The aim of this study was to provide a basis for studying the molecular mechanism of pharmacological action of chlorhexidine digluconate. Fluorescence polarization of n-(9-anthroyloxy)stearic acid was used to examine the effect of chlorhexidine digluconate on differential rotational mobility of different positions of the number of membrane bilayer phospholipid carbon atoms. The six membrane components differed with respect to 2, 3, 6, 9, 12, and 16-(9-anthroyloxy)stearic acid (2-AS, 3-AS, 6-AS, 9-AS, 12-AS and 16-AP) probes, indicating different membrane fluidity. Chlorhexidine digluconate increased the rate of rotational mobility of hydrocarbon interior of the cultured Porphyromonas gingivalis outer membranes (OPG) in a dose-dependent manner, but decreased the mobility of surface region (membrane interface) of the OPG. Disordering or ordering effects of chlorhexidine digluconate on membrane lipids may be responsible for some, but not all of its bacteriostatic and bactericidal actions.