This study aimed to evaluate the effects of supplementation with different concentrations of L-carnitine (L-car) during in vitro maturation (IVM) of bovine oocytes on their mitochondrial membrane potential (MMP), reactive oxygen species (ROS) levels, and subsequent embryonic development. Cumulus-oocyte complexes (COC) were matured for 22 h at 38.5°C, 5% CO₂ in air, on IVM medium (TCM-199 with bicarbonate, hormones and 10% FCS) supplemented with 0 (Control), 1, 5 or 10 mM of L-car. Matured and immatured oocytes (0 h) were stained with 500 nM of MitoTracker Red (Molecular Probes, Invitrogen), or with 5 mM of H₂DCFDA (Molecular Probes, Invitrogen). Stained oocytes (MMP: n = 191; and ROS: n = 250) were evaluated under an epifluorescence inverted microscope (excitation 579/495nm and emission 599/404nm, respectively) and the images analyzed by Q-Capture Pro image software to measure the arbitrary fluorescence units. The fluorescence intensity values were subtracted from mean values of “background” in the images. The 0h group was chosen as the calibrator, and each treatment value was divided by the mean of the 0h. In a second trial, COC (n = 1875) were IVM as above and then fertilized. The presumptive zygotes were cultured in SOF medium at 38.5°C, 5% CO₂ in air, for 7 d (Day 0 = IVF), when the blastocysts (Bl) rates were evaluated. The averages were compared by ANOVA followed by the Tukey’s test and data are presented as mean ± SEM. It was found a reduction (P < 0.05) on MMP after IVM with L-car 10mM (0.4 ± 0.0^c) in comparison to 1 mM (1.1 ± 0.1^ab) and 5 mM (1.0 ± 0.0^ab), as well as to 0h (1.0 ± 0.1^a); however, no treatment differed from the Control (0.8 ± 0.0^abc). In respect to ROS levels, we found an increase (P < 0.05) in oocytes matured with 10 mM (2.3 ± 0.1^c), in comparison to Control (1.8 ± 0.1^b), 1 mM (1.9 ± 0.1^b), 5 mM (2.0 ± 0.1^b) and 0 h (1.0 ± 0.0^a). Bl rates were similar (P > 0.05) in all treatments (25.2% ± 3.7 to 37.1% ± 2.7). In conclusion, the reduction on MMP after IVM of bovine oocytes with L-car 10 mM was followed by an increase of ROS level. However, there was no influence on their acquisition of capacity to Bl development. Financial support: FAPESP (#2012/10084–4 and #2013/07382–6)