Semiquantitative analysis of isomeric oligosaccharides by negative‐ion mode UV‐MALDI TOF postsource decay mass spectrometry and their fragmentation mechanism study at N‐acetyl hexosamine moiety

Abstract

Postsource decay (PSD) spectra of isomeric neutral lactooligosaccharide mixtures were measured from the chlorinated molecules [M + Cl]- by negative-ion mode ultraviolet matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (UV-MALDI TOF MS) to estimate quantitatively the mixing ratios in their mixtures. The PSD ions specific to each isomeric structure were used to distinguish the linkage and branching isomers, and the molar ratios of the isomers were estimated from their ion abundances. The relative ion abundances changed linearly in the PSD spectra of the mixtures of the isomers as their molar ratio was varied in the analyte solutions. Therefore, the molar ratios of the isomers in the analyte mixtures could be estimated semiquantitatively. In addition, we studied their fragmentation mechanisms in N-acetyl hexosamines such as GlcNAc, which enabled us to quantitatively analyze the structures of the isomers of lactooligosaccharides. The conjugated systems elongate in the chemical species of the Z-type fragmentation on the 3-linked GlcNAc owing to the acetoamido groups at the C-2 positions, which made the chemical species of the Z-type ions stable. The glycosyl bonds of the front of GlcNAc cleaved easily as a C-type fragmentation because the negative charge at the anomeric position could be delocalized to the carbonyl oxygen atom at the acetoamido group of GlcNAc. These factors caused the stabilization of the chemical species of the C/Z fragment ions produced by the double cleavage around GlcNAc.

MALDI linear-field reflectron TOF post-source decay analysis of underivatized oligosaccharides: Determination of glycosidic linkages and anomeric configurations using anion attachment

“…On the contrary, when subjected to PSD fragmentation in the negative ion mode, prominent cross-ring cleavages are usually produced along with glycosyl bond cleavages. PSD of both deprotonated molecules and anionic adducts of oligosaccharides has been performed and some success for differentiation of linkage positions as well as anomeric configurations in oligosaccharides has been shown [35][36][37][38][43][44][45].…”

Assignments of B-Type Fragments in Post-Source Decay of Negative-Ion Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry of Neutral Lactooligosaccharides

“…1)῍5), 7) On the basis of these results, we discarded the possibility of the assignment to a Y-type fragmentation.…”

Challenges in Separations of Proteins and Small Biomolecules and the Role of Modern Mass Spectroscopy Tools for Solving Them, as Well as Bypassing Them, in Structural Analytical Studies of Complex Biomolecular Mixtures

“…Second important property of commercially available purified compounds is their known amount. Thus, appropriate calibration curves/mixtures may be constructed for advanced MALDI MS studies as well [82,83]. Although LC-MS combined with bioinformatic tools represents a state-of-the-art screening tool, state-of-the-art MALDI MS methods are more appropriate for detailed structural characterization of previously uncharacterized biomolecules.…”

“…However, for more detailed analyses, and especially quantitative ones, appropriate calibration mixtures with known amounts of individual structures are required. An example of such study is shown in Figure 19 [83], where various mixtures of three isomers, lacto-N-monofucosylpentaose I, II and III were analyzed, based on which the respective calibration curves were constructed. Although these proof-of-concept studies show encouraging data, in order to be able to utilize this approach reliably, all~500 HMO structures must be commercially available in a purified form.…”