Effect Of Genotype And Nitrogen On Grain Phenolic Compound And Amino Acid Of Japonica Rice

Rice,with high nutritional value, is one of the major food crops on earth and of importance for improving human health. Cereals are rich in physiological active components, for example, phenolic compounds, vitamins, mineral, and fiber. In the present study, we conducted a field experiment:two japonica rice cultivars, Wuyunjing7 and Wuyujing3, and the mutants of Wuyujing3 induced by EMS were used, nitrogen fertilization were applicated with different rate and timing. The objective of this study were to elucidate the effect of genotype and nitrogen fertilization on rice grain nutrition quality. We hope the results obtained would be helpful for rice improvement aiming at high nutrition quality.The main results and conclusions are as follows:1. Determination of phenolic acids in rice by ultra-high performance liquid chromatographyThis study aims to develop an ultra-high performance liquid chromatography method for the determination of phenolic acids in brown rice and milled rice, and analyze the tissue distribution of phenolic acids in rice grain. Standard compounds of the target phenolic acids, including gallic acid, protocatechuic acid, gentisic acid,2,5-dihydroxybenzoic acid, 4-hydroxybenzoic acid, chlorogenic acid, vanillic acid, caffeic acid, syringic acid, p-coumaric acid, ferulic acid, sinapic acid, isoferulic acid,2-hydroxycinnamic acid, and trans-cinnamic acid, were purchased from Sigma-Aldrich. The liquid-chromatographic separation was performed on Agilent ZORBAX Eclipse Plus C18 (2.1*50 mm,1.8 μm-Micron) column kept at 30℃, using acetonitrile/acetic acid/water mixture as the mobile phase with a flow rate of 0.5 mL·min-1 through gradient elution, and the detection wavelength was set at 280 nm and 325 nm. Results showed that a good separation of 14 phenolic acids was achieved within 8 min. Eleven phenolic acids were detected in brown and milled rice by mass spectrometry, with eight of them can be used for quantitative analysis. The proposed method exhibited a linear range of 5-220μg·mL-1 (r2=0.9994-0.9999), with limits of quantification ranging from 0.002 to 0.03μg·mL-1, recovery rates from 84.11% to 114.43%, and relative standard derivation ranging from 1.28% to 12.94%. Significant difference existed in the distribution of 4-hydroxybenzoic acid, vanillic acid, syringic acid, p-coumaric acid, ferulic acid, sinapic acid, isoferulic acid, trans-cinnamic acid between brown and milled rice, with 4-hydroxybenzoic acid, vanillic acid, syringic acid, p-coumaric acid, ferulic acid, sinapic acid, trans-cinnamic acid mainly concentrated in the outer endosperm, whereas a considerable part of isoferulic acid accumulated in the endosperm. This method is simple, rapid, accurate, reliability, and can be used for the determination of phenolic acids in rice grain.2. Effect of genotype and N fertilization on phenolic compounds of japonica riceTo investigate the influence of panicle nitrogen fertilization on phenolic compounds of japonica rice, a field experiment was conducted:two japonica rice cultivars, namely Wuyunjing7 and Wuyujing3, were used and nitrogen fertilization was applied as spikelet-promoting and spikelet-sustaining, respectively. And three levels (0,75,150 kg.hm-2) of N-fertilization application were conducted. The results showed significant effect of nitrogen panicle fertilization on the content of total phenolic contents (TPCs), DPPH scavenging activity (DPPH·), Oxygen radical absorbance capacity (ORAC). But such effect was not the same among different compounds, because of variety difference, the change of the total phenolic contents was not obvious. Antioxidant activity of Wuyujing3 was greatly influenced by nitrogen. Despite the nitrogen application timing, with the treatment of nitrogen, the TPCs of Wuyujing7 increased in both brown and milled rice; While, that of Wuyujing3 only increased in milled rice. And when nitrogen was applied as spikelet-promoting, the TPCs of Wuyujing3 in brown rice increased. DPPH’and ORAC in milled rice and brown rice affected by nitrogen were not consistent. With high level of nitrogen input, the DPPH of milled rice decreased, in brown rice the opposite. With low level of nitrogen input, the DPPH of Wuyujing3 decreased in both milled and brown rice, which the DPPH of Wuyujing7 increased. ORAC increased with high level fertilization applied as spikelet-promoting. Wuyunjing7 had relative higher TPCs and ORAC than Wuyujing3. It was beneficial to improve the antioxidant quality with high level fertilization applied as spikelet-promoting.3. Effect of genotype and nitrogen on amino acidsTo investigate the influence of nitrogen fertilization mode on grain amino acids, Wuyujing 3 and 11 mutants of it with contrasting agronomic traits were used and two treatments of nitrogen with the ratios of basal to topdressing (N5:5 and N10:0) were performed. The results showed the contents of 17 amino acids in W31 and W84 were higher than the wild-type. Among all the materials used, the total amino acid of W31 and W84 were ranged to the highest, no matter with or without nitrogen fertilization input. The contents of threonine and lysine had a tiny difference among genotypes, and methionine had a significant difference among genotypes. Compared with wild-type, the content of methionine in W31 increased 43.6%. With nitrogen fertilization input, the content of methionine increased prominently, and which in W80, W38, W10 increased 51.00%、 42.68%、36.57%. Among all the mutants, W34, W62 had a higher amino acid content and better nutritional value. In addition, histidine, phenylalanine, tyrosine, arginine were great influenced by nitrogen, but cystine was slight influenced by nitrogen.