Industrial Production and Clinical Application of L-Asparaginase: A Chemotherapeutic Agent
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Industrial Production and Clinical Application of L-Asparaginase: A Chemotherapeutic Agent

Authors: Soni Yadav, Sitansu Kumar Verma, Jitendra Singh, Ajay Kumar

Abstract:

This article comprises detail information about L-asparaginase, encompassing topic such as various sources of L-asparaginase, mechanism and properties of L-asparaginase. Also describe the production, cultivation and purification of L-asparaginase along with information about the application of L-asparaginase. L-asparaginase catalyzes the conversion reaction to convert asparagine to aspartic acid and ammonia. Asparagine is a nutritional requirement for both normal and tumor cell. Present scenario has found that L-asparaginase has been found to be a best anti tumor or antileukemic agent. In the recent years this enzyme gained application in the field of clinical research pharmacologic and food industry. It has been characterized based on the enzyme assay principle hydrolyzing L-asparagine into L-aspartic acid and ammonia. It has been observed that eukaryotic microorganisms such as yeast and filamentous fungi have a potential for L-asparaginase production. L-asparaginase has been and is still one of the most lengthily studied therapeutic enzymes by scientist and researchers worldwide.

Keywords: L-asparaginase, antitumor, solid state fermentation, chemotherapeutic.

Digital Object Identifier (DOI): doi.org/10.5281/zenodo.1091554

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[1] A. Abuchowski, G. Kazo, and C. Verhoest, Cancer therapy with chemically modified enzymes. I. Antitumor properties of polyethylene glycol L-asparaginase conjugates. Cancer Biochem Biophys, 7, 175-180. 1984.
[2] R. Alegre, L. A. Monte and L. A. Minim, Cheese whey utilization for L- asparaginase production from Erwnia arodeae NRRL B-138 in pilot plant. Arquivos de Biologia e Technology (Curitiba), 36(3): 525-534. 1993.
[3] E.A. M. Ali. Purification and Characterization of Vignaunguiculata Cultiver Asparaginase. The Egyptian Journal of Biochemistry & Molecular Biology. 2(1):145-162. 2009.
[4] S. S. Ali, V.Rai, K. Soni, P. Kulshrestha, S. K. Lai. A fungal L-asparaginase with potential antitumor activity. Ind. J. Microbiol.. 34, 73–76. 1994.
[5] I. M. Appel, C. van Kessel-Bakvis, R. Stigter, R. Pieters. "Influence of two different regimens of concomitant treatment with asparaginase and dexamethasone on hemostasis in childhood acute lymphoblastic leukemia". Leukemia. 21 (11): 2377–80. PMID: 17554375. 2007.
[6] K. Arima. Microbial enzyme production in Global Impact of Applied Microbiology, M.P. Starr (eds.), pp. ,279-299, John Willey, New York, USA. 1964.
[7] A. Ashraf, S. El-Bessoumy, Mohamed, and M. Jehan. Production, Isolation, and Purification of L-Asparaginase from Pseudomonas Aeruginosa 50071 Using Solid-state Fermentation, Journal of Biochemistry and Molecular Biology, Vol. 37, No. 4, pp. 387-393. PMID: 15469724. 2004.
[8] W. Azmi, Studies on production of L- asparaginase an antiumour enzyme by a new bacterial isolate. 42 Conf Assoc Microbiol India, Gulbarga University, Gulbarga, 9-11. 2001.
[9] S. M . Badr El-Din, M. S. Foda., Kineties and properties of L-asparaginase and L-glutaminase activities of Pseudomonas ovalis. Zentralbl Bakteriol parasetenkd Infektionskr Hyg, ,131, 489-496. PMID: 13588. 1976.
[10] S. Bascomb, G. T. Banks, M. T. Skarstedt, A. Fleming, K. A. Bettelheim. The properties and large scale production of L-asparaginase from citrobacter.J Gen Microbiol, 91, 1-16. PMID: 465. 1975.
[11] N. Basha Saleem, R. Rekha, M . Komala and S. Ruby. Production of Extracellular Antileukaemic Enzyme L- asparaginase from Marine Actinomycetes by Solidstate and Submerged Fermentation: Purification and Characterisation. Tropical Journal of Pharmaceutical Research. 8 (4): 353-360. 2009.
[12] T. L . Bell, and M. A. Adams. Ecophysiology of ectomycorrhizal fungi associated with Pinus spp. Plant Ecology.171(1-2): 228-231. 2004.
