Typical modes of bond cleavages of organic compounds in mass spectrometry are briefly summarized. Although these fragmentation rules can be quite useful for interpreting mass spectra of simple compounds, application to structurally complex molecules that contain multiple hetero atoms such as nitrogen or oxygen becomes increasingly difficult, because the exact location of an unpaired electron or positive or negative charges becomes obscure in precursor ions.
About a decade ago, we proposed “a rule of mass shift,” which correctly predicts the m/z for observed peaks corresponding to singly charged even-electron fragment ions. The basis of the rule postulates that ions observed as peaks in an ordinary mass spectrum should be sufficiently stable to survive during the flight path in a mass spectrometer.
The important recognition is that each atom in a stable ion should be in an ordinary valence state, and no free valence should be allowed. Therefore, if the cleavage of a bond leads to an ion with an unstable structure, some structural changes must take place in order for the ion to be observed in the mass spectrum. Such structural changes can be the addition of hydrogen atom(s) and/or a proton for positive ions, and the addition of a hydrogen atom and/or the elimination of two hydrogen atoms in the case of negative ions. These required structural changes in each case are schematically depicted and discussed in detail.
Two typical examples are shown, in which m/z’s of the observed peaks are correctly predicted. The scope and limitations, as well as the significance of the rule for analyzing fragmentations in organic mass spectrometry are also discussed.
A sheet-like ultraviolet (UV) probe laser is used to investigate the ejection and propagation of ion packets of matrix CHCA, which are produced by matrix-assisted laser desorption and ionization (MALDI). Laser irradiation of the expanding MALDI plume induced photodissociation of the CHCA-related ions, which existed in a sheet-like volume, leading to their absence in their MALDI signal profiles. The MALDI spectra were measured under varying conditions: the temporal delay of the lasers and the distance of the sheet-like probe laser from the MALDI sample surface. It was found that the center of the (CHCA)H+ packets were ejected at 46±11 ns after MALDI laser irradiation, while the (CHCA)2H+ packets were ejected at 64±12 ns, regardless of the magnitude of acceleration static high-voltage in 3.5–5.5 kV. This suggests that (CHCA)2H+ is formed by a proton transfer reaction from (CHCA)H+ to (CHCA)2 in the heated condensed phase and/or near the surface. This study represents the first experimental determination of ion ejection time in the MALDI process, which is also applicable to other species in the MALDI plume.
In the urine of a Niemann–Pick disease type C (NPC) patient, we have identified three characteristic intense peaks that have not been observed in the urine of a 3β-hydroxysteroid-Δ5-C27-steroid dehydrogenase deficiency patient or a healthy infant and adult. Based on accurate masses of the protonated molecules, we focused on two of them as candidate NPC diagnostic markers. Two synthesized authentic preparations agreed with the two compounds found in NPC patient urine in regard to both chromatographic behavior and accurate masses of the deprotonated molecules. Moreover, the isotopic patterns of the deprotonated molecules, twin peaks unique to the sulfur-containing compounds appearing in their second isotope positions, and accurate masses of product ions observed at m/z 97 also agreed between the target compounds and authentic preparations. We identified the two compounds as the sulfated cholesterol metabolites as 3β-sulfooxy-7β-hydroxy-5-cholen-24-oic acid and 3β-sulfooxy-7-oxo-5-cholen-24-oic acid. These two compounds represent more promising candidate diagnostic markers for NPC diagnosis than three other candidates that are multiple conjugates of cholesterol metabolites, 3β-sulfooxy-7β-N-acetylglucosaminyl-5-cholen-24-oic acid and its glycine and taurine conjugates, although we have reported an analytical method for determining the urinary levels of these compounds using liquid chromatography/electrospray ionization tandem mass spectrometry, because of their lack of N-acetylglucosamine conjugation.
Mass spectrometry imaging (MSI) with ambient sampling and ionization can rapidly and easily capture the distribution of chemical components in a solid sample. Because the spatial resolution of MSI is limited by the size of the sampling area, reducing sampling size is an important goal for high resolution MSI. Here, we report the first use of a nanopipette for sampling and ionization by tapping-mode scanning probe electrospray ionization (t-SPESI). The spot size of the sampling area of a dye molecular film on a glass substrate was decreased to 6 μm on average by using a nanopipette. On the other hand, ionization efficiency increased with decreasing solvent flow rate. Our results indicate the compatibility between a reduced sampling area and the ionization efficiency using a nanopipette. MSI of micropatterns of ink on a glass and a polymer substrate were also demonstrated.
Mycolic acids (MAs) are characteristic components of bacteria in the suborder Corynebacterineae, such as Mycobacterium. MAs are categorized into subclasses based on their functional bases (cyclopropane ring, methoxy, keto, and epoxy group). Since MAs have heterogeneity among bacterial species, analyzing of MAs are required in the chemotaxonomic field. However, their structural analysis is not easy because of their long carbon-chain lengths and several functional groups. In this study, total fatty acid (FA) methyl ester (ME) fraction of M. tuberculosis H37Rv was analyzed by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOFMS) with a spiral ion trajectory (MALDI spiral-TOFMS). The distributions of carbon-chain length and their relative peak intensities were confirmed with those obtained by analysis of each subclass fraction which was separated from total FA ME fraction using thin-layer chromatography (TLC). The observed major peaks were reliably assigned as MAs owing to the high mass accuracy (error<3 ppm). The types of MA subclasses, their distributions of carbon-chain lengths, their relative peak intensities, and the ratio of even- and odd-numbered carbon-chain MAs for the total FA ME fraction were consistent with those of MA subclass fractions. To visualize whole MAs, contour maps of relative peak intensities for whole MAs were created. The contour maps indicated the MA subclasses and their distributions of carbon-chains with relative peak intensities at a glance. Our proposed method allows simple characterization in a short time and thus enables the analysis of large numbers of samples, and it would contribute to the chemotaxonomy.