Isolation, screening, optimization and production of Anti-tumor L-Asparaginase by fungi from karwar coastal region
- 1Department of Biotechnology, Government Arts and Science College Karwar-581301, Karnataka, India
- 2Department of Biotechnology, Government Arts and Science College Karwar-581301, Karnataka, India
Res. J. Recent Sci., Volume 6, Issue (3), Pages 1-7, March,2 (2017)
L-asparaginase is an extracellular enzyme that converts L-asparagine to L-Aspartic acid and has gained considerable attention in the recent years. In the present study, soil samples were gathered from different areas in and around the Karwar regions, Karnataka, India. Several fungal species were isolated and identified using standard manuals, screened primarily for the production of L-Asparaginase on Czapek’s Dox medium containing L- Asparagine as main source of carbon. A total of 50 fungal cultures were isolated from soil. The fungal isolates obtained were selected by plate assay method for screening of potential L-Asparaginase production on Modified Czapek Dox’s (mCD) medium. Enzyme production was carried out by submerged fermentation process and was performed by using mCD liquid media. Quantitative enzyme assay was performed to determine the rate of hydrolysis of L-Asparagine by measuring the liberated ammonia by nesslerization. From the 50 total isolates 10 fungal cultures showed L-Asparaginase activities. The cultures were identified as Penicillum species, Basidiomycetes species. Aspergillus species, Mucor species. Fusarium species. Among the above species Aspergillus sp showed potential L-Asparaginase production (155U/ml). An attempt is made to optimize the cultural conditions for the production of potential L-Asparaginase by using submerged fermentation. Different temperature (150C, 250C, 350C, 450C) different pH (3.5, 5.5, 7.5, 9.5, and 11.5) different carbon and nitrogen source were used. The highest amount of enzyme production was observed at pH 7.5(155 U/ml) and temperature at 35°C (160 U/ml), among the various carbon sources dextrose promoted maximum enzyme activity (176 U/ml) and highest activity was obtained when nitrogen source ammonium sulphate was used (185 U/ml).
- Jemal A., Bray F., Center M.M., Ferlay J., Ward and E. Forman D. (2011)., Global cancer statistics., CA Cancer J Clin., 61(2):69-90. doi: 10.3322/caac.20107. Erratum in: CA Cancer J Clin. 2011 Mar-Apr; 61(2):134.
- Weinberg R.A. (1996)., How cancer arises., Scientific American, 275(3), 62-71.
- Kotzia G.A. and Labrou N.E. (2007)., L-Asparaginase from Erwinia chrysanthemi 3937, Cloning, Expression and Characterization., Journal of Biotechnology, 127(4), 657-669.
- Berenbaum M.C., Ginsburg H. and Gilbert D.M. (1970), Effects of L-asparaginaseon lymphocyte target cell reactions in Vitro., Nature, 227(5263), 1147-1148.
- Broome J.D. (1963)., L-asparaginase EC-II from Escherichia coli.Some substrate specificity characteristics., Biochemistry 8, 3766-3772.
- Wriston J. and Yellin T. (1973)., L-Asparaginase: A Review., Adv Enzymol Relat Areas Mol Biol, 39, 185-248.
- Yellin T. and Wriston J. (1973)., Purification and Properties of Guinea Pig Serum Asparaginase., Biochemistry, 5(5), 1605-1612.
- Capizzi R.L., Poole M., Cooper M.R., Richards F., Stuart J.J., Jakson D.V., White D.R., Spurr C.L., Hopkins J.O. and Muss H.B. (1984)., Treatment of poor risk acute leukaemia with sequential hig-done ARA-C and asparaginase., Blood 63(3), 694-700.
- Lubkowski J., Palm G.J., Gilliland G.L., Derst C., Röhm K.H. and Wlodawer A. (1996)., Crystal structure and amino acid sequence of Wolinella succinogenes l-asparaginase., European Journal of Biochemistry, 241(1), 201-207.
- Clementi A. (1922)., La désamidation enzymatique de l, Archives Internationales de Physiologie, 19(4), 369-398.
- Gulati R., Saxena R.K. and Gupta R. (1997)., A rapid plate assay for screening L-asparaginase producing micro-organisms., Lett Appl Microbiol, 24(1), 23-26
- Dhevagi P. and poorani E. (2006)., Isolation and characterization of l asparaginase from marine actinomycetes., international journal of biotechnology, 5, 514-520
- Selvakumar N. (1991)., vanajakumar and Natarajan R., Partial purification, characterization and anti tumor properties of l asparaginase from vibrio in bioactive compounds from microorganisms, 289-300.
- Tiwari N. and Dua R.D. (1996)., purification and preliminary characterization of l asparaginase from Erwinia aroideae NRRL B- 138., Ind. J Biochem Biophys, 33(5), 371-376.
- Wade H.E., Robinson H.K. and Philips B.W. (1971)., Asparaginase and glutaminase activities of bacteria., J.Gen Microbiol., 69(3), 299-312.
- Wiame J.M., Grenson M. and Arst N.H. Jr. (1985)., Biodiversity of higher yeasts and filamentous fungi., AdvMicrobial Physiol., 26, 1-88.
- Pinheiro I.O., Araujo J.M., Ximenes E.C.P.A., Pinto J.C.S. and Alves T.L.M. (2001)., Production of L-Asparaginase by zymononas mobiles strain Cp4., Biomaterial and Diagnostic, 6, 243-244.
