In vitamin A-replete populations, increased concentrations of serum carotenoids have been associated with a decreased risk of degenerative diseases. The mechanism of action of carotenoids in determining antioxidant activity is largely unknown. The aim of the study was to examine the effect of carotenoid supplementation and spinach intake on erythrocyte enzyme antioxidant activities, serum or plasma nonenzymatic antioxidant concentrations, and concentrations of oxidatively damaged amino acids in plasma. Subjects received for 3 wk a basic diet (n = 10), a basic diet with a carotenoid supplement (n = 12) or with a spinach product (n = 12 per group), i.e., whole-leaf, minced, liquefied or liquefied spinach plus added dietary fiber. After 3 wk of dietary intervention, changes in serum or plasma concentrations of ascorbic acid, alpha-tocopherol, FRAP (ferric reducing ability of plasma) and uric acid and erythrocyte enzyme activities were assessed, and differences among experimental groups were tested. Consumption of spinach resulted in greater (P < 0.01) erythrocyte glutathione reductase activity and lower (P < 0.05) erythrocyte catalase activity and serum alpha-tocopherol concentration compared with the control group. Consumption of the carotenoid supplement led to lower alpha-tocopherol responses (P = 0.02) compared with the basic diet only. Our data suggest that the short-term changes in erythrocyte glutathione reductase activity and serum alpha-tocopherol concentration can be attributed to an increased carotenoid (lutein and zeaxanthin) intake, but beta-carotene is unlikely to be a causative factor. Lower erythrocyte catalase activity after intervention with spinach products may be related to other constituents in spinach such as flavonoids.