Immobilized Pichia pastoris in alginate beads for continuous culture

Conditions for immobilizing Pichia pastoris GS115pPIC9g by calcium alginate gel entrapment were determined. After mixture immobilization with 1.5 × 10⁹ CFU/ml, free biomass concentration decreases from 2.3 × 10⁴ to 2.2 × 10³ CFU/ml when alginate concentration rises from 20 to 40 g/l. Bead diameter with cells was 3.08 ±0.18 mm. Immobilized biomass reutilization power was demonstrated with repeated replacement cultures. Glycerol uptake in a continuous culture with 100 ml of immobilized biomass with 500 ml of medium (36.3 g/l glycerol), reached the stationary state the third day on 52.9 ±1.5 g/d, with a dilution rate of 0.12 h^-1. Biomass concentration at stationary phase was 10 ⁷-10⁸ CFU/ml for free cells and 10⁹ CFU/ml for immobilized ones. The distribution pattern of entrapped cells in alginate beads is showed by scanning electron microscopy (SEM) and the morphology of free and immobilized cells by optical microscopy.