To evaluate the association between serum carotenoids and quantitative measures of retinal vasculature in elderly Singapore Chinese subjects. The following details were collected in 128 healthy subjects: sociodemographics, lifestyle information, medical and drug history, and anthropometric measurements. Serum concentrations of carotenoids were estimated in fasting venous blood using high performance liquid chromatography. Retinal vascular parameters were quantitatively measured from retinal photographs using a computer-assisted program (Singapore I Vessel Assessment). The mean age of the population was 54.1 years (range 40 to 81 years). In multiple linear regression analysis, per SD decrease in retinal arteriolar caliber [β = 0.045 (0.003 to 0.086), p = 0.036], per SD increase in retinal venular caliber [β = -0.045 (-0.086 to -0.003), p = 0.036] and per SD increase in arteriolar branching angle [β = -0.039 (-0.072 to -0.006), p = 0.021] were associated with decreased serum lutein. Per SD increase in retinal venular tortuosity [β = -0.0075 (-0.0145 to -0.0004), p = 0.039] and per SD increase in arteriolar branching angle (β = -0.0073 [-0.0142 to -0.0059], p = 0.041) were associated with decreased serum zeaxanthin. None of the other carotenoids demonstrated meaningful relationship with quantitative measures of retinal vasculature. Lower levels of lutein and zeaxanthin demonstrated significant relationship with adverse quantitative measures of retinal vasculature in elderly healthy subjects.