The risk of infection by HIV-1 through transfusion of contaminated blood products has been markedly decreased but not eliminated by serological screening of donors. Methods are required to further minimize or eliminate the risk of infection of blood product recipients. We therefore examined the capacity of alkylureas to inhibit infectivity of HIV-1. Incubation of free HIV-1 virions with alkylureas suppressed their infectivity, and the minimal inhibitory concentration of the alkylureas was related to the length of the alkyl chain. Butylurea, the most potent inhibitor of HIV-1, inhibited the infectivity of 105median tissue culture infective dose (TCID)50of HIV-1, chronically HIV-1-infected H9 cells and mononuclear cells from two HIV-1-infected patients. Size fractionation of HIV-1 following incubation with butylurea indicated that the structure of the virus was disrupted by butylurea. This study demonstrates that butylurea, at a concentration that has been shown not to affect red blood cell function, can inhibit infectivity of extracellular and intracellular HIV-1. Since the HIV-1 inhibitory capacity of the alkylureas increases with the length of the alkyl side chain, it is likely that hydrophobic interactions between the alkylureas and HIV-1 are responsible for the observed effect.