Synthesis of poly(ɛ-caprolactone) nanospheres in the presence of the protective agent poly(glutamic acid) and their cytotoxicity, genotoxicity and ability to induce oxidative stress in HepG2 cells

Abstract

Nanospheres of poly (ɛ-caprolactone) (PCL) with sizes smaller than 200 nm were produced by combining the freeze drying method and the physicochemical solvent/non-solvent approach. The influence of various types of cryoprotectants (poly(glutamic acid) (PGA) or sacharose) and their concentrations on the outcome of freeze-dried poly(ɛ-caprolactone) particles was evaluated. The physiochemical properties, structural and morphological characteristics of thereby obtained PCL particles were determined by scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). The cytotoxicity of the samples was examined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT assay). The formation of intracellular reactive oxygen species was measured spectrophotometrically using a fluorescent probe (DCFH-DA assay). In addition, the genotoxic response of PCL particles obtained using PGA as a cryoprotectant was investigated by the Comet assay. This paper focuses on the role of PGA in the synthesis of PCL particles and demonstrates that PGA plays a dual role in the synthesis, i.e. it acts as a stabilizer but also as a cryoprotective agent. The sufficient and optimal concentration of PGA for producing uniform, spherical but also biocompatible PCL nanoparticles is established to be 0.05%.