Glutaraldehyde cross-linking of immobilized thermophilic esterase on hydrophobic macroporous resin for application in poly(ε-caprolactone) synthesis

Abstract

The immobilized thermophilic esterase from Archaeoglobus fulgidus was successfully constructed through the glutaraldehyde-mediated covalent coupling after its physical adsorption on a hydrophobic macroporous resin, Sepabeads EC-OD. Through 0.05% glutaraldehyde treatment, the prevention of enzyme leaching and the maintenance of catalytic activity could be simultaneously realized. Using the enzymatic ring-opening polymerization of ε-caprolactone as a model, effects of organic solvents and reaction temperature on the monomer conversion and product molecular weight were systematically investigated. After the optimization of reaction conditions, products were obtained with 100% monomer conversion and Mn values lower than 1010 g/mol. Furthermore, the cross‑linked immobilized thermophilic esterase exhibited an excellent operational stability, with monomer conversion values exceeding 90% over the course of 12 batch reactions, still more than 80% after 16 batch reactions.

Scheme 1

Schematic representation for constructing the cross-linked immobilized thermophilic esterase via glutaraldehyde chemistry.

Figure 1

Effect of glutaraldehyde concentration on the specific activity of immobilized enzymes.

Figure 2

Enzymatic ring-opening polymerization of ε-caprolactone using different immobilized enzymes. The reactions were carried out using 60 mg immobilized enzymes, 200 μL ε-caprolactone and 600 μL toluene at 80 °C for 72 h.

Figure 3

Immobilized enzyme-catalyzed ring-opening polymerization of ε-caprolactone at different temperatures. The reactions were carried out using 60 mg immobilized enzymes, 200 μL ε-caprolactone and 600 μL toluene for 72 h.

Figure 4

Monomer conversion and M_n values for a series of consecutive batch reactions conducted using 60 mg immobilized enzyme as catalyst; on completion of one reaction, the catalyst was washed and recycled for use in the next. The reactions were carried out using 60 mg immobilized enzymes, 200 μL ε-caprolactone and 600 μL toluene at 80 °C for 72 h.