Immobilization of β-galactosidase retained in yeast: adhesion of the cells on a support

Summary

Treatment of Kluyveromyces cells with several polycations partially neutralized their negative charge, even after washing; chitosan at 0.5 mg/ml rendered the zeta potential positive. Adhesion of the cells to various supports (glass, polycarbonate, polystyrene) was promoted by treating the cells or the support with chitosan; this confirms the importance of electrostatic interactions in the adhesion process. When the support was treated, a dense and regular monolayer of adhering cells was obtained; when the cells were treated the adhering layer was more heterogeneous due to floc formation. Cell ghosts of K. lactis were prepared by CHCl₃/ethanol permeabilization and glutaraldehyde treatment. Treating the ghosts with chitosan provided immobilization of 1.9 to 7.3×10⁶ ghosts per cm² according to the support; treating the support allowed to obtain 5.5 to 5.7×10⁶ ghosts per cm². The lactase activity of the immobilized ghosts was found to be comparable to that of ghosts in suspension, i.e. in the range of 0.9 to 2.2 U/mg dry cell weight, at 30°C, corresponding to 0.02 to 0.14 U/cm² of support. A test in microreactor with a lactase solution showed that 50% of the initial activity was still found, after about 25 days of continuous operation at 30°C.