The health-promoting effects of whole-grain consumption have been attributed in part to their unique phytochemical contents and profiles that complement those found in fruits and vegetables. Wheat is an important component of the human diet; however, little is known about the phytochemical profiles and total antioxidant activities of milled fractions of different wheat varieties. The objectives of this study were to investigate the distribution of phytochemicals (total phenolics, flavonoids, ferulic acid, and carotenoids) and to determine hydrophilic and lipophilic antioxidant activity in milled fractions (endosperm and bran/germ) of three different wheat varieties, two of which were grown in two environments. Grain samples of each of the wheat varieties were milled into endosperm and bran/germ fractions. Each fraction was extracted and analyzed for total phenolics, ferulic acid, flavonoids, carotenoid contents, and hydrophilic and lipophilic antioxidant activities. Total phenolic content of bran/germ fractions (2867-3120 micromol of gallic acid equiv/100 g) was 15-18-fold higher (p < 0.01) than that of respective endosperm fractions. Ferulic acid content ranged from 1005 to 1130 micromol/100 g in bran/germ fractions and from 15 to 21 micromol/100 g in the endosperm fractions. The bran/germ fraction flavonoid content was 740-940 micromol of catechin equiv/100 g. On average, bran/germ fractions of wheat had 4-fold more lutein, 12-fold more zeaxanthin, and 2-fold more beta-cryptoxanthin than the endosperm fractions. Hydrophilic antioxidant activity of bran/germ samples (7.1-16.4 micromol of vitamin C equiv/g) was 13-27-fold higher than that of the respective endosperm samples. Similarly, lipophilic antioxidant activity was 28-89-fold higher in the bran/germ fractions (1785-4669 nmol of vitamin E equiv/g). Hydrophilic antioxidant activity contribution to the total antioxidant activity (hydrophilic + lipophilic) was >80%. In whole-wheat flour, the bran/germ fraction contributed 83% of the total phenolic content, 79% of the total flavonoid content, 51% of the total lutein, 78% of the total zeaxanthin, 42% of the total beta-cryptoxanthin, 85% of the total hydrophilic antioxidant activity, and 94% of the total lipophilic antioxidant activity. Our results showed that different milled fractions of wheat have different profiles of both hydrophilic and lipophilic phytochemicals. These findings provide information necessary for evaluating contributions to good health and disease prevention from whole-wheat consumption.