Abstract Background Consumption of a cholesterol lowering dietary portfolio including plant sterols (PS), viscous fibre, soy proteins and nuts for 6 months improves blood lipid profile. Plant sterols reduce blood cholesterol by inhibiting intestinal cholesterol absorption and concerns have been raised whether PS consumption reduces fat soluble vitamin absorption. Objective The objective was to determine effects of consumption of a cholesterol lowering dietary portfolio on circulating concentrations of PS and fat soluble vitamins. Methods Using a parallel design study, 351 hyperlipidemic participants from 4 centres across Canada were randomized to 1 of 3 groups. Participants followed dietary advice with control or portfolio diet. Participants on routine and intensive portfolio involved 2 and 7 clinic visits, respectively, over 6 months. Results No changes in plasma concentrations of α and γ tocopherol, lutein, lycopene and retinol, but decreased β-carotene concentrations were observed with intensive (week 12:p = 0.045; week 24:p = 0.039) and routine (week 12:p = 0.031; week 24:p = 0.078) portfolio groups compared to control. However, cholesterol adjusted β-carotene and fat soluble compound concentrations were not different compared to control. Plasma PS concentrations were increased with intensive (campesterol:p = 0.012; β-sitosterol:p = 0.035) and routine (campesterol: p = 0.034; β-sitosterol: p = 0.080) portfolio groups compared to control. Plasma cholesterol-adjusted campesterol and β-sitosterol concentrations were negatively correlated (p