS Mass spectral conditions were optimized for model solutions of the flavonoid naringenin and used for the identification of phenolic compounds in green leaves extracts of Bergenia crassifolia. OpenSpot sample cards with a metal grid surface were used for sample introduction into the ID-CUBE ion source on an Obitrap mass spectrometer. The samples were applied as 5-μL aliquots of the extract onto the metal grid of the card. Sample ionization was stimulated in the ion source within 20 s by applying an electric current to the metal grid to thermally desorb the analytes into the gas flow of metastable helium atoms from the ID-CUBE. RESULTS Elemental compositions were assigned to abundant ions in the mass spectra of the extracts. The major phenolic components were confirmed by their [M-H](-) ions. Thirty-six other marker ions were found, and elemental compositions were suggested for 30% of them, based on a search for compounds found in herbal extracts. CONCLUSIONS The ID-CUBE-Orbitrap MS coupling allowed the rapid accurate mass determination of the phenolic components (and other compounds) in herbal extracts. Higher confidence in component identification could be provided by using additional structural elucidation methods, including tandem mass spectrometry (MS/MS), and this will be the focus of future studies.

Some new features of Direct Analysis in Real Time mass spectrometry utilizing the desorption at an angle option

The present study is a first step towards the unexplored capabilities of Direct Analysis in Real Time (DART) mass spectrometry (MS) arising from the possibility of the desorption at an angle: scanning analysis of surfaces, including the coupling of thin-layer chromatography (TLC) with DART-MS, and a more sensitive analysis due to the preliminary concentration of analytes dissolved in large volumes of liquids on glass surfaces. In order to select the most favorable conditions for DART-MS analysis, proper positioning of samples is important. Therefore, a simple and cheap technique for the visualization of the impact region of the DART gas stream onto a substrate was developed. A filter paper or TLC plate, previously loaded with the analyte, was immersed in a derivatization solution. On this substrate, owing to the impact of the hot DART gas, reaction of the analyte to a colored product occurred. An improved capability of detection of DART-MS for the analysis of liquids was demonstrated by applying large volumes of model solutions of coumaphos into small glass vessels and drying these solutions prior to DART-MS analysis under ambient conditions. This allowed the introduction of, by up to more than two orders of magnitude, increased quantities of analyte compared with the conventional DART-MS analysis of liquids. Through this improved detectability, the capabilities of DART-MS in trace analysis could be strengthened.

Coupling of planar chromatography with Direct Analysis in Real Time mass spectrometry

AbstractDirect Analysis in Real Time mass spectrometry (DART-MS) is an emerging and rapidly developing area of ambient desorption ionization mass spectrometric techniques. Its coupling with planar chromatography is especially promising, as compared to other ambient desorption ionization techniques, because it does not require the use of liquids that may distort the shape of a spot by diffusion effects. In the first publications on TLC/HPTLC-DART-MS, due to the fixed, horizontally aligned supply of the gas flow from the DART ionization source to the MS inlet, the introduction of HPTLC/TLC plates as cut strips was inconvenient for quantitation, and the repeatability was very low due to the manual positioning. Recently a new version of the DART ion source was suggested, which allows adjusting the angle of the DART gas stream and the use of a motorized rail, thereby, improving highly the capabilities of TLC/HPTLC-DART-MS. This comprehensive review describes the development and analytical capabilities of TLC/HPTLC-DART-MS, and the general DART-MS perspectives for surface analysis or imaging MS.

Letter: Characterization of Volatile and Semi-Volatile Compounds in Green and Fermented Leaves of Bergenia Crassifolia L. by Gas Chromatography-Mass Spectrometry and ID-CUBE Direct Analysis in Real Time-High Resolution Mass Spectrometry:

Chemical compositions of volatile and semi-volatile components in green and fermented leaves of Bergenia crassifolia L. were studied. Leaf components were identified using gas chromatography with low resolution mass spectrometry and direct analysis in real time (DART) high resolution mass spectrometry with an ID-CUBE ion source. Phytol, nerolidol, geraniol, linalool, α-bisabolol, α-bisabololoxide B, α-cadinol, δ-cadinene, α-terpineol and several other marker compounds of special interest were defined, for which the process of fermentation significantly changed their content in the leaves. Low resolution EI GC-MS and ID-CUBE DART-HRMS were found to be complementary methods, as they provide different information, helpful to increase the confidence of identification.

