Spray treatment of leaves with Fe 2+ promotes procyanidin biosynthesis by upregulating the expression of the F3H and ANS genes in red rice grains ( Oryza sativa L.)

Red rice (RR) is bred for the high-quality proanthocyanidins (PAs) deposition and colouration in the pericarp, which contribute to its potential as a functional food ingredient. The study has investigated the pigment characteristics attributed to PAs enriched in grains with a 2:1 ratio of oligomeric to polymeric PAs. Based on metabolomic analysis, we characterized six marker PAs responsible for grain pigment, including three procyanidins and three prodelphinidins/propelargonidins. During production, cultivation environments substantially affect the grain redness quality. To relieve these negative impacts on RR high-PA characteristics, we have employed a Fe 2+ -spraying cultivation method. When RR was treated by the leaf FeSO 4 spraying, it exhibited a significant increase in the PA contents in grains, particularly for polymeric components, which increased twofold. Through transcriptomic analysis and molecular validation, we determined that the foliar-applied FeSO 4 significantly affected the flavonoid pathway, flowing to substantial substrates of (epi)catechin units by upregulating the expression of F3H and ANS, thereby leading to the grain procyanidin accumulation. Foliar-applied FeSO 4 can be employed as an effective cultivation method to enhance the high-PA characteristics of cereal crops.