Determination of distinctive carbohydrate signatures obtained from the Aeromonas hydrophila (chemotype II) core oligosaccharide pinpointing the presence of the 4‐O‐phosphorylated 5‐O‐linked Kdo reducing end group using electrospray ionization quadrupole orthogonal time‐of‐flight mass spectrometry and tandem mass spectrometry

Abstract

The electrospray quadrupole orthogonal time-of-flight mass spectrometric (ESI-QqTOF-MS) structural elucidation of the core oligosaccharide of Aeromonas hydrophila (chemotype II) lipopolysaccharide has been investigated and it was demonstrated that it contained an 4-O-phosphorylated Kdo reducing end group, which was glycosylated by the remaining outer core oligosaccharide through its O-5 position. After releasing the core oligosaccharide from the native LPS with acid, the phosphorylated Kdo residue eliminated phosphoric acid, to produce a core oligosaccharide containing a mixture of diastereomeric 4,8- and 4,7-anhydro-alpha-keto acids and an open-chain olefinic Kdo residue. The characteristic glycone sequence was elucidated by collision-induced dissociation tandem mass spectrometry (CID-MS/MS) of the protonated molecule of the native core oligosaccharide. In addition, the analysis of the Hakomori permethylated core oligosaccharide was carried out by electrospray ionization quadrupole orthogonal time-of-flight mass spectrometry (ESI-QqTOF-MS) and matrix-assisted laser desorption/ionization (MALDI)-QqTOF-MS analyses. The presence of more than nine isobaric isomers of this core was detected. The CID-MS/MS analysis of the various protonated permethylated core oligosaccharide molecules showed a similar and diagnostic fragmentation pattern. The over-methylation of the permethylated core oligosaccharide containing either the 4,7- or the 4,8-anhydro-alpha-keto acid unit and the open-chain olefinic Kdo unit was reported. It was realized that the extra minor satellite signals obtained in the ESI-QqTOF-MS and MALDI-TOF-MS analyses were dimethyl sulfoxide (DMSO) stable covalent addition products, which have occurred by a Michael addition on the 4,8-Kdo exocyclic double bond. The occurrence of this series of covalent addition products during the MS analysis of a permethylated core oligosaccharide should be considered as 'carbohydrate-distinctive signatures' establishing and confirming the presence of a 4-O-phosphorylated-5-O-linked Kdo reducing end group.

Analysis of carbohydrates and glycoconjugates by matrix‐assisted laser desorption/ionization mass spectrometry: An update for 2009–2010

“…These ions were determined to be dimethylsulfoxide (DMSO) covalent addition products resulting from the Michael addition of the dimsyl anion on the C‐2–C‐3 double bond of a 4,8‐anhydro Kdo ( 1/13 ) residue followed by an addition of a proton on the double bond. Corresponding ions were also seen in methylations performed using the NaOH technique and represent the first characterization of these addition products (Sioud et al, ).…”

Synthesis and Antibacterial Evaluation of Some New Fused Cyclic Sulfoxides

“…Michael addition of DMSO to permethylated core of oligosaccharides was used in the formation of covalent DMSO products adducts as shown below (Sioud et al, 2010).…”

Distinguishing isomeric aldohexose-ketohexose disaccharides by electrospray ionization mass spectrometry in positive mode

“…Tandem mass spectrometry (MS/MS) is particularly useful, because it is simple, fast and sensitive. Collision‐induced dissociation (CID) is the most common fragmentation technique; however, it provides limited structural information at low collision energy . Recently, other alternative techniques, such as post‐source decay, electron capture dissociation, electron transfer dissociation, infrared multiple‐photon dissociation and higher collision energy dissociation (HCD) have been proposed to distinguish oligosaccharide isomers.…”