Heterofunctional epoxy support development for immobilization of fructosyltransferase from Pectinex® Ultra SP-L: batch and continuous production of fructo-oligosaccharides

Abstract

In this study, a heterofunctional carrier was obtained by modification of a macroporous polystyrene ion exchange resin with primary amino groups (Purolite® A109) with epichlorohydrin, in view of achieving covalent immobilization of fructosyltransferase (FTase) from a complex enzyme mixture, Pectinex® Ultra SP-L, that is responsible for the synthesis of functionally active fructo-oligosaccharides (FOS). A two-step immobilization protocol, comprising the physical adsorption of FTase performed at pH 4 and subsequent buffer exchange to promote the establishment of covalent bonds at pH 9, was proposed for obtaining a highly active immobilized preparation (243 IU g⁻¹ of support). Additionally, this protocol provided development of a preparation with 20 times more prominent expressed activity of FTase compared to the commercial preparation with predominant pectinase activity. The obtained immobilized preparation was further tested in batch and air-lift reactor systems for FOS synthesis, yielding 52.8% and 54.7% of FOS in total carbohydrates, respectively. Finally, the continuous production of FOS in the air-lift reactor was established for 7 days, with an average FOS yield of 52.5%. Accordingly, it is demonstrated that the immobilization process enabled the development of preparations with exceptional potential for industrial implementation.