Flavonoid and isoflavonoid distribution in developing soybean seedling tissues and in seed and root exudates

The distribution of flavonoids, isoflavonoids, and their conjugates in developing soybean (Glycine max L.) seedling organs and in root and seed exudates has been examined. Conjugates of the isoflavones daidzein and genistein are major metabolites in all embryonic organs within the dry seed and in seedling roots, hypocotyl, and cotyledon tissues at all times after germination. Primary leaf tissues undergo a programmed shift from isoflavonoid to flavonoid metabolism 3 days after germination and become largely predominated by glycosides of the flavonols kampferol, quercetin, and isorhamnetin by 5 days. Cotyledons contain relatively constant and very high levels of conjugates of both daidzein and genistein. Hypocotyl tissues contain a third unidentified compound, P19.3, also present in multiple conjugated forms. Conjugates of daidzein, genistein, and P19.3 are at their highest levels in the hypocotyl hook and fall off progressively down the hypocotyl. These isoflavones also undergo a programmed and dramatic decrease between 2 and 4 days in the hypocotyl hook. All root sections are predominated by daidzein and its conjugates, particularly in the root tip, where they reach the highest levels in the seedling. Light has a pronounced effect on the distribution of the isoflavones; in the dark, isoflavone levels in the root tips are greatly reduced, while those in the cotyledons are higher. Finally, the conjugates of daidzein and genistein and several unidentified aromatic metabolites are selectively excreted into root and seed exudates. Analysis of seed exudates suggests that this is a continuous, but saturable event.     

Distribution of isoflavones in lupin hypocotyls. Possible control of cell wall peroxidase activity involved in lignification

The distribution of 4 key isoflavones (luteone, genistein, 2'-hydroxygenistein and wighteone) in lupin ( Lupinus albus L. cv. multolupa) hypocotyls shows a gradient that diminishes from young to old tissues. A spatial gradient occurs within the hypocotyl, and a temporal gradient in both the outermost vascular and epidermal tissues. Not only does a gradient exist in respect to the quantity of isoflavones, but there is also a gradient in respect to the type of isoflavone. Thus, wighteone is mainly associated with the non-meristematic zones of the lupin hypocotyl. A close relationship was found between the distribution and the localization in the walls of phloem cells of both peroxidase (EC 1.11.1.7) and isoflavones. This observation suggests an in vivo peroxidase-isoflavone interconnection. In fact, lupin isoflavones are able to inhibit the peroxidase-catalyzed oxidation of the lignin precursor coniferyl alcohol, probably due to the co-oxidation of isoflavones in the reaction media. The results are discussed on the basis of a possible role for isoflavones in controlling cell wallperoxidase activity involved in the lignification of phloem cells.     

Production of free and glycosylated isoflavones in in vitro soybean (Glycine max L.) hypocotyl cell suspensions and comparison with industrial seed extracts

Glycine max is one of the major sources of phytochemicals, in particular of isoflavones, a class of phytoestrogens with ascertained beneficial effects on human health. In the present study, in vitro callus production from soybean hypocotyl seedling explants and cell suspensions were optimized. Time-courses having 20, 40 and 60 g/L of initial cell inoculum were performed to determine the concentration most suitable for isoflavone production. The amount of total polyphenols and total flavonoids as well as the antioxidant capacity of both cell and culture media fractions were measured by means of spectrophotometric methods. The levels of aglycone and glycosylated isoflavones (didzein, genistein, glycitein, didzin, genistin, glycitin), as well as of ferulic acid, vanillic acid and vanillin, were determined by HPLC–DAD. On average, 93.5 % of the total (cells plus media) isoflavones in soybean cell suspensions were detected as aglycones. Concentrated cell cultures as well as industrial soybean seed extracts were enzymatically hydrolyzed to release the aglycones and their metabolic profiles were analysed by HPLC–DAD. In contrast to cell suspensions, in undigested seed extract the aglycon form represented only 16.8 % of the total isoflavones amount. After enzymatic treatment, the antioxidant capacity increased by 30 and 33 %, respectively, in concentrated cell and seed extracts, demonstrating the presence of a larger amount of bioactive metabolites after digestion. At the present extraction conditions, soybean concentrated cell suspensions yielded 5.8-fold more total isoflavones (mostly in the free form) than seed extracts, leading to hypothesise their possible use as ingredients for cosmetic and nutraceutical applications.     

