Yoichi Nishiba

Fluorometric Determination of a 1,3-Diethyl-2-thiobarbituric Acid–Malondialdehyde Adduct as an Index of Lipid Peroxidation in Plant Materials

A fluorometric method has been developed to measure a 1,3-diethyl-2-thiobarbituric acid (DETBA)-malondialdehyde (MDA) adduct as an index of lipid peroxidation in plant materials. Plant tissue samples were prepared under nitrogen gas and then added to an assay system containing butylated hydroxytoluene. Following the reaction between DETBA and the plant tissue samples, the DETBA-MDA adduct was extracted with ethyl acetate and measured by spectrofluorometry or high-performance liquid chromatography (HPLC) with a fluorescence detector. The species of influencing substances with spectrofluorometry were fewer and their interfering concentration was higher than that by traditional colorimetry. When this method was applied to plant materials, the detection limits for the DETBA-MDA adduct were 2.5 nmol/g of fresh weight and 0.0625 nmol/g of fresh weight by spectrofluorometry and HPLC with a fluorescence detector, respectively. Using this sensitive, specific and simple fluorometric method, DETBA-MDA adducts ranging from 0.8 to 18.0 pmol/g of fresh weight could easily be detected from vegetables, fruits, and potatoes.

Use of Near-infrared Reflectance Spectroscopy for the Estimation of the Isoflavone Contents of Soybean Seeds

The consumers’ needs for agricultural products are now widely diversifi ed with attention focused not only on the major constituents, but also on the physiological functions. In Japan, soybean (Glycine max L.) is used to produce various foods. Increase of nutritional functions is important to promote the consumption of soybean. However, the conventional method for estimation of elements of physiological functions such as isoflavone is labor intensive. Isoflavones may help prevent osteoporosis (Yamori, 2001). A simple and rapid method for estimating their contents is necessary for screening soybean varieties for breeding. Near infrared spectroscopy (NIRS) has been used as one of the most powerful analytical tools in the agricultural and food fi elds (Williams, 2006). NIRS is an offi cial method for analyzing the major constituents of soybean in the trade based on the quality in USA. Then, if other criteria can be estimated by the NIRS method, this method will become more useful. In this study, the feasibility of NIRS for the estimation of the isofl avone contents of soybean seeds was examined.

Effect of repeated harvesting on the content of caffeic acid and seven species of caffeoylquinic acids in sweet potato leaves

The purpose of this study was to investigate the effect of repeated harvesting on the content of caffeic acid (CA) and seven species of caffeoylquinic acids (CQAs) in sweet potato leaves using a newly developed high-performance liquid chromatography method. Six cultivars and two breeding lines were used in this study. Leaves were collected at monthly intervals from 1st harvest (May) to 4th harvest (August) in 2011 and 2012. ANOVA analysis revealed that the contents of CQAs were significantly different among all cultivars and breeding lines, but no significant differences were found for CA. No annual variation was confirmed in CA and CQAs. Repeated harvest of sweet potato leaves affected the content of only 4-CQA and 5-CQA. Post-hoc comparisons using Tukey’s method indicated that the contents of 4-CQA and 5-CQA in sweet potato leaves harvested at first time were significantly higher compared to those at the other harvest times. Graphical Abstract Contents of the article titled “Effect of repeated harvesting on the content of caffeic acid and seven species of caffeoylquinic acids in sweet potato leaves”

Buckwheat R2R3 MYB transcription factor FeMYBF1 regulates flavonol biosynthesis

Buckwheat (Fagopyrum esculentum) contains high amounts of flavonoids, especially flavonols (e.g., rutin), which are thought to be highly beneficial for human health. Little is known, however, about the regulation of flavonol synthesis in buckwheat. We identified a buckwheat gene encoding an R2R3 MYB transcription factor, and named this gene FeMYBF1. Analysis of the deduced amino acid sequence and phylogenetic analysis suggested that FeMYBF1 encodes an ortholog of the Arabidopsis flavonol regulators AtMYB11, AtMYB12 and AtMYB111. Expression of FeMYBF1 in a flavonol-deficient Arabidopsis triple mutant (myb11 myb12 myb111) restored flavonol synthesis. Constitutive expression of FeMYBF1 driven by the CaMV 35S promoter in Arabidopsis resulted in over-accumulation of flavonol glycosides and upregulation of the expression of AtFLS1. Transient expression assays showed that FeMYBF1 activated the promoter of the Arabidopsis gene encoding AtFLS1, and the promoters of buckwheat genes related to anthocyanin and proanthocyanidin synthesis such as dihydroflavonol 4-reductase (DFR) and leucoanthocyanidin dioxygenase (LDOX) in addition to genes encoding FLS. The results indicate that FeMYBF1 regulates flavonol synthesis and may have a role in synthesis of other flavonoid compounds, and also that buckwheat may have alternative pathway of flavonol synthesis through DFR and LDOX.