MALDI-MS analysis of sialylated N-glycan linkage isomers using solid-phase two step derivatization method

Area of Science:

• Glycomics
• Analytical Chemistry
• Biochemistry

Background:

• Sialic acids are terminal glycans with distinct biological roles based on α(2,3) or α(2,6) linkages.
• Existing methods for discriminating sialic acid linkages, like ethyl esterification, face challenges with derivative stability and byproducts.

Purpose of the Study:

• To develop a robust method for distinguishing and quantifying α(2,3)- and α(2,6)-linked sialic acids.
• To improve upon the limitations of prior carboxyl derivatization techniques for sialic acid linkage analysis.

Main Methods:

• A novel two-step solid-phase derivatization strategy was employed.
• α(2,6)-linked sialic acids were converted to ethyl esters, and α(2,3)-linked sialic acids to N-methyl amides.
• Optimized conditions enabled selective derivatization and subsequent analysis.

Main Results:

• The developed method achieved accurate relative quantification of N-glycans and their sialylated linkage types.
• This approach demonstrated superior performance compared to the ethyl esterification method.
• The strategy was successfully applied to analyze N-glycans in a biosimilar antibody drug and human serum samples.

Conclusions:

• The solid-phase derivatization method offers enhanced selectivity and stability for sialic acid linkage analysis.
• This technique provides accurate glycan quantification, aiding in the differentiation of linkage types.
• The method holds significant potential for biomarker discovery and applications within the pharmaceutical industry.