CROSS-LINKED POTATO STARCH AS AN AFFINITY ADSORBENT FOR BACTERIAL ALPHA-AMYLASE

Crosslinked potato starch was prepared as an affinity adsorbent for bacterial alpha-amylase. To this end, reaction parameters for crosslinking in an ethanol/water solvent were investigated. The degree of crosslinking, and consequently the suitability of crosslinked starch as an adsorbent for alpha-amylase, changed by altering these parameters. An increase in the degree of crosslinking of the adsorbent caused lower affinity for bacterial alpha-amylase which resulted in an unfavourable decrease in adsorption capacity and a favourable decrease in the degradation of the adsorbent by the enzyme. 1 g of a suitable adsorbent for bacterial alpha-amylase, prepared with an epichlorohydrin/glucose monomer ratio of 0.65 (starch concentration 150 mg/ml, ethanol/water ratio 2.0, sodium hydroxide/epichlorohydrin ratio 1.0), can adsorb 9.8 mg of an alpha-amylase from B. licheniformis at 4-degrees-C in 20 h. The equilibrium constant between bound and unbound alpha-amylase is dependent on the temperature. An effective desorption was possible by a shift to higher temperatures. Degradation values smaller than 0.1% were measured after an incubation of 1 h at 70-degrees-C in a desorption buffer with 20% glycerol. It was concluded that coulombic interactions and hydrogen bonds are of no or little importance in enzyme adsorption. Van der Waals forces, which are responsible for the large temperature effect, are the main forces in the interaction between alpha-amylase and crosslinked starch.