Polyphenols In Rice(Oryza Sativa L.):Identification,Distribution,Genetics And Gene Exoression

Phytochemicals such as phenolics and flavonoids in rice grain are antioxidants that can reduce the risks of developing chronic diseases including cardiovascular diseases, type-2diabetes, obesity, and some cancers. In this study, we identified some phenolic acids and anthocyanins, and clarified the accumulation and distribution pattern in white, red, and black rice grain. The genetics and gene expression were also performed. The main results are shown as follows:1. Many common phenolic acids, such as protocatechuic, p-hydroxybenzoic, vanillic, syringic, trans-coumaric, ferulic and trans-sinapic acid in rice grain were detected by HPLC-QTOF-MS/MS, among which protocatechuic acid existed in red and black rice, and vanillic acid only existed in black rice. Besides these common phenolic acids,3rare phenolic acids,3coupled diferulic acids, and1rare anthocyanin in balck rice were also identified. The B-type of proanthocyanidins was found only in red rice.2. The accumulation of polyphenol and its antioxidant capacity at different development stages after flowering were different among white (9311), red (SB7), and black rice grain (Yunanheixiannuo). Unlike TPC of white and red rice which was significantly higher at1-week stage, TPC of black rice was highest at maturity. In white rice, ferulic and isoferulic acid were significantly higher at3-week stage and matured stage, respectively. In red rice, protocatechuic,p-coumaric, ferulic, and isoferulic acid were significantly higher at1-week stage. In black rice, vanillic and ferlic acid were significantly higher at matured stage, while,p-coumaric and syringic acid were higher at1-week stage. Cyanidin-3-glucoside and peonidin-3-glucoside were the main anthocyanins in black rice showing significantly higher levels at2-and3-weeks.3. The distributions of polyphenols were different in white, red, and black grains. TPC was highest in the bran, which accounted for60-86%of whole grains, and mainly existed in free and conjugated, and bound form. Free and conjugated phenolic acids in white, red and black bran accounted for41,65and85%of total acids in whole grain, and that in embryo accounted for59,35, and15%, respectively. Bound phenolic acids in rice bran accounted for90%of total acids in whole grain. Total anthocyanin, cyanidin-3-O-glucoside and peonidin-3-O-glucoside were identified mainly in black rice bran which accounted for more than93%.4. Analyses of the effects of genotype, environment, and their interaction showed that TPC, TFC, TPAC, ABTS and DPPH were mainly affected by genotypic variance which accounted for about95%. Although the environmental and G×E effects accounted for about5%, they were at significant levels (P<0.001). High correlations among these five parameters were observed in Proc corr analysis (P<0.001). The rice materials were divided into three subgroups:Japonica, Indica and aus. By using association study, twenty-three putative unique loci were identified in two years. Among them, five loci (qPAC7-3, qPC4, qPC12, qFCl-1and qPAC2-1) were close to previously identified genes or QTLs detected by the linkage or association mapping.5. The accumulation of anthocyanin was controlled by many genes, which had temporal and spatial expression specificities. OsPAL, OsCHS and OsCHI were the basic genes involved in anthocyanin biosynthesis. OsF3H, OsF3’H, OsDFR, OsANS, OsBl, OsB2and OsCl had different expression patterns in stems, leves, flowers, and seeds of white, red, black, and purple-leaf with white grain (IR1552) rice plants, some of which were associanted with the development time. We found3indels by comparing Ra sequence between black grain rice and purple-leaf with white grain rice. According to these polymorphisms, we designed three markers, which were used to detect the genotyope of the RIL of balck-grained rice and white-grained rice. We found that all the balck-grained rice had the genotype of Ra/Ra, while brown-grained rice had the genotype of Ra/Ra or Ra/ra, and white-grained rice had the genotype of ra/ra. By RT-PCR, we found that Ra could be digested by BamW I in brown-grained and black-grained rice plants with different expression patterns. For the former, it expressed higher in stem and at12-20days after flowering, and for the latter it expressed higher in seedling and at16-26days after flowering. Knockout of Ra by RNAi showed that the knockout of Ra did not lead to the color change of black bran although C1and Ra together lead to the pigmentation in aleurone layer of rice grain.