Abstract: β-carotene is the most widely studied carotenoid that has provitamin A properties beside its great promise for cancer chemoprotective and human physiological functions. β-carotene can be cleavaged to retinal by some enzymes in wide variety of cells especially in intestinal cells. The retinal is further metabolized to retinoic acid by Aldehyde Dehydrogenase (ALDH) family of enzymes. In this study we use CMT-93 cell line as a cell model of intestinal epithelial cell to study the effect of β-carotene on cells morphology, proliferation and Aldehyde Dehydrogenase (ALDH) gene expression. These cells were grown in DMEM supplemented with 10% heat-inactivated Fecal Bovine Serum (FBS), 4 mM L-glutamine, 50 U/ml penicillin and 50 mg/ml streptomycin. Tetrahydrofuran (THF) (1.25%) was used to solve β-carotene (Sigma). Final concentration of β-carotene in medium for proliferation were 0.5, 1.0, 1.5 and 5.0 μM. The results showed that in higher concentration (1.5 μM), β-carotene can alter the morphologycal structure of monolayer epithelial cells of CMT-93. Meanwhile, β-carotene in supra physiological concentration (5.0 μM) significantly (p<0.05) decreased proliferation, exhibited ALDH1A2 gene expression and changed morphological structure.