In this study, the possible mechanisms were investigated with chitosan nanoparticles using sodium tripoly-phosphate and effects on human lymphoma SUDHL-4 in vitro. It was characterized by XRD, FTIR, TGA, particle Size, zeta potential, SEM & TEM. Different techniques such as cell proliferation, ultra structure changes, DNA fragmentation, phase distribution of cell cycle, MTT assay, MMP, agarose gel electrophoresis of DNA, flow cytometry and electron microscopy were used with treatment of different concentrations of CH-NPs (25, 50, 75, 100 μg/ml) at different time periods. Electron microscopy study revealed that the chitosan nanoparticles showed 78 nm particle size which is a high surface charge as 52 mV. Inhibition of chitosan nanoparticles after 48h treatment was marked in cell proliferation of SUDHL-4 with an IC₅₀ value of 5 μg/ml. Electron microscopy showed typical necrotic cell morphology after treatment of chitosan nanoparticles. The DNA degradation related with necrosis was determined using agarose electrophoresis and loss of MMP & occurrence of apoptosis was analyzed by flow cytometry. Chitosan nanoparticles with low molecular weight (LMW) were comparatively stable in medium containing aqueous and rate of dissolution was slow in acidic medium. Results of this present study clearly provided information that the chitosan nanoparticles effectively inhibit the proliferation of SUDHL-4 through multiple mechanisms in vitro and this novel formulation can open a new avenue against human Lymphoma.