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The effect of a singular amino acid, asparagine (Asn), glutamine (Gln), or proline deletion in a cultured medium (RPMI 1640 supplemented with 10% fetal calf serum and other ingredients) on adriamycin (ADR) cytotoxicity was evaluated in the growth of P388 murine leukemia cells and CEM human acute lymphoblastic leukemia cells over a 3 day period. No enhancement of ADR cytotoxicity was observed in the assay of IC50 values under the amino acid deleted condition. Singular deletion of Gln or Asn from ADR-free medium apparently inhibited the proliferation of both cells, i.e. both cell lines strongly require them. The cytotoxicity of 5 nM ADR was then examined in medium which included one or the other of them in stepwise levels varied at 80, 60, 40, 20 and 0% of the ordinary level. Change of Asn level caused a difference in ADR toxicity; also, the change of Gln level, especially the 60% level caused ADR toxicity of 5 nM, which is less than the IC50 value, in the proliferation of both cells. This suggested the usefulness of glutamine level modification on the enhancement of ADR cytotoxicity.