Short-chain fatty acids as sodium salts (pH 7.0) induced hemoglobin synthesis in cultured Friend leukemia cells (T-3-Cl-1) in concentrations of 0.5 mM butyrate, 6mM isobutyrate, 2mM propionate, and 30mM acetate. Relative number of cells decreased by the addition of these fatty acids. Addition of 0.5mM butyrate to culture medium resulted in the highest hemoglobin synthesis as found by the extracted hemoglobin content by spectrophotometry. The activity of delta-aminolevulinic acid synthetase increased on day 4 by the addition of any of these fatty acids into the medium. Hemoglobin synthesis was initiated on day 3 of culture by the addition of either butyrate or isobutyrate, and on day 4 by the addition of dimethyl sulfoxide. Hemoglobin accumulation reached the maximum on day 5 of culture in all cases when the fatty acid was added. Reasonably similar values were obtained for hemoglobin synthesis determined by either benzidine staining or hemoglobin quantitation. Formate, succinate, and citrate were ineffective for the induction.