Seed Coat Retention and Hypocotyl Hook Development in Mutants of Arabidopsis thaliana L.

Germination and growth of wild-type and two mutant strains (aux-1and Dwf) of Arabidopsis thaliana L. have been examined. Seedlingsof aux-1 exhibit agravitropic roots whereas Dwf display bothagravitropic roots and shoots. Wild-type seedlings retained the seed coat at the root-hypocotyltransition zone and developed hypocotyl hooks. In contrast,aux-I and Dwf seedlings did not retain their seed coats andlacked hypocotyl hooks. A positive gravitropic response of theroots was essential for the retention of the seed coat at theroot—hypocotyl transition zone by the attachment of roothairs to the seed coat. The development of the hypocotyl hookwas aided by the retention of the seed coat. The apical regionof the hypocotyl apparently remained agravitropic during formationand maintenance of the hypocotyl hook. Arabidopsis thaliana L., auxins, gravitropism, hypocotyl hook, mutants, peg formation, germination     

Chemotaxis of Bradyrhizobium japonicum to soybean exudates.

The chemotactic response of Bradyrhizobium japonicum toward soybean seed and root exudates was examined. Assays using various isoflavones and fractionated exudate indicated that isoflavones are not the principal attractants in exudates. Likewise, induction of nod genes with isoflavones or seed exudate before assay did not enhance chemotaxis. Screening of numerous compounds revealed that only dicarboxylic acids and the amino acids glutamate and aspartate were strong attractants. The presence of glutamate, aspartate, and dicarboxylic acids in appreciable concentrations in soybean seed and root exudates indicates that these compounds likely represent natural chemoattractants for B. japonicum.     

Chemotaxis of Bradyrhizobium japonicum to soybean exudates.

The chemotactic response of Bradyrhizobium japonicum toward soybean seed and root exudates was examined. Assays using various isoflavones and fractionated exudate indicated that isoflavones are not the principal attractants in exudates. Likewise, induction of nod genes with isoflavones or seed exudate before assay did not enhance chemotaxis. Screening of numerous compounds revealed that only dicarboxylic acids and the amino acids glutamate and aspartate were strong attractants. The presence of glutamate, aspartate, and dicarboxylic acids in appreciable concentrations in soybean seed and root exudates indicates that these compounds likely represent natural chemoattractants for B. japonicum.     

Hypocotyl of seedlings of the large-seeded species Araucaria angustifolia: an important underground sink of the seed reserves

Araucaria angustifolia exhibits cryptogeal germination, where the root–hypocotyl axis emerges first and penetrates into the soil. In Araucaria bidwillii, the whole process of transferring reserves from the seed to the seedling takes place before shoot emergence, and there is a major storage of these reserves in the underground hypocotyl, which assumes a tuberous form. In A. angustifolia, the shoot emerges before seed reserves are depleted. Though it does not grow like a tuber, the hypocotyl of A. angustifolia grows thicker than the adjacent taproot during initial growth, and we hypothesize that it may act as a major sink for seed reserves during this stage. The study tests this hypothesis by evaluating changes in the mass of different plant parts during initial growth. Four harvests were conducted during a ~6-month period to compare the dry mass of different fractions (attached seed, seedling, its shoot and root and the hypocotyl) of seedlings growing under darkness and high light. While seed reserves were still being depleted, the hypocotyl mass showed an initial increase and then a reduction. This was more abrupt when light was available. After seed mass had stabilized, the mass of the hypocotyl continued to decrease in the dark-grown seedlings, but showed a second increase in the light-grown ones. Results confirm the hypothesis that the hypocotyl represents a major sink for the seed reserves of A. angustifolia, acting as an underground storage structure for the growing seedling. Its reserves seem to be important for sustaining initial shoot growth and might also act as a storage sink for photosynthates.     

