Nanoparticular systems, which have made great advances with the development of nanotechnology, have been used extensively recently in pesticide, herbicide plant growth regulators, fertilizer applications, gene transfer technologies and agriculture. It was aimed to synthesize and characterize 2,4-D loaded PLGA nanoparticles and investigate their biological activity in comparison with its free form. Here, the effects of 2,4-D loaded poly (lactic-co-glycolic) acid (PLGA) nanoparticles on biomass in Medicago sativa cell suspension cultures were investigated. Single emulsion solvent evaporation method is used in nanoparticle synthesis. As a result of the characterization of nanoparticles, 63.82% encapsulation efficiency, 60.73% reaction efficiency and 10.51% drug loading capacities were calculated. Particle size was measured as 181.7 ± 3.74 nm, zeta potential-18.3 ± 1.48 and polydispersity index as 0.081. Compared with the free 2,4-D molecule, it was observed that the addition of 2,4-D to the medium using the nanoparticles drug release system increased the growth of plant cells and the yield of biomass in M. sativa cell suspension cultures.