| Popis: |
Glycans are molecules that contain multiple monosaccharides linked by glycosidic bonds and often bind to proteins or lipids to form various glycoconjugates. Rising awareness of the vital role that glycosylation plays in protein functioning governs the need for further advances in glycan analysis. Because free glycans lack chromophore or fluorophore properties, and do not ionize well, they are often derivatized, to facilitate their separation or detection, and to enhance the sensitivity of the analysis. Released glycans are usually derivatized using a fluorescent tag, which enables their optical detection in LC profiling. Some fluorescent labels can also improve ionization efficiency, which then facilitates MS detection. For this reason, there is a need to design fluorophores that will contribute more to the fluorescence and ionization of glycans and the need to quantify these contributions to improve glycan analysis methods in negative MS mode as these methods are more informative than methods involving positive MS mode, allowing for a less ambiguous elucidation of detailed glycan structures. Therefore, eleven fluorescent labels of different chemo-physical properties were chosen in order to study the influence of label hydrophobicity and presence of a negative charge on glycan ionization in negative MS mode. N-glycans released from IgG sample were labeled with one of the eleven labels, purified with HILIC-SPE and analyzed with HILIC-UHPLC-FLR- MS. The results show that hydrophobicity significantly contributes more to signal-to-noise intensity ratios in the mass spectrum than the negative ion charge in the case of negative MS mode. With this in mind, further research and development of fluorophores that will be suitable for glycan analysis in the negative MS mode are proposed. |