Electron ionization mass spectra and their reproducibility for trialkylsilylated derivatives of organic acids, sugars and alcohols.

Mass spectra of trialkylsilyl derivatives of fatty acids, dicarboxylic acids, hydroxyacids, oxoacids, sugars, amino acids and alcohols were obtained. Amino acids were analyzed as tert-butyldimethylsilyl derivatives; all other model compounds were analyzed as trimethylsilyl derivatives. Reproducibility of the electron ionization (EI) mass spectra for the derivatives obtained was discussed. It was shown that, for many investigated derivatives, composition of the respective mass spectra depended greatly on ion source contamination. The trimethylsilylated α-tocopherol mass spectrum composition was most significantly influenced by ion source contamination. This compound can be used to test ion source contamination.

Aspects of surface scanning by direct analysis in real time mass spectrometry employing plasma glow visualization.

RATIONALE Visual monitoring of the Direct Analysis in Real Time (DART) gas impact region during sampling was demonstrated via its metastable plasma glow. It is known that adding neon into helium for DART leads to plasma glow, but this effect has not been used in practice and discussed in the literature so far. METHODS A single quadrupole mass spectrometer with a DART SVPA ion source was used for recording of DART mass spectra from different surfaces, using galangin and p-coumaric acid as model analytes. In specific cases, the composition of the mass spectra was clarified using an Orbitrap mass spectrometer. RESULTS Plasma glow visualization made it possible to track the metastable gas distributions during surface scanning. The influence on the composition of the mass spectra was studied for different carrier gases, i.e. pure helium versus a helium-neon mixture, and for the vacuum pumping rate. The spatial resolution was substantially improved via a DART cap with a narrowed internal diameter, but impaired by a decreased sensitivity. Comparably low signal intensities were obtained for analytes on porous layers due to analyte penetration and metastable gas scattering. CONCLUSIONS Visualization through the plasma glow enables the optimal selection of the coordinates for DART-MS analysis and thus it will support scanning and imaging MS on surfaces, including porous planar chromatographic separation materials.

An ultra superfast identification of low-molecular components of pharmaceuticals by DART mass spectrometry

1537 Direct analysis in real time (DART) was invented and patented in 2005 as a new ionization method for mass spectrometry [1, 2]. The abbreviation DART was also selected, because the word dart has another meaning, that is, arrow, which reflects the promptness and high accuracy of this method. The method offers the possibility of analyzing solid and liquid samples without a sample preparation, introducing samples directly into the ionization region. The ionization occurs immediately under atmospheric pressure; because the ionization takes place in the vapor phase, high molecular matrix components do not interfere. DART mass spectrometry attracts considerable inter est, because various samples—pharmaceutical prepa rations, narcotics, beverages, foodstuff, explosives, inks, biological fluids and tissues, and others—can be analyzed without a sample preparation [1]. The number of publications on the application of DART mass spectrometry remains very small. In par ticular, publications concerning the analysis of phar maceuticals are limited by the data for some solid pharmaceuticals and substances and for only one oint ment [1, 3–6]. Therefore, further studies of various pharmaceutical preparations by DART mass spec trometry are very important for the reliable evalution of the potential of this method. In the present work, we studied the DART mass spectra of tablet of Anapryline, Aspirin, Biseptol, Gly cine, Mexidol, Nootropyl, Furacillin, Erespal, and Ersefuryl, as well as ointments Tetracycline, Syntho mycin, and Levomecol. The corresponding prepara tions were purchased in pharmacies in Moscow. The samples were selected randomly. We used an AccuTOF time of flight mass spec trometer (JEOL, Japan) equipped with a DART ion source (IonSense, United States). The sample tablets were placed into the gap between the DART ion source and the inlet of the mass spectrometer by for ceps. In the cases of ointments, a small amount of a sample (3–5 μL) was placed into the ion source using a glass melting point stick. Helium 5.0–6.0 grade (more than 99.999% purity), heated to 260°C, was used for ionization (Moscow Gas Processing Plant, Russia). The helium flow rate was 2 L/min. Positive ion mass spectra were recorded. The voltage on the discharge needle of the DART source was 4 kV. The An Ultra Superfast Identification of Low Molecular Components of Pharmaceuticals by DART Mass Spectrometry

Organic elemental analysis: a new universal approach to authenticity/quality control of pharmaceuticals.

