Interplay of H‐Bonds with Aromatics in Isolated Complexes Identifies Isomeric Carbohydrates

Abstract

The tremendous isomeric diversity of carbohydrates enables awide range of their biological functions but makes the identification and study of these molecules difficult. We investigated the ability of intermolecular interactions to communicate structural specificity of carbohydrates to protonated aromatic molecules in non-covalent complexes,i solated and cooled in the gas phase.Our study revealed that small structural differences between carbohydrate isomers of any type,i ncluding enantiomers,a re accurately communicated by these interactions to aromatic molecules as detectable changes in their electronic excitation spectra. The specific response of the aromatics to the isomers of carbohydrates is fine-tuned by the interplay of the various involved non-covalent bonds.These findings enable the gas-phase identification and relative quantification of any isomers of oligosaccharides in their solution mixtures using the 2D UV-MS fingerprinting technique.Carbohydrates are by far the most abundant organic molecules present in living organisms. [1] Existing in myriads of structural forms,m any of which are isomeric,c arbohydrates may function as messenger and recognition molecules in signaling the type and the state of living cells in complex chains of biological processes. [2] Based on this,cancer cells can be identified, for instance,b yd etecting their specific membrane glycoproteins, [3] which may differ from those of the healthy cells by the isoform of the glycan only,w hile viruses and bacteria adhere to appropriate cells for invasion by selective binding to specific membrane glycans, [4] distinguishing them from many other structurally similar (for example, isomeric) saccharides.T hese and many other examples illustrate the importance of comprehensive structural studies of carbohydrates,i ncluding the identification of their isoforms. [5] Them onomeric structural units of carbohydrates exist in av ariety of stereoisomeric forms,s uch as d/l-enantiomers, epimers,a nd a/b-anomers.F or instance,acyclic aldohexo-pyranose has 5stereogenic centers,which implies an existence of 2 5 = 32 stereoisomers (for example, a-d-glucose, b-lgalactose,e tc.), although not all of them are essential in nature.I nc ontrast to amino acids and nucleotides,t hese isomeric units can interconnect through different OH groups, forming regioisomers,but also at multiple points,assembling into linear and branched structural isomers.N atural modifications (for example, N-acetylation) of different units further multiply the number of possible isoforms.Human milk alone, for instance,c ontains at least 200 different oligosaccharides, many of which are essential not only for supplying energy to newborns,but also for their antibacterial defense. [6] Theenormous isomeric diversity of carbohydrates makes their identification and study exceptionally challenging.Most common analytical methods of isomeric identifications of carbohydrates,such as chromatography [7] and, more recently, ion mobility, [8] demonstrate ah igh capability in separating small is...

Identification and Quantification of Any Isoforms of Carbohydrates by 2D UV-MS Fingerprinting of Cold Ions

“…The isomeric structure of a guest carbohydrate is communicated to the host aromatic molecule by noncovalent interactions, which induce changes in the UV absorption of the latter. In particular, an interplay of different types of H-bonds in isomeric complexes make the position of their UV absorption onset very sensitive to the isoform of a guest carbohydrate . We also found that the number of low-energy conformers is much lower for noncovalent complexes of glycans with aromatic molecules than for glycan–alkaline ion adducts, , which are often used for identifications of isomeric carbohydrates by IR spectroscopy.…”

“…2D UV-MS fingerprinting is a recently developed approach that is capable of highly accurate identification of isomeric biomolecules. [17][18][19][20][21][22] This analytical technique integrates UV photofragmentation (UVPD) spectroscopy of cryogenically cooled ions with broadband highresolution (e.g. Orbitrap-based) mass spectrometry.…”

“…Instead, we make use of non-covalent complexes of glycans with protonated aromatic molecules, such as phenylalanine, tyrosine and its decarboxylated analog tyramine. 22,[25][26][27] These complexes can be readily prepared by dissolving a glycan and an aromatic molecule in a standard for MS methanol/water solution.…”

“…Herein, we explore the use of the recently developed approach, named 2D UV-MS fingerprinting, for identification and quantification of isomeric carbohydrates. This technique integrates UV photofragmentation (UVPD) spectroscopy of cryogenically cooled ions with broadband high-resolution (e.g., Orbitrap-based) MS. − It measures abundance of all UV-induced photo fragments at once as a function of UV wavelength, thus providing a two-dimensional data array, named 2D UV-MS fingerprint, as a characteristic of an ion. Cryogenic cooling of mid-size biomolecular ions often enables vibrational resolution in their electronic spectra, which makes them highly distinguishable.…”

“…A straightforward application of 2D UV-MS fingerprinting to glycans is challenging because they do not absorb in UV. Instead, we make use of noncovalent complexes of glycans with protonated aromatic molecules, such as phenylalanine, tyrosine, and its decarboxylated analogue tyramine. ,− These complexes can be readily prepared by dissolving a glycan and an aromatic molecule in a standard for MS methanol/water solution. The complexes appear to be strong enough to survive electrospray ionization that brings them to the gas phase.…”