Aim To study the effect of phosphalipids on the cholesterol efflux from rat macrophages mediated by HDL 3. Methods Measure the cholesterol content, cellular phospholipid content, and the balance between free cholesterol and cholesteryl ester when phosphatidylcholine (PC) or sphingomyelin (SPM) was added to the mediun. Results The cholesterol content changed from 79.95 μg/plate to 42.79, 37.16 and 31.53 in HDL 3, HDL 3+PC and HDL 3+SPM group respectively. The cellular phosphorous content in PC (PC-p) were 13.95 and 16.28 μg/plate in HDL 3 and HDL 3+PC group respectively. The cellular phosphorous content in SPM (SPM-p) were 5.03 and 7.01 μg/plate in HDL 3 and HDL 3+SPM group respectively. After macrophages were incubated with BSA, PC and SPM alone, the cellular cholesterol content were 75.30, 74.65 and 73.09μg/plate respectively and the ratio of free cholesterol to total cholesterol were 59.42%, 56.95% and 64.77% respectively. Conclusion PC and SPM couldn't mediate directly cellular cholesterol efflux, but they could enhance cellular cholesterol efflux from macrophages mediated by HDL 3, and the ability of SPM was stronger than PC. SPM could induce cholesteryl ester convert into free cholesterol. In macrophages and PC could induce free cholesterol convert into cholesteryl ester.