Calibration-Curve-Locking Database for Semi-Quantitative Metabolomics by Gas Chromatography/Mass Spectrometry.

Calibration-Curve-Locking Databases (CCLDs) have been constructed for automatic compound search and semi-quantitative screening by gas chromatography/mass spectrometry (GC/MS) in several fields. CCLD felicitates the semi-quantification of target compounds without calibration curve preparation because it contains the retention time (RT), calibration curves, and electron ionization (EI) mass spectra, which are obtained under stable apparatus conditions. Despite its usefulness, there is no CCLD for metabolomics. Herein, we developed a novel CCLD and semi-quantification framework for GC/MS-based metabolomics. All analytes were subjected to GC/MS after derivatization under stable apparatus conditions using (1) target tuning, (2) RT locking technique, and (3) automatic derivatization and injection by a robotic platform. The RTs and EI mass spectra were obtained from an existing authorized database. A quantifier ion and one or two qualifier ions were selected for each target metabolite. The calibration curves were obtained as plots of the peak area ratio of the target compounds to an internal standard versus the target compound concentration. These data were registered in a database as a novel CCLD. We examined the applicability of CCLD for analyzing human plasma, resulting in time-saving and labor-saving semi-qualitative screening without the need for standard substances.

Atmospheric Pressure Dark-Current Argon Discharge Ionization with Comparable Performance to Direct Analysis in Real Time Mass Spectrometry.

Herein, a dark-current discharge state created by combining argon flow with a needle electrode in ambient air is described that has an ionization efficiency and mechanism comparable to those of conventional helium direct analysis in real time (DART), without requiring dopants and DART glow discharge. Using this method, polar compounds such as α-amino acids were ionized in the dark-current argon discharge via (de)protonation, molecular anion formation, fragmentation, (de)protonation with the attachment of oxygen, deprotonation with hydrogen loss and negative ion attachment. In contrast, nonpolar compounds (e.g., n-alkanes) were detected as positive ions via hydride abstraction and oxidation. Major background ions observed were H3O+(H2O) n , O2 ·+, O2 ·-(H2O) n and CO3 ·-. These results indicate that the present dark-current discharge efficiently generates resonance-state argon with an internal energy of ∼14.2 eV, higher than that of the well-known metastable state (∼11.6 eV). It is therefore suggested that ionization reactions occurring there can be attributed to the Penning ionization of atmospheric H2O and O2 by resonance-state argon, analogous to helium DART.

Improvement in ionization efficiency of direct analysis in real time-mass spectrometry (DART-MS) by corona discharge.

Herein it is shown that a combination of direct analysis in real time (DART) with a corona discharge system consisting of only a needle electrode easily improves DART ionization efficiency. Positive and negative DC corona discharges led to a formation of abundant excited helium atoms as well as the reactant ions H3O(+)(H2O)n and O2˙(-) in the DART analyte ionization area. These phenomena resulted in an increase in the absolute intensities of (de)protonated analytes by a factor of 2-20 over conventional DART. The other analyte ions detected in this corona-DART system (i.e., molecular ions, fragment ions, oxygenated (de)protonated analytes, dehydrogenated deprotonated analytes, and negative ion adducts) were quite similar to those obtained from DART alone. This indicates a lack of side reactions due to the corona discharge. The change in the relative intensities of individual analyte-related ions due to the combination of a corona discharge system with DART suggests that there is no effect of the abundant excited helium in the analyte ionization area on the fragmentation processes or enhancement of oxidation due to hydroxyl radicals HO˙. Furthermore, it was found that the corona-DART combination can be applied to the highly sensitive analysis of n-alkanes, in which the alkanes are ionized as positive ions via hydride abstraction and oxidation, independent of the type of alkane or the mass spectrometer used.

Ionization characteristics of amino acids in direct analysis in real time mass spectrometry.

The positive and negative ionization characteristics of 20 different α-amino acids were investigated using Direct Analysis in Real Time (DART) mass spectrometry. Almost all of the amino acids M were ionized to generate the (de)protonated analytes [M ± H](±)via proton transfer reactions with the typical background ions H3O(+)(H2O)n and O2˙(-) and resonant electron capture by M. The application of DART to amino acids also resulted in molecular ion formation, fragmentation, oxidations involving oxygen attachment and hydrogen loss, and formation of adducts [M + R](-) with negative background ions R(-) (O2˙(-), HCO2(-), NO2(-) and COO(-)(COOH)), depending on the physicochemical and/or structural properties of individual amino acids. The relationship between each amino acid and the ionization reactions observed suggested that fragmentation can be attributed to pyrolysis during analyte desorption as well as excess energy obtained via (de)protonation. Oxidation and [M + R](-) adduct formation, in contrast, most likely originate from reactions with active oxygen such as hydroxyl radicals HO˙, indicating that the typical background neutral species involved in analyte ionization in DART mass spectrometry contain HO˙.