[13] M. C. Berenbaum, H. Ginsburg, D. M.Gilbert. Effects of L-asparaginase on lymphocyte target cell reactions In Vitro. Nature. 227, 1147–1148. PMID: 4915992. , 1970.
[14] P . Blánquez, M. Sarrà, M. T. Vicent. "Study of the cellular retention time and the partial biomass renovation in a fungal decolourisation continuous process,”Water Research,40, no. 8, pp. 1650–1656, PMID: 16616292. ., 2006.
[15] E. P. Bon, E. Carvajal, M. Stanbrough, D. Rowen, B. Magasanik. Asparaginase II of Sacharomyces cerevisiae. GLN3/URE2 regulation of a periplasmic enzyme. Appl Biochem Biotechnol,. 63/65, 203-212. PMID: 9170245. 1997.
[16] J. Boos, G. Werber, E. Ahlke, P. Schulze-Westhoff, U. Nowak-Göttl, G . Würthwein, E. J. Verspohl, J. Ritter, H. Jürgens. Monitoring of asparaginase activity and asparagine levels in children on different asparaginase preparations. Eur J Cancer, 32, 1544-1550. PMID: 8911116. 1996.
[17] D. Borek, K. Michalska, K. Brezezinski, A. Kisiel, J. Podkowinski, D. T. Bonthron, D. Krowarshch, J. Otlewski. and M. Jaskolski. Expression, purification and catalytic activity of Lupinus luteus aspergines beta. amidohydrolaseand its Escherichia coli homolog. Eur J Biochem, 271(15):3215-3226. PMID: 15265041. 2004.
[18] D. Borek, J. Podkowinski, A. Kisiel and M.Jasloski. Isolation and characterization of cDNA encoding L- asparaginase from Lupinus luteus (Accession No. AF112444), Plant Physiol. 119: 1568-1570. 1999.
[19] B. Borkotaky, and RL.Bezbaruah. Production and properties of asparaginase from a new Erwinia sp. Folia Microbiologica,47(5):473-476. PMID: 12503389. 2002.
[20] G. E. P. Box, W. G. Hunder, S. J. Hunder. Statistics for Experiments. John Wiley & Sons Inc New York. 1978.
[21] K. S.Chang, and K. J. F. Farnden. Purification and properties of Asparaginase EC-3.5.1.1 from Lupinus arboreus and Lipinus angustifolius. Archives of biochemistry and biophysics, 208(1): 49-58. PMID: 7259189. 1981,
[22] M. P. Curran, R. M. Daniel, G. R. Guy, H. W. Morgan. A specific L-asparaginase from Thermus aquaticus. Arch Biochem Biophys,. 241,571-576. PMID: 3929688. 1985.
[23] R. Datar. Economic of primary separation steps in relation to fermentation and genetics engineering. Process Biochem, 21, 19-26. 1986.
[24] L. Davidson, M. Burkorn, S. Ahn, L. C. Chang and B. Kitto. L- asparaginase from Citrobactor freundii. Biochim. Biophs Acta. 480:282-94. PMID: 401650. 1977.
[25] Dejong. L- asparaginase production by Streptomyces griseus. Appl. Microbiol. 23; 1163-64. PMID: 4626231. 1972.
[26] K . Dhevendaram and K . Annie. Antibiotic and L- asparaginase of Streptomycetes isolated from fish, Shellfish and sediments of veli estuarine along Kerala coast, Indian J Mar Science. 28: 335-37. 1999.
[27] D. C. Dietrich, M. Landwehr, C. Reissner, R. H. Smalla, K. Richter, G. Wolf, T. M. Bockers, E. D. Grundelfinger, and M. R. Kreutz. Gliap-a novel untypical L- asparaginase localized to rat brain astrocytes. J Neurorochem, 85(5): 1117-1125. PMID: 12753071. 2003.
[28] J. A. Distasio, R. A. Niederman, D. Kafkewitz, D. Goodman. purification and characterization of L-asparaginase with antilymphoma activity from Vibrio succinogenes. J Biol Chem. 251, 6929-6933. PMID: 11211. 1976.
[29] D. Drainas & C. Drainas, A conductimetric method for assaying asparaginase activity in Aspergillus nidu L-asparaginase. Eur J Biochem,. 151, 591-593. PMID: 3896790. 1985.
