Kinetics of an extracellular exo-inulinase production from a 5-flourocytosine resistant mutant of Geotrichum candidum using two-factorial design.

In the present study, eight different strains of Geotrichum candidum were isolated and screened for an extracellular exo-inulinase production using chemically enriched sucrose-mineral media. The isolate (Zool-3i) with a better enzyme activity (1.38 IU/ml) was subjected to induced mutagenesis using methyl methane sulphonate (MMS) and a mutant with an enzyme activity of 32.06 IU/ml was obtained. Further exposure to ethyl methane sulphonate (EMS) and ultraviolet (UV) radiations yielded a mutant exhibiting an improved activity of 39.34 IU/ml. The potential mutant was cultured overnight and plated on 5fc-YPR agar medium and thus made resistant against 5-flourocytocine. Over 50-fold enhancement in enzyme production (71.85 IU/ml) was achieved when the process parameters including incubation period (48 h), sucrose concentration (5.0 g/L), pH (6.0), inoculum size (2.0%, 16 h old) and urea (0.2%) were identified using Plackett-Burman design. On the basis of kinetic variables, notably Q(p) (0.723 U/g/h), Y(p/s) (2.036U/g) and q(p) (0.091U/g cells/h), the mutant MEU-5fc-6 was found to be a hyper producer of exo-inulinase (HS, LSD 0.045, p< or =0.05).