The aim of this study was to evaluate the effect of a synbiotic containing Lactobacillus acidophilus ATCC 4962, fructooligosaccharide, inulin and mannitol on plasma lipid profiles and erythrocyte membrane properties in hypercholesterolaemic pigs on high- and low-fat diets. Twenty-four white male Landrace pigs were randomly allocated to four treatment groups for 8 weeks (n 6). Treatment factors were the supplementation of synbiotic (with and without) and dietary fat (5 and 15 %). The supplementation of synbiotic reduced plasma total cholesterol (P = 0·001), TAG (P = 0·002) and LDL-cholesterol (P = 0·045) for both dietary fats. A higher concentration of esterified-cholesterol in HDL of pigs supplemented with synbiotic than the control regardless of dietary fat (P = 0·036) indicated that cholesterol was reduced in the form of cholesteryl esters. Reduced concentration of cholesteryl esters (P < 0·001) and increased concentration of TAG (P = 0·042) in LDL of pigs on synbiotic suggested that LDL-cholesterol was reduced via the hydrolysis of smaller and denser LDL particles. The erythrocytes of pigs without any synbiotic showed more prevalence of spur cells than those given the synbiotic, as supported by the higher cholesterol: phospholipid ratio in erythrocytes (P = 0·001). Also, membrane fluidity and rigidity were improved as supported by the decreased fluorescence anisotropies in the Hb-free erythrocyte membrane of pigs given synbiotic (P < 0·001). The administration of the synbiotic reduced plasma TAG, total cholesterol and LDL-cholesterol in hypercholesterolaemic pigs, possibly in the form of cholesteryl esters, via the interrelated pathways of lipid transporters (VLDL, LDL and HDL). The synbiotic also reduced deformation of erythrocytes via improved membrane fluidity and permeability.