Direct recovery of β-galactosidase from yeast homogenate: Comparison of expanded bed and stirred fluidized bed ion-exchange chromatography.

This study investigates the purification of β-D-galactosidase from disrupted Kluyveromyces marxianus using STREAMLINE DEAE adsorbent in expanded bed adsorption (EBA) and stirred fluidized bed adsorption (SFBA) systems. A key feature of this work is the direct comparison of feed complexity under industrially relevant conditions. The clarified lysate (50% w/v) contained approximately 1-2% residual cells and cell debris, representing a semi-clarified industrial feedstock, whereas the unclarified lysate consisted of a highly concentrated suspension (50% w/v solids). Batch and column experiments were conducted to optimize adsorption and elution conditions, including pH, the effects of cell debris, adsorption kinetics, equilibrium isotherms, superficial velocity, ionic strength, and buffer pH. Adsorption kinetic and equilibrium analyses showed that the Avrami kinetic model and the Sips isotherm model best described the experimental data, suggesting a multi-stage adsorption mechanism on a heterogeneous adsorbent surface. The optimized conditions were subsequently applied to the EBA and SFBA systems. Breakthrough analysis indicated that the presence of cell debris caused slightly earlier breakthrough; however, high adsorption efficiencies were maintained. In the EBA system, β-galactosidase adsorption efficiency reached 95.41% for the unclarified feedstock. In the SFBA system, adsorption efficiencies of 94.47% and 93.82% were achieved for the clarified and unclarified lysates, respectively. Two-step elution using 0.5 M NaCl (pH 7.0) at 150 cm/h yielded recoveries of 45-48% with purification folds of 4-5. The SFBA system exhibited superior tolerance to viscous, particulate-rich feedstocks and maintained stable bed expansion, enabling efficient direct enzyme capture without prior clarification.