The effects of selected antitumor agents--methotrexate, 6-mercaptopurine, cyclophosphamide, hydroxyurea and cytosine arabinoside on hemopoietic progenitor cells were determined by femoral and splenic colony forming cell (CFC) content after single and multiple injections of the agents. Mice were injected in groups of three with the agents on different days and killed on the same day. Major findings were: 1) All the antitumor agents (by single or multiple injections), with the exception of 6-mercaptopurine, caused an early reduction in the number of CFC per femur and an increase in CFC levels per spleen on day 7.2)CFC levels per 10(5) cells were significantly lower at 1 hour after single injections of hydroxyurea or methotrexate than at 24 hours after a single injection of either drug. 3) Multiple vs. single doses of hydroxyurea and cytosine arabinoside showed that multiple doses were more effective in reducing femoral CFC levels on day 1 and elevating splenic CFC content on day 7 than were single doses. 4) Multiple doses of 6-mercaptopurine were more effective than single doses in lowering femoral CFC levels on day 1, but all doses of 6-mercaptopurine failed to evoke a change in splenic CFC levels. 5) Multiple doses of methotrexate were not more effective than single doses in decreasing femoral CFC content and raising splenic CFC levels.