[30] J. Dunn. Research in Brief. Published June 1,. www.foodmanufacture.co.uk/news/fullstory.php/aid/B1/Rearch.in.brief.html. 2004.
[31] P. C. Dunlop, G. M. Meyer, D. Ban, and R. J. Roon. Characterization of two forms of asparaginase in Saccharomyces cerevisiae. Journal of Biological Chemistry, 253(4): 1297-1304. PMID: 342521. 1978.
[32] A. A. El-Bessoumy, S. Mohamed and M. Jehan. Production, Isolation, and Purification of L- asparaginase from Pseudomonas Aeruginosa 50071 Using Solid-state Fermentation. Journal of Biochemistry and Molecular Biology. 37(4): 387-393. 2004.
[33] M. S. Foda, H. H. Zedan, and S. A. E. M. Hashem. Characterization of novel L- asparaginase produced by Rhodotorula rubra. Revista Latinoamericana de Microbiologia, 22(2): 87-96. PMID: 7027389. 1980.
[34] B. H. Frank, A. H. Pekar A. J. Veros. Ho PPK, Crystalline L-asparaginase from Escherichia coli B. II. Physical properties, subunits, and reconstitution behavior. J. Biol. Chem, 245, 3716. PMID: 4919215. 1977.
[35] M. P. Gallogher, R. D. Murshall, and R.Wilson. Asparaginase drug for treatment of acute lymphoblastic leukemia. Essays Biochem., 24, 1-40. 1989.
[36] M. M.Garaev, and E. I. Golub. Mechanism of the effect of glucose on L- asparaginse synthesis by Escherichia coli bacteria, Mikrobiologia, 46(3):433-439. 1977.
[37] L. Giovanni, V. G. F. Pagliardi, Gavosto. Mechanism of Action of L-Asparaginase on the Cell Cycle and Growth in Acute Lymphoblastic Leukemia. Acta Haematol, 50:257-268. PMID: 4202629. 1973.
[38] C. R. Goward, G. B. Stevens, I. J . Collins, I. R.Wilkinson, and M. D. Scawen. Use of macrosorb keiselguhr composite and CM-sepharose fast flow for the large-scale purification of L- asparaginase from Erwinia chrysanthemi. Enzyme and Microbial Technology. 11(12): 810-814, 1989.
[39] S. Gunasekaran, L. McDonald, M. Manavathu, E. Manavathu, and M. Gunasekaran. Effect of culture media on growth and L- asparaginase production in Nocardia asteroids. Biomedical letters, 52(207): 197-201. 1995.
[40] B. Gurunathan,and S. Renganathan. Optimization of Culture Conditions and Bench-Scale Production of L- asparaginase by Submerged Fermentation of Aspergillus terreus MTCC 1782. J. Microbiol. Biotechnol, 22(7), 923–929. PMID: 22580311. 2012.
[41] V.Heesgen, J.Matlok, S.Schrader, and H.Rudolph. Asparagine catabolism in bryophytes: Purification and characterization of two L- asparaginase isoforms from Sphagnum fallex. Physiologia Plantarum. 97(2): 402-410. 1996.
[42] H. V.Hendriksen, B. A. Kornbrust, P. R. Oestergaard, M. A. Stringer, , "Evaluating the Potential for Enzymatic Acrylamide Mitigation in a Range of Food Products Using an Asparaginase from Aspergillus oryzae". Journal of Agricultural and Food Chemistry. 57 (10): 4168–4176. doi: 10.1021/jf900174q. Epub 2009 Apr 23. PMID: 193886392009.
[43] K . Hellman, D. S. Miller, and K. A. Cammack. The effect of freeze drying on the quarternary structure of L- asparaginase EC-3.5.1.1 from Erwinia carotovora, Biochemica-et-Biophysia-Acta. 749(2):133-142. PMID: 6652094. 1983.
[44] J. Hess, C. Leitner, C. Galhaup, Enhanced formation of extracellular laccase activity by the white-rot fungus Trametes multicolor, Applied Biochemistry and Biotechnology—Part A Enzyme Engineering and Biotechnology, 98-100, pp. 229– 241. PMID: 12018250, 2002.
[45] J. Hill, J. Roberts, Loeb, E. A. Kahn and R. Hill. L- asparaginase therapy for leukemia and other malignant neoplasm. JAMA . 1967: 882. 2002.
[46] C. Howard and J. H. Schwartz, Production of L-asparaginase II by Escherichia coli. J. Bacteriol. 96:2043-2048. PMID: 4881701. 1968.
