Statistical optimization of L-asparaginase production by using Fusarium solani

Abstract

Background and objective The enzyme l-asparaginase is important as a therapeutic agent in the treatment of acute lymphocytic leukemia. It has been observed that microorganisms such as yeast and filamentous fungi such as Aspergillus spp., Penicillium spp., and Fusarium spp. are commonly produced asparaginases with fewer adverse effects. Fusarium solani was selected to be the most potent microbial isolate for l-asparaginase production. The factors controlling l-asparaginase production, such as containing media, were implemented to increase the yield of l-asparaginase. Statistical experimental design such as the Plackett–Burman enables finding out the most effective factors that increase the yield of l-asparaginase. Materials and methods Aspergillus rubber, Aspergillus terreus, Epico niger, Penicillium cyclopium, and F. solani were chosen for screening the l-asparaginase activity using phenol red – a pH indicator. l-Asparagine and the modified Czapek Dox media were tested for the production of l-asparaginase from F. solani under different incubation times. To optimize l-asparaginase production by using F. solani, the combined effect of seven variables (l-asparagine, sucrose, NaNo3, K2HPO4, MgSO4, KCl, and time) were assessed using the central composite design. Results and conclusion All the four fungal strains screened for l-asparaginase activity showed positive results in the rapid plate assay method. The medium employed contained asparagine with phenol red, and after incubation, pink zones around the colonies were observed. F. solani was a potential source for a high yield of l-asparaginase enzyme and high substrate specificity. Modified Czapek medium II was the most suitable for l-asparaginase, showing an activity of 121 U/ml. The result on l-asparaginase activity according to the screening Plackett–Burman experiments gives a medium composed of the following (g/l): sucrose (1); l-asparagine (0.5); KH2PO4. (0.75), MgSO4 (0.7); H2O (0.72); KCl (0.72); (NH4)2SO4 (11.7) at an incubation time of 3 days. For further investigation, x1 K2HPO4, x2 sucrose, and x3 time were found to be the most significant variables affecting l-asparaginase activity. The second optimization step was to identify optimal values of the three factors that bring about maximum l-asparaginase activity, using the central composite designed experiment. The results showed that K2HPO4 (2.5 g/l), sucrose (4 g/l), and the time (8 days) were critical in the production of l-asparaginase.