- Saxena R.K. and Sinha U. (1981)., L-asparaginase and glutaminase activities in the culture filtrates of Aspergillus nidulans., Cur Sci., 50, 218-219.
- Imada A., Igarasi S., Nakahama K. and Isono M. (1973)., Asparaginase and glutaminase activities of micro-organisms., J Gen Microbiol, 76(1), 85-99.
- Ellis M.B. (1976)., More “dematiaceous hyphomycetes”. Commonwealth Mycological Institute, Kew, Survey., England 507, ISBN 0-79235957-7.
- Raper K.B. and Fennell D.I. (1965)., The Aspergillus, Baltimore: Williams and Wilkins., 1-686.
- Lapmak Kodchakorn, Lumyong Saisamorn, Thongkuntha Sutheera, Wongputtisin Pairote and Sardsud Uraporn (2010)., L-ASPARAGINASE PRODUCTION BY BIPOLARIS SP. BR438 ISOLATED FROM BROWN RICE IN THAILAND., Chiang Mai Journal of Science., 37, 160-164.
- Hosamani R. and Kaliwal B.B. (2011)., Isolation, molecular identification and optimization of fermentation parameters for the production of L-asparaginase, an anticancer agent by Fusarium equiseti., Int J Microbiol Res, 3(2), 108-119.
- Rani S.A., Lalitha S. and Praveesh B.V. (2011)., In vitro antioxidant and anticancer activity of L-asparaginase from Aspergillus flavus (KUFS20)., Asian J. Pharm. Clin. Res., 4, 174-177.
- Sarquis M.I., Oliveira E.M., Santos A.S. and Costa G.L. (2004)., Mem Inst Oswaldo Cruz., 99(5), 489-92.
- Siddalingeshwara K.G. and Lingappa K. (2011)., Production and characterization of L-asparaginase-a tumour inhibitor., Int J Pharm Tech Res, 3(1), 314-319.
- Monica T., Lynette L., Niyonzima F.N. and Sunil S.M. (2013)., Isolation, Purification and Characterization of Fungal Extracellular L- Asparaginase from Mucor Hiemalis. J Biocatal Biotransformation 2: 2., 9, 12-14.
- Kotra S.R., Prudvi N., Sada Sai K.R.A., Mannava K.K., Peravali J.B., Kumar Anmol, Sambasiva Rao K.R.S. and Pulicherla K.K. (2013). Cost effective process for the production of fungal L-asparaginases from Penicillium sps isolated from local soil sample. Mintage Journal of Pharmaceutical and Medical Sciences, 2(1), 45-50., undefined, undefined
- Jalgaonwala R.E. and Mahajan R.T. (2014)., A review: bacterial endophytes and their bioprospecting., J Pharm Res., 4, 795-799.
- Chandrasekhar A.P. (2012)., Isolation, purification and characterization of asparaginase from aspergillus species., Int J Res Chem Environ, 2, 38-43.
- Thirunavukkarasu N., Suryanarayanan T.S., Murali T.S., Ravishankar I.P. and Gummadi S.N. (2011). Mycosphere, 2(2), 147-15., undefined, undefined
- Mohsin S.M., Dutt Sunil, Siddalingeshwara PLNSN, Karthik K.G., Jayaramu J., Naveen M., Vishwanatha M. and Prathiba T.K.S. (2012). J Acad , Ind Res,1(4), 180-182., undefined, undefined
- Yadav Niharika and Sarkar Supriya (2014). Intl J Pharmaceutical Science Invention, 3(6), 32-40., undefined, undefined
- Thirumurugan G., Moses Jeyakumar Rajesh, Leelavathy Rajesh, Vanapalli VSV and Rajarammohan Sivasubramanian (2011)., Effect of Inducers and Physical Parameterson the Production of L-Asparaginase Using Aspergillus Terreus., J Bioprocess Biotechniq, 1, 1-6. doi:10.4172/2155-9821.1000110.
- Dange V. and Peshwe S. (2015)., Purification and biochemical characterization of L-asparaginase from Aspergillus niger and evaluation of its antineoplastic activity., Int. J. Sci. Res., 4(2), 564-569.
- Kalyanasundaram J., Nagamuthu B., Srinivasan A., Pachayappan S. and Muthukumarasamy (2015)., Production, purification and characterization of extracellular L-asparaginase from salt marsh fungi endophytes., World J. Pharma. Sci., 4(3), 663-667.
- Baskar G. and Renganathan S. (2011)., Design of experiments andartificial neural network linked genetic algorithm formodelling and optimization of L-asparaginase production by Aspergillus terreus MTCC 1782., Biotechnology and Bioprocess Engineering, 16(1), 50-58.
- Murali T.S. (2011)., L-Asparaginase from marine derived fungal endophytes., Mycosphere, 2(2), 147-155.
- Kalyanasundaram I., Nagamuthu J., Srinivasan B., Pachayappan A. and Muthukumarasamy S. (2015)., Production, purification and characterization of extracellular L-asparaginase from salt marsh fungi endophytes., World J. Pharma. Sci., 4(3), 663-667.
- Gaffar S.A. and Shethna Y.I. (1977)., Purification and Some Biological Properties of Asparaginase from Azotobactervinelandii., Appl Environ Microbiol., 33(3), 508-514.
- Sreenivasulu V., Jayaveera K.N. and Mallikarjuna Rao (2009)., Optimization of process parameters for the production of Lasparaginasefrom an isolated fungus., Research J.Pharmacognosy and Phytochemistry, 1(1), 30-34.