Mitomycin C (MMC) is antitumor-antibiotic, currently in use for treatment of various solid tumors. The effect of different dosages of MMC was tested in human lymphocyte culture after exposure for different duration. The exposure of 0.05μg/ml MMC for 24 hours was not sufficient to induce mitotic inhibition and chromosomal aberrations. The cells exposed to concentration 2.0μg/ml and above for last 24 hours showed complete inhibition of mitotic activity. Low concentration exposure for long duration inhibits mitosis rather than exposure to high concentration for short duration. Mitotic indexes in after exposure of MMC for same duration were decreased significantly with the increase in the dose of MMC. The highest frequency of chromosomal aberrations was observed in cultures treated with 0.5μg/ml MMC for the last 24 hours of culture. MMC did not induce chromosomal aberrations randomly; which more commonly affected chromosome 1, 9 and 16. Most commonly affected was chromosome 9 followed by the involvement of chromosome 1 and least was the involvement of chromosome 16. The non random interchange breakages induced in these chromosomes were significantly at secondary constriction region i.e., site of paracentric heterochromatin, most commonly that of chromosome 9.