This study aims to evaluate the effects of N-acetylcysteine (NAC) on bovine oocyte maturation, mitochondrial activity and transzonal projections (TZP), as well as on the levels of reactive oxygen species (ROS) and messenger RNA (mRNA) for catalase (CAT) superoxide dismutase (SOD), periredoxin-6 (Prdx6), glutathione peroxidase (GPx), growth and differentiation factor-9 (GDF9), histone H1Foo, cyclin B1 (CCNB1) and c-Mos. Bovine cumulus–oocyte complexes (COC) of medium-sized antral follicles (3.0–6.0 mm) were prematured in TCM-199 for 8 h at 38.5°C in 5% CO2. After prematuration in the presence of forskolin and C-type natriuretic peptide, COCs were matured in TCM-199 alone or with 0.1, 0.5 or 2.5 mM NAC. Then, oocytes were classified according to the stage of chromatin. Furthermore, mitochondrial activity and intracellular levels of ROS and TZP were also evaluated. The levels of mRNAs for CAT, SOD, Prdx6, GPx, GDF9, H1Foo, CCNB1 and c-Mos were evaluated using real-time polymerase chain reaction (RT-PCR). The results showed that NAC significantly increased the percentages of oocytes with resumption of meiosis when compared with those oocytes matured in control medium. Oocytes had homogeneous mitochondrial distribution, and those cultured with 0.1 and 0.5 mM NAC had lower levels of ROS when compared with the control. In addition, 0.5 mM NAC reduced TZP and the levels of mRNA for CCNB1. In contrast, NAC did not influence the expression of CAT, GPx, Prdx6, SOD, GDF9, H1Foo, and c-Mos. In conclusion, 0.5 mM NAC reduced the levels of ROS, TZP and mRNA for CCNB1, and improved in vitro resumption of meiosis in oocytes from medium-sized bovine antral follicles.