Dimethylhydrazine (DMH) is a potent procarcinogen with selectivity for the colon. To determine whether alterations in the lipid composition and fluidity of rat colonic brush border membranes existed before the development of DMH-induced colon cancer, rats were injected s.c. with this agent (20 mg/kg body weight per wk) or diluent for 5, 10, and 15 wk. Animals were killed at these time periods and brush border membranes were prepared from proximal and distal colonocytes of each group. The "static" and "dynamic" components of fluidity of each membrane were then assessed, by steady-state fluorescence polarization techniques using limiting hindered fluorescence anisotropy and order parameter values of the fluorophore 1,6 diphenyl-1,3,5-hexatriene (DPH) and fluorescence anisotropy values of DL-2-(9-anthroyl) stearic acid and DL-12-(9-anthroyl) stearic acid, respectively. Membrane lipids were extracted and analyzed by thin-layer chromatography and gas-liquid chromatography. Phospholipid methylation activity in these membranes was also measured using S-adenosyl-L-methionine as the methyl donor. The results of these studies demonstrate that: the lipid composition and both components of fluidity of proximal DMH-treated and control membranes and their liposomes were similar at all time periods examined; at 5, 10, and 15 wk the "dynamic component of fluidity" of distal DMH-treated membranes and their liposomes was found to be higher, similar, and lower, respectively, than their control counterparts; the "static component of fluidity" of distal DMH-treated membranes and their liposomes, however, was similar to control preparations at all three time periods; and alterations in the lipid composition and phospholipid methylation activities appeared to be responsible for these differences in the "dynamic component of fluidity" at these various time periods.