[47] N. G. Illarionova, L. N. Petrov, L. V. Olennikova, S. N. Roshchin,. A. Pasechnik, B. D . Khalyapin, A. E. Polotskii, N. E. Voinova, and TB T. B. Shtukina, Study of L- asparaginase EC-3.5.1.1 secondry structure in a wide pH region. Molekulyamaya Biologiya (Moscow), 14(4): 951-955. 1980.
[48] P. E. Joner. Purification and properties of L-asparaginase B from Acinatobacter calcoaceticus. Biochem Biophys Acta. , 438,287-295. PMID: 938683. 1976.
[49] G. E. Jones. Genetic and physiological relationships between L- asparaginase I and L- asparaginase II Saccharomyces cerevisiae. Journal of biotechnology, 130(1): 128-130. PMID: 323221. 1977.
[50] K. D. Kamble, P. R. Bidwe1, V. Y. Muley, L. H . Kamble, D. G. Bhadange and M . Musaddiq, Characterization of L-asparaginase Producing Bacteria from Water, Farm and Saline Soil. Bioscience discovery, 3(1):116-119. . 2012.
[51] M. H. Kang, Y. H. Kang,. B. U. Szymanska,. W . Kalak,. M. A. Sheard &.T. M. Harned. Activity of vincristine, L-ASP, and dexamethasone against acute lymphoblastic leukemia is enhanced by the BH3-mimetic ABT-737 in vitro and in vivo. Blood Journal., 110(6), 2057. PMID: 17536015. 2007.
[52] J. O. Kill, G. N. Kim, I. Park. Extraction of extracellular L-asparaginase from Candida utilis. Biosci Biochem, 1995. 59, 749-750. PMID: 7772845.
[53] A. N. Kondart Eva. Comparative evaluation of the clinical efficacy of 3 preparations of L- asparaginase from Escherichia coli. Antibiotiki-(Moscow). 29(7): 527-531. 1984.
[54] S. Kumar , V. V. Dasu and K . Pakshirajan. Localization and production of novel L-asparaginase from Pectobacterium carotovorum MTCC 1428. Process Biochem, 45: 223-229. . 2010.
[55] J. M . Lema, E . Roca, A . Sanroman, M. J. Nunez, M. T. Moreira and G . Feijoo. "Pulsating bioreactors,” in Multiphase Bioreactor Design, J. M. S. Cabral,M.Mota, and J. Tramper, Eds., 309–329, 2001.
[56] C. Liu, J. D.Kawedia, C. Cheng, D. Pei, C. A. Fernandez & X .Cai. Leukemia.,
[Epub ahead of print]. 2012.
[57] S.Manna, A. Sinha, R. Sadhukhan, S. L. Chakrabarty. Purification, characterization and antitumour activity of L- asparaginase isolated from Pseudomonas stuzeri MB-405. Curr Mocrobiol,. 30, 291-198. PMID: 7766157. 1995.
[58] N. K. Maladkar, V. K. Singh, S. R. Naik. Fermentative production and isolation of L- asparaginase from Erwinia carotovora, EC-3 Hindustan Antibiotic Bull, 35, 77-86. PMID: 8181956. 1993.
[59] S. R. Mardashev, A. Y. Nikolaev, N. N. Sokolov, E. A. Kozlov, and M. E. Kutsman . Isolation and properties of an homogeneous L asparagenase prepration from Pseudomonas flourescens AG. Biokhimia. 40(5); 984-989. PMID: 2329. 1975.
[60] J. M. Mesas, J. A.Gil, J. F. Martín. Characterization and Partial purification of L- asparaginase from Cornebacterium glutamicum. J Gen Microbiol, 136, 515-519. PMID: 2391490. 1990.
[61] J. Mikucki, J. Szarapińska-Kwaszewska, Z. Krzemiński. Factors influencing L-asparaginase production by Staphylococci. Zentralbl Bakteriol Parasetenkd Infektionskr Hyg,. 132, 135-142. PMID: 17983. 1997.
[62] B.R .Mohapatra, M .Bapuji, U. C.Banerjee. Production and properties of L-asparaginase from Mucor sp associated with a marine sponge (Spirastrella sp.). Cytobios. 92, 165-173. 1997.
[63] B. R. Mohapatra, R. K.Sani, and U. C.Banerjee. Characterization of L-asparaginase from Bacillus sp isolated from an intertidal marine alga(Sargassum sp). Lett Appl Microbiol.,21, 380-383. 1995.
