This study aimed to determine the effect of L-carnitine on the growth and subsequent nuclear maturation of buffalo small growing oocytes (92–108 µm in diameter) in vitro. Oocyte-granulosa cell complexes (OGCs) were dissected from early antral follicles of slaughtered buffaloes and cultured in in vitro growth (IVG) medium with the supplementation of different concentrations (0, 1.25, 1.875 or 2.5 mM) of L-carnitine for 6 days. The results revealed that L-carnitine increased the diameter of buffalo oocytes in vitro. The degeneration rate was significantly (p < 0.05) lower in 2.5 mM of L-carnitine-treated oocytes (10%) than others (55%, 45% and 32.5% in 0, 1.25 and 1.875 mM of L-carnitine-supplemented groups, respectively). The OGCs showed antrum-like structures significantly (p < 0.05) higher in the 2.5 mM of L-carnitine group (74.0%) than the 0- and 1.25-mM groups (34.6% and 38.1%, respectively). Furthermore, in vitro grown oocytes were placed in in vitro maturation (IVM) medium for 24 h to examine meiotic competence of in vitro grown oocytes with L-carnitine. The L-carnitine (1.875 and 2.5 mM) treated oocytes showed a higher rate of nuclear maturation up to the metaphase II (MII) stage and a lower rate of degeneration. In conclusion, L-carnitine enhances the growth, prevents degeneration, promotes the formation of antrum-like structures and supports nuclear maturation of buffalo oocytes in vitro.