The purpose of the present investigation is to enhance extracellular L-asparaginase production by subjecting isolated Pseudomonas aeruginosa strain F1 (Gene Bank Accession Number: JN412064.1) to improvement by random mutagenesis by ultra-violet (UV) irradiation, Nitrous acid and N-Methyl-N’-nitro-Nnitroso guanidine (NTG) treatment. Mutants were screened as L-asparaginase producers on the basis of enzyme activity in submerged fermentation. UVS-7 mutant obtained after 15-18 minutes of UV exposure showed 1.26 times more enzyme activity (76.6 IU/ml). UVS-7 mutant further mutated by Nitrous acid and UVS7-N-4 mutant obtained after 210-240 Sec Nitrous acid exposure showed 1.58 times increase in enzyme activity (98.41 IU/ ml). Then UVS7-N-4 mutant was treated with NTG. UVS7-N-4-N-8 mutant obtained after 210-240 minute NTG exposure showed 2.05 times increase in enzyme activity (124.28 IU/ml) in compared to the isolated wild Pseudomonas aeruginosa strain F1(60.64 IU/ml). The results indicated that UV radiation, Nitrous acid and NTG were effective mutagenic agents for strain improvement.Thus these findings have more impact on enzyme economy by enhancing the production of extracellular L-asparaginase for anticancer applications.