[64] Z. B. Moola, M. D. Scawen, T. Atkinson, D. J. Nicholls. Erwinia chrysanthemi L-asparaginase, epitope mapping and production of antigenically modified enzyme. Biochem J. 302, 921-927. 1994.
[65] S. A. Mostafa, and M. S. Salama. L- asparaginase producing Streptomyces from soil of Kuwait. Zentralbl Bakteriol (Naturwiss),134. 325-334. 1979.
[66] S. A. Mostafa. Properties of L- asparaginase in cell free extract of Streptomyces karnatakensis. Zentralbl Mikrobiol, 137, 63-71. 1982.
[67] J. Mukherjee, S. Majumdar, T. Scheper. Studies on nutritional and oxygen requirements for production of L-asparaginase by Enterobacter aerogenes. Appl Microbiol Biotechnol. 53, 180-184. 2000.
[68] M. S. Nawaz, Isolation and characterization of Enterobacter cloacae capable of metabolizing L-asparaginase. Appl Microbiol Biochem. 50, 568-572. 1998.
[69] M. V. Nefelova, S. G. Ignatov, A. G. Chigalenchik, B. D. Vinogradov, N. S. Egorov. Biosynthesis of L-asparaginase II by cultures of Bacillus polymyxa var Ross. Prikl Biokhim Mikrobiol. 14,510-514. 1978.
[70] J. Netrval. Stimulattion of L-asparaginase in E.coli by organic and amino acid. Folia Microbiol (Praha), 22, 106-116. 1977.
[71] I. Pastuszak & M. Szymona. Purification and properties of L-asparaginase from Mycobacterium phlei. Acta biochem Pol, . 23, 37-44. 1976.
[72] K. K. R Patro, S. Satpathy and N. Gupta. Evaluation of Some Fungi For L-Asparaginase Production. Indian Journal of Fundamental and Applied Life Sciences. 1(4): 219-221. 2011.
[73] J. H. Paul and K. E. Cooksey. Regulation of L- asparaginase EC-3.5.1.1 in a chlamydomonas species in response to ambient concentration of combined nitrogen. Journal of Bacteriology,147(1):9-12. 1981.
[74] R. E. Peterson and A. Cieglar. L- asparaginase production by Erwinia aroidae. Appl. Microbiol. 18: 64-67. 1996.
[75] R. Pieters, S. P. Hunger, J. Boos, C. Rizzari, L. Silverman & A. Baruchel, 2011 , L-asparaginase treatment in acute lymphoblastic leukemia: a focus on Erwinia asparaginase. Cancer., 117(2), 238. doi: 10.1002/cncr.25489. Epub. 2010.
[76] S. M. Pradeep, R. Mahmood and K. S. Jagadeesh. Screening and characterization of L- asparaginase producing microorganisms from tulsi (Ocimum sanctum. L). Karnataka J. Agric. Sci. 23 (4): 660-661. 2010.
[77] A. A. Pritsa and D. A. Kyriakidis. L-asparaginase of Thermus thermophillus: Purification, Properties and identification of essential amino acid for its catalytic activity. Molecular and cellular Biochemistry, 216(1-2): 93-101. 2001.
[78] S. K. Raha, S. K. Roy, S. K. Dey, S. L. Chakrabarty. Purification and properties of L-asparaginase from Cylindrocarpon obtusisporum MB-10.J Biochem Int. 21, 987-1000. 1990.
[79] N. Ramaiah, D. and Chandramohan. Production of L- asparaginase from marine Luminous bacteria. Indian Journal of Marine Science, 21(3): 212-214. 1992.
[80] M. S. Ramakrishnan, and R. Joseph. Characterization of an extracellular asparaginase of Rhodosporidium toruloides CBS14 exhibiting unique physicochemical properties. Canadian Journal of microbiology, 42(4): 316-325. 1996.
[81] V. K. Redd, and S. M. Reddy. Effect of carbon and nitrogen source on L- asparaginase production by Bacteria. Indian J Microbiol, 30,81-83. 1990.
[82] J. Roberts, G. Bursen, and M. H. Joseph, New procedures for purification of L-asparaginase with high yield from Escherichia coli. J Bacterial, 95, 2117-2123. PMID: 4970225. 1968.
[83] J. Roberts, J. S. Holcenberg, W. C. Dolowy, Isolation, characterization and properties of Achromobacteriaceae glutaminase-asparaginase with anti tumor activity. J Biol Chem. 247, 84-90. 1972.
