Production and optimization of glucoamylases from plants and aspergillusniger for starch hydrolysis in a batch bioreactor

Thesis Abstract

Glucoamylases were produced from both plants and microorganisms and were optimized for starch hydrolysis in batch bioreactor. Amylase activity was monitored in germinating guinea corn seeds for seven days. Highest amylase activity was observed on days 3 and 7. A study of the amylopectin content of millet, guinea corn, cassava, corn and tigernut starch showed that tiger nut had the highest amylopectin content while cassava starch had the lowest. Moist amylopectin frommillet, guinea corn, cassava, corn and tiger nut starch were exposed on the shelf to triger microbial growth. Luxurial growths were noticed on amylopectin from guinea corn, tigernut and cassava starch. Pure isolates were obtained by subculturing and identified as Aspergillus niger. A 14 day fermentation study to determine the optimal production time using the organism and amylopectin from guinea corn, tigernut and cassava starch was carried out. The fermentation studies showed a two peak profile for each amylopectin used.The first on day 3 or 4, while the second peak on day 11 and 12, respectively. Large scale production of glucoamylase was carried out on these days of highest enzyme production. Glucoamylase activities from both germinating guinea corn seeds and Aspergillusniger were enhanced by calcium (Ca2+), zinc (Zn2+), cobolt (Co2+), iron (Fe2+) and manganespre (Mn2+) ionbut Lead ion (Pb2+) completely inactivated the enzymes. The Michaelismenten constant (Km) and the maximum velocity (Vmax)obtained from Lineweaver-Burk plot of initial velocity data at different substrate concentrations showed high affinity of the glucomylases for their substrates. The optimal pH and temperature of glucoamylases from both germinating seeds and Aspergillusniger were in the range of 4.5-8.5 and 45-60 ËšC, respectively.The glucoamylases were screened for α and β glucosidase activities and glucoamylase obtained on day 7 from germinating guinea corn seeds (GluGERGC7) and that obtained on days 11 and 12 from Aspergillusniger grown in broth containing amylopectin from cassava and tiger nut starch (GluAgCSV11) and (GluAgTN12), respectively were found to exhibit high α glucosidase activity. The rate of substrate utilization or the efficiency of batch bioreactor at the predetermined optimal conditions was predicted using Kmand Vmaxobtained from a modified form of Michaelis-Menten’s equation and were found within the range of 40- 460mg/ml and 0.811-50 µmol/min, respectively using cassava, guinea corn and tiger nut starch as substrates. The results suggest that the glucoamylases obtained from both germinating guinea corn seeds and Aspergillusniger possess the qualities of biotechnological applications in which the optimal conditions could be predicted.