Folic is essential for sustaining cell activity, promoting cell growth, and participating in cell division and proliferation. The demand for colored wheat is increasing due to its richness in anthocyanin, iron, zinc, selenium and other useful minerals. To research the content of folate in colored wheat grains,this study employed a total of 42 wheat varieties (lines) with varying grain colors as materials to ascertain the contents of 4 folic acid derivatives in 3 grains under 3 growing conditions: tetrahydrofolate (THF), 5-methyltetrahydrofolate (5-CH3-THF), 5-formyltetrahydrofolate (5-CHO-THF) and 5,?10-methylenetetrahydrofolate (5,?10-CH+THF). The results indicated that the 4 folic acid derivatives were distributed across wheat varieties with varying grain colors, with a coefficient of variation ranging from 18.34% to 21.10%, among which Lin 4179 had high folic acid content, and with a total content of 66.99 μg·100 g-1. The contents of 5-CH3-THF and 5-CHO-THF accounted for approximately 70% of the total folic acid content. The mean total folate level in purple and blue grains was 62.31 and 62.97 μg·100 g-1, respectively, which was significantly higher than that in white (41.90 μg·100 g-1) and red grains (44.36 μg·100 g-1). Genotype effect was the main factor affecting total folic acid content, environmental factors have less impact. The results of this study provided an important insight for the development and breeding of folate biofortified wheat varieties.