[84] M. Rozalska & J. Mikucki. Staphylococcal L-asparaginase, catabolic repression of synthesis. Acta Microbiol Pol, 1992. 145-150. 1992.
[85] B. Rowly & J. C. Wriston, L-asparaginase from Serratia marcescens. Biochem Res Commun. 28, 160-171. 1967.
[86] T. Sakamoto, C. Araki, T. Beppu, and K. Arima, Partial purification and some properties of extracellular Asparaginase from Candida utilis. Agricultural and Biological Chemistry, 41 (8): 1359-1364. 1977
[87] M. I. Sarquis, E. M. Oiiiviera, A. S. Santos, and G. L. da-Costa, Production of L-asparaginase by filamentous fungi. Memorias-do-Instuto-Oswaldo-Cruz, 99(5): 489-492. 2004.
[88] N. A. Savitri, and W. Azmi, Microbial L- asparaginase: A potent antitumor enzyme. Indian J. Biotechnol, 2: 184-194. 2003.
[89] M. L. Shwu, T. John, H. Marie. Process of manufacture of L- asparaginase from Erwinia chrysanthemi. US Pat. 4729957. 1998.
[90] A. Sonawane, U. Kloppner, S. Hovel, U. Volker, and K. H. Rohm. Identification of Pseudomonas proteins coordinately induced by acidic amino acids and their amides: a two-dimentional electrophoresis study, Microbiology,149(Pt 10): 2909-2918. 2003.
[91] R. M. Stark, M. S. Suleiman, I. J. Hassan, J. Greenman, M. R. Millar, Amino acid utilization and deamination of glutamine and asparagines by Helicobacter pylori, J Med Microbiol,46, 793-800. 1997.
[92] K. R. Stepanyan, & M. A. Davtyan. Some question of thermostability of L-asparaginase of yeast Candida guilliermondii. Biologicheskii Zhurnal Armenii. 41, 599-603. 1998.
[93] M. Sugumaran , L. Giglio, H. Kundzicz, S. Saul, and V. Semensi, Studies on the enzymes involved in puparial cuticle sclerotization in Drosophila melanogaster,” Archives of insect biochemistry and physiology, 19, no. 4, pp. 271–283, 1992.
[94] E. S. Tiul'panova, V. V. Eremenko, S. R. Mardashev, Activity and properties of l-asparaginase from Bacillus mesentericus. 43A. Microbiologika. 41 423-429. 1972.
[95] N. Tiwari & R. D. Dua, Purification and preliminary characterization of L-asparaginase from Erwinia aroideae.Indian J Biochem Biophys.33, 371-376. 1996.
[96] T. Tosa, R. Sano, K. Yamamoto, M. Nakamura and I. Chibata, L- asparaginase from Proteus vulgaris. Appl. Microbiol. 22: 387-92. 1971.
[97] D. J. Triantafillou , J. G. Georgatsos, Purification and properties of a membrane-bound L-asparaginase of Tetrahymena pyriformis. Molecular and cellular Biochemistry, 81(1): 43-51. 1998.
[98] S. A. Tsirka, & D. A. Kiriakidis, L-asparaginase of Tetrahymena pyriformis is associated with a kinase activity. Mol Cell Biochem. 95, 77-78. 1990.
[99] D. Z. Wei, and H. Liu, Promotion of L- asparaginase using n-dodecane. Biotechnology Techniques, 12(2): 129-131. 1998.
[100] M. Wetzler, B. L. Sanford, J. Kurtzberg, D. Oliveira, S. R. Frankel, & B. L. Powell, Effective asparagine depletion with pegylated asparaginase results in improved outcomes in adult acute lymphoblastic leukemia: Cancer and Leukemia Group B Study 9511. Blood. 109(10), 4164. 2007.
[101] S. T. Williams, and JC Vikers. The ecology of antibiotics production. Microbial Ecology, 12, 43-52. 1986.
[102] F. Zaho, and J. Yu, L-asparaginase release from Escherichia coli cells with K2HPO4 and Triton X-100. Biotechnol progr, 17, 3, 490-494. 2001.
[103] S. K. Verma, & A. Kumar. Therapeutic uses of Withania somnifera (Ashwagandha) with a note on withanolides and its pharmacological actions. Asian Journal of Pharmaceutical and Clinical Research, 4(1), 01-04. 2011.