Quantitative analysis of the degree of silanization by the ninhydrin method and its application to the immobilization ofGL-7-ACA acylase and cellulolytic enzyme

A simple quantitative method to measure the degree of silanization was developed, based on the reaction of ninhydrin with the silanization reagent (3-aminopropyltriethoxysilane, 3-APTES). At low concentrations (0.001-0.005%, v/v) of 3-APTES, a good linearity was obtained when 3-APTES reacted with undiluted ninhydrin for 30 min. On the other hand, at high levels of 3-APTES, a linearity was obtained when 3-APTES reacted with 3-fold diluted ninhydrin for 20 min. The reliability of regression curves mentioned above was expressed as a regression coefficient (R²) of more than 0.99. Immobilization of different enzymes was introduced via silanization by using the 3-APTES in order to confirm the validity of the ninhydrin method. When yields for each step in the immobilization process were compared, yields of both glutaraldehyde and protein were found to have the same tendency to silanization. These results show that the ninhydrin method was suitable for quantitative analysis of silanization and that yields of immobilization could be pre-estimated by measuring silanization levels using the ninhydrin method.