6. Linkage Pattern Analysis

Methylation analysis, a powerful tool for primary structure characterization, has been used for decades and is still widely used nowadays (Chandra et al. in Carbohydrate Research 344(16):2188–2194, 2009; Pereira et al in Phytochemistry 71(17–18), 2132–2139 2010; Wu et al. in Food Hydrocolloids 23(6), 1535–1541 2009). As shown in Fig. 6.1, it includes four main steps: (1) methylation reaction using methyl iodide to convert all free hydroxyl groups of the polysaccharide molecules into methoxy groups; (2) acidic hydrolysis (usually by TFA) to convert the polymer into monomer; (3) reduction (using sodium borodeuteride) and acetylation (by acetic anhydride) to give volatile products: partially methylated alditol acetates (PMAA); (4) GC-MS analysis to identify and quantify the produced PMAAs (Dell in Meth Enzymol 193:647–660, 1990; Liu in Food carbohydrates: chemistry, physical properties, and applications. Taylor and Francis, Boca Raton, pp 309–355, 2005). The linkage patterns for each monomer and the molar ratios can be obtained using this method (Ciucanu & Kerek in Carbohydr Res 131(2):209–217, 1984).