Purification, Characterization and In vitro Cytotoxicity of L-asparaginase from Withania somnifera L. Against Acute Lymphoblastic Leukemia

Withania somnifera L. has been traditionally used as a sedative and hypnotic. Withania somnifera L. is reported to have anti-carcinogenic effects in animal and cell cultures by decreasing the expression of nuclear factor-kappaB, suppressing intercellular tumor necrosis factor, and potentiating apoptotic signalling in cancerous cell lines [1]. The present study was carried out on the purification, characterization and in vitro cytotoxicity of L-asparaginase from Withania somnifera L., a popular medicinal plant. L-asparaginase was purified from the crude extract of the fruits of Withania somnifera L. up to 95% through column chromatography. The purified L-asparaginase was characterized by size exclusion chromatography, PAGE and 2-D PAGE. The antitumor and growth inhibition effect of the L-asparaginase was assessed using MTT colorimetric dye reduction method. The purified enzyme is a homodimer, with a molecular mass of 72 ± 0.5 kDa, and pI value of the enzyme was around 5.1. It is the first report for plant L-asparaginase with antitumor activity. Data obtained from the MTT assay indicated that L-asparaginase significantly (P < 0.05) reduced the viability of lymphocyte cells in a dose-dependent manner, showing a LD50 value of 1.45 ± 0.05 IU/ml. Withania somnifera L. proved to be an effective and a novel source of L-asparaginase, furthermore it shows lot of similarity with bacterial L-asparaginases which have already been commercialized for the treatment of acute lymphoblastic leukemia. References: [1] Ichikawa H, et al. (2006), Molecular Cancer Therapeutics, 5(6): 1434–1445.