초록
<P><B>Abstract</B></P> <P>A tri-enzyme biocatalyst “combi-CLEAs” with starch hydrolytic activity was prepared from commercially available alpha amylase, glucoamylase and pullulanase preparations by aggregating enzymes with ammonium sulphate followed by cross-linking formed aggregates for 4.5h with 40mM glutaraldehyde. The effects of precipitant type and cross-linking were studied and the biocatalyst was characterized. Scanning electron microscopy analysis showed that tri-enzyme biocatalyst was of spherical structure. For one pot starch hydrolytic activity, shift in optimum pH from 6 to 7 and temperature from 65 to 75°C were observed after co-immobilization of enzymes. After one pot starch hydrolysis reaction in batch mode, 100%, 60% and 40% conversions were obtained with combi-CLEAs, separate CLEAs mixture and free enzyme mixture, respectively. Co-immobilization also enhanced the thermal stability of enzymes. Finally, the catalytic activity of enzymes in combi-CLEAs during one pot starch hydrolysis was well maintained up to five cycles without performance changes.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Development of combi-CLEAs of alpha amylase, glucoamylase and pullulanase. </LI> <LI> Combi-CLEAs improved optimal conditions for one pot starch hydrolysis. </LI> <LI> Co-immobilization as combi-CLEAs enhanced thermal stability of enzymes. </LI> <LI> One pot starch hydrolysis in batch mode using combi-CLEAs was studied. </LI> <LI> Reusability of enzymes during one pot starch hydrolysis was determined. </LI> </UL> </P>