Expedient and reliable methods for the quantitation of active ingredients in pharmaceutical dosage forms and substances are essential due to the growing problem of counterfeit pharmaceuticals and ineffective quality control in pharmaceutical production.[1] This problem arises from the fact that the active ingredient content is often determined by different methods such as high performance liquid chromatography (HPLC), titrimetry, or potentiometry. Using these methods, different determination conditions are required, which are specific to determining the targeted active ingredient.[2,3] In such cases, the reference samples are used and the calibration is performed for each individual compound. These methods are very time-, labour-, and cost-consuming. As such, they are inappropriate for high-throughput quality control at the site of production. Organic elemental analysis (OEA) is an alternative, fast and relatively inexpensive method for active pharmaceutical ingredient content determination. We have reviewed the structures of 2500 pharmaceutical compounds which are used as active ingredients in many pharmaceuticals. It was found that approximately 80% of these compounds contain nitrogen, although in the majority of cases the excipients of pharmaceutical dosage forms do not. Thus by determining the nitrogen content it is possible to find out the content of the active ingredient in the respective dosage form, if we determine the nitrogen content in that dosage form. This indirect approach does not require extraction of the active ingredient, sample referencing, or the necessity for accurate and reproducible extract sample injection and calibration. The duration of one analysis depends only on the preparation and weighing of the small part of the grounded solid matrix (or solution) and determination of nitrogen content in that sample by elemental analysis. In addition the determination of nitrogen content by elemental analysis takes only five minutes or less. Therefore, the accuracy and throughput of such determination is much higher than that of the commonly used methods such as HPLC, titrimetry, and potentiometry. In our opinion, the advantages of such an approach are apparent. Nevertheless, there are no publications in which active pharmaceutical ingredients in dosage forms are quantified by elemental analysis. The purpose of the present work was to study the possibility of fast active ingredient content determination in different pharmaceuticals and substances using an automated elemental analyzer, by means of nitrogen content determination in respective samples. Helium (99.9999%) and oxygen (99.999%) were obtained from PromGasService (Moscow, Russia). Cystine (C6H12N2O4S2) purchased from ThermoFinnigan (Milan, Italy) was used as the only reference compound for all studied samples. Atropine, sulfanilamide, pentoxifylline, captopril, and BBOT were used as target substances. Target pharmaceutical dosage forms were obtained from different pharmacies in Moscow, Russia. Organic elemental analyzers model Flash EA 1112 (ThermoFinnigan, Milan, Italy) and model Dumatherm (Gerhardt, Germany) were configured as N-analyzers. They were used for the determination of active ingredient of target pharmaceutical dosage forms and substances by means of nitrogen quantitation. Analytical balance model MX5 Mettler Toledo, Greifensee, Switzerland was used for the weighing of samples. All samples were weighed and introduced into the OEA autosampler in tin containers (ThermoFinnigan, Milan, Italy). A 4-point calibration was performed each day prior to the analysis of target pharmaceuticals. The weight of cystine sample (reference compound) was at least 0.5 mg. Substances or powders from capsules were analyzed directly. In the case of tablets, sample preparation included homogenization (grinding) of 3 tablets of the respective pharmaceutical. Portions of the respective powder (not less than 0.3 mg and not more than 6.5 mg) were analyzed by OEA. The weight of the analyzed sample portion was dependant on the nitrogen content in the active compound molecule and also on the specified content of the active ingredient in its dosage form. It was chosen according to the range of nitrogen content used for the calibration curve in such a manner that the expectable weight of nitrogen in a sample portion was within the minimal and maximal nitrogen weights used for the calibration curve. In the case of liquid sample analysis, a sorbent was used. Ethanol and methanol evaporated too quickly from the sorbent. Therefore, model solutions used for analysis were prepared in water. Model water solution of the active ingredient with known concentration was placed on the sorbent immediately prior to the analysis. The same sample preparation was used for the liquid pharmaceuticals from ampoules. The quantitation of the active N-containing ingredient in various pharmaceutical dosage forms and substances using elemental analysis was investigated. Using the proposed OEA approach, analysis of pharmaceutical substances and various pharmaceutical dosage forms was carried out. The data obtained for solids are presented in Tables 1 and 2. As presented in Table 1, the difference between the specified and experimental values of nitrogen content in substances was within the limits of allowable error. The determined degree of purity was approximately 100%. The overestimated values for the content of active ingredient could be explained with the presence of N-containing impurities.