Investigation of the enzyme Bacillus agaradhaerens Cel 5a as an analytical tool in mass spectral characterisation of methylcelluloses

Two methylcelluloses (MC) were investigated by partial hydrolysis followed by direct infusion electrospray ionisation mass spectrometry (ESI-MS). Partial hydrolysis was achieved either enzymatically or by acid hydrolysis. Enzyme hydrolysis was performed using the endoglucanase Bacillus agaradhaerens Cel 5a. Three different hydrolysis buffer solutions were investigated in order to determine which was most compatible with the subsequent MS analysis. ESI-MS experiments showed that when using enzymatic hydrolysis as an analytical tool for characterisation of modified cellulose one has to be watchful for the investigated polymer to have a sufficient amount of available cleavage sites in order to ensure that the original ends of the polymer do not influence the results. MS2 experiments confirmed that the detected products originated from methylcellulose and provided structural information on the investigated products. MS data proved that B. agaradhaerens Cel 5a is able to cleave β1 → 4 glycosidic linkages adjacent to a permethylated glucosyl unit. MS2 data showed that methylation on the C-2 and C-6 hydroxyl groups of the glucosyl unit on the non-reducing side of the cleavage site hindered the enzyme.