Retarded development and blastomere fragmentation of human preimplantation embryos represent a common phenomenon in in-vitro culture systems. Even though media composition is generally formulated to meet embryo nutritional requirements, the influence of antibiotic supplementation has not been investigated thoroughly. The present study was performed to evaluate the effects of antibiotics on embryo morphology and growth in modified culture media. A total of 196 zygotes from 18 couples was cultured in three different media: (i) conventional medium (n = 99, control group); (ii) medium modified with half the standard antibiotic concentration (n = 54; and (iii) antibiotic-free medium (n = 43); 49 embryos from the control group were selected at the zygote stage and transferred to the patients on day 2. The remaining 147 zygotes were cultured to the blastocyst stage for cryopreservation; their morphology and cell number were assessed daily at 40, 64, 88 and 112 h post-insemination. Overall cleavage rate was 95% and embryo scoring revealed 91% grade 1 embryos throughout the culture period in the three media. Significantly higher cleavage rates were obtained in the antibiotic-free medium at each observation, including the blastocyst stage, when compared to the other two groups. In addition, no notable improvement was observed in the embryos cultured in a reduced concentration of antibiotics. In conclusion, antibiotic supplementation of media has an adverse effect on the growth rate of preimplantation embryos, even in reduced concentrations, suggesting that antimicrobial drugs may interfere with the timing of cleavage events either by delaying or blocking embryo development.