Abstract :

Actinomycetes are a group of filamentous, Gram-positive bacteria widely distributed in soil and recognized as prolific producers of bioactive secondary metabolites, including antibiotics and enzymes of therapeutic importance. Among these, L-asparaginase is a clinically significant enzyme used as an antineoplastic agent in the treatment of acute lymphoblastic leukemia. The enzyme catalyzes the hydrolysis of L-asparagine into L-aspartic acid and ammonia, thereby depriving tumor cells of an essential amino acid required for their growth and survival. Although L-asparaginase derived from Escherichia coli and Erwinia carotovora is commercially available, these bacterial sources often cause hypersensitivity and other side effects. Hence, there is a growing need to explore alternative microbial sources such as actinomycetes, which can produce the enzyme with better therapeutic efficiency and fewer side effects. This study aims to identify potent L-asparaginase-producing actinomycetes and their production under submerged fermentation conditions. The   study was designed with four major objectives, to isolate actinomycetes from soil samples, to perform qualitative and quantitative assays for L-asparaginase production, and to evaluate the production of L-asparaginase by submerged fermentation using boiled rice water as the substrate. Soil sample was collected, and actinomycetes were isolated using standard microbiological techniques. The qualitative screening of isolates was carried out using a plate assay method to identify potential L-asparaginase-producing strains based on the formation of color zones. Quantitative estimation of enzyme activity was performed by measuring the amount of ammonia released during enzymatic reaction. For enzyme production, submerged fermentation was employed, utilizing boiled rice water as a nutrient-rich, cost-effective substrate to enhance enzyme yield.

---

Keywords :

Actinomycetes, L- Asparaginase, Qualitative and quantitative assay, submerged fermentation.

---

References :

1. El-Refai, H. A., Abdel Naby, M. A., Gaballa, A. A., El-Araby, M. H., & Ammar, H. S. (2020). L-Asparaginase from microbial sources: Production, optimization, and applications. Journal of Genetic Engineering and Biotechnology, 18(1), 1–12.
2. Sasidharan, S., & Priya, K. (2021). Actinomycetes as a promising source of novel L-asparaginase with improved biochemical properties. Biotechnology Reports, 30, e00615.
3. Manisha, M., Kumar, S., & Singh, P. (2022). Exploration of Streptomyces species for extracellular L-asparaginase production and characterization. Journal of Applied Microbiology, 133(1), 45–56.
4. Verma, N., & Kumar, K. (2018). Thermostable L-asparaginase from actinomycetes: Production, characterization, and industrial relevance. International Journal of Biological Macromolecules, 117, 1084–1092.
5. Sheik, G. B., M. S. Maqbul, G. S. S., and R. M. S. Isolation and characterization of actinomycetes from soil of ad-dawadmi, saudi arabia and screening their antibacterial activities. International Journal of Pharmacy and Pharmaceutical Sciences, vol. 9, no. 10, Oct. 2017, pp. 276-9.
6. Noura EE, Hassan M, Nancy ME, Sara ME. (2015) Optimization of culture conditions for production of the anti- leukemic glutaminase free Lasparaginase by newly isolated Streptomyces olivaceus NEAE-119 using response surface methodology. BioMed Res Int 2015; 2015:627031.
7. Ashok, A., Doriya, K., Rao, J. V., Qureshi, A., Tiwari, A. K., & Kumar, D. S. (2019). Microbes producing L-asparaginase free of glutaminase and urease isolated from extreme locations of Antarctic soil and moss. Scientific Reports, 9 (1), 1423.
8. Deshpande, N., Choubey, P., & Agashe, M. (2014). Studies on optimization of growth parameters for L-asparaginase production by Streptomyces ginsengisoli. The Scientific World Journal, 2014, Article ID 895167.
9. Narayana KJP et al. (2007) L-asparaginase production by Streptomyces albidoflavus. Indian J.Microbiol. 48(3): 331-336.
10. Mostafa SA and Salama M S (1979) L-asparaginase producing Streptomyces from soil of Kuwait. Zentralbl Bakterol 134:325–334.
11. Lopes Fernanda Cortez, Ligabue-Braun Rodrigo. 2021., Agro-Industrial Residues: Eco-Friendly and Inexpensive Substrates for Microbial Pigments Production Frontiers in Sustainable Food Systems, VOL 5: 2021.589414.