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In this paper, we describe the development of an efficient enzyme immobilization procedure based on the activation of epoxy carriers with glucosamine. This approach aims at both creating a hydrophilic microenvironment surrounding the biocatalyst and introducing a spacer bearing an aldehyde group for covalent attachment. First, the immobilization study was carried out using penicillin G acylase (PGA) from <i>Escherichia coli</i> as a model enzyme. PGA immobilized on glucosamine activated supports has been compared with enzyme derivatives obtained by direct immobilization on the same non-modified carriers, in the synthesis of different 3&#8242;-functionalized cephalosporins. The derivatives prepared by immobilization of PGA on the glucosamine-carriers performed better than those prepared using the unmodified carriers (i.e., 90% versus 79% cefazolin conversion). The same immobilization method has been then applied to the immobilization of two other hydrolases (neutral protease from <i>Bacillus subtilis</i>, PN, and bromelain from pineapple stem, BR) and one transferase (&#947;-glutamyl transpeptidase from <i>Bacillus subtilis</i>, GGT). Immobilized PN and BR have been exploited in the synthesis of modified nucleosides and in a bench-scale packed-bed reactor for the protein stabilization of a Sauvignon blanc wine, respectively. In addition, in these cases, the new enzyme derivatives provided improved results compared to those previously described.