花生种子内源ABA含量变化及其与活力的关系

花生（Ａｒａｃｈｉｓ ｈｙｐｏｇａｅａ Ｌ．）种子发育过程中,胚轴内源ＡＢＡ 含量一直是增加的;种皮内源ＡＢＡ含量在果针入土后４０ ｄ 最大,然后急剧下降;子叶内源ＡＢＡ 含量在果针入土后６０ ｄ 出现高峰,然后有轻微下降。种子活力指数和萌发时内源ＡＢＡ 的净下降量有密切关系。甘露醇可促进离体胚内源ＡＢＡ 合成,１－甲基－３－苯基－５（３－［三氟甲基］－苯基－４－（１氢）－吡啶）抑制子叶内源ＡＢＡ 的合成,子叶和胚轴存在不同的ＡＢＡ 合成途径。种子早熟和早萌处理时,内源ＡＢＡ 含量都下降,胚轴在种子由发育向萌发转换中起着十分重要的作用     

Antioxidant activity of soya hypocotyl tea in humans

Antioxidative activity of isoflavones has not been shown in humans. Newly-developed isoflavone-rich soya hypocotyl tea contains about 12 mg isoflavones per liter. 15 tea drinkers and 23 control young female students were randomly selected from volunteers, and underwent physical examination, blood chemistry and urinary analysis before and after one month of tea drinking. A three-day dietary record was taken before each physical examination. The tea drinkers showed a lower level of phosphatidylcholine hydroperoxide (PCOOH) and phosphatidyl-ethanolamine hydroperoxide (PEOOH) in the red blood cells and a significant reduction of 8-hydroxydeoxyguanine (8ohdG) in the urine compared to the controls.     

发育和早萌中花生胚胚轴与子叶内BAPAase活性的差异

花生胚发育过程中,子叶和胚轴中都出现BAPAase活性。花生种子萌发时,子叶和胚轴中的BAPAase活性迅速上升,子叶中无新的BAPAase合成,胚轴中能重新合成BAPAase。ABA抑制了子叶和胚轴中BAPAase的活性,抑制胚轴中BAPAase活性所需的外源ABA浓度更高,Act-D和CHM不能逆转ABA对BAPAase活性的抑制作用。     

Changes of Endogenous ABA Content in Relevance to the Vigor of Peanut Seeds

During the development of peanut (Arachis hypogaea L. ) seed, the endogenous ABA content increased steadily'in hypocotyl, increased to a peak at 40 d after pegging with a drastic decline afterwards in testa; and in cotyledon, increased to a peak at 60 d after pegging but with a slight fall afterwards. There seemed to be a close relationship between the increose of vigor index and net loss of endogenous ABA content in the peanut seed germinating in vitro. Osmoticum (mannitol) promoted the endogenous ABA in the cotyledon and hypocotyl. and Fluridone inhibited that in the cotyledon. There were two different paths of the endogenous ABA synthesis in peanut seed, C40 in the cotyledon and C15 in the hypocotyl. When peanut seeds were put in the conditions of precocious maturation or germination tine endogenous ABA content fell down. Result from this experiment concluded that the hypocotyls played an important role in the transition from development to germination of. peanut seed.     

Intricate environment-modulated genetic networks control isoflavone accumulation in soybean seeds

Background  

Soybean (Glycine max [L] Merr.) seed isoflavones have long been considered a desirable trait to target in selection programs for their contribution to human health and plant defense systems. However, attempts to modify seed isoflavone contents have not always produced the expected results because their genetic basis is polygenic and complex. Undoubtedly, the extreme variability that seed isoflavones display over environments has obscured our understanding of the genetics involved.     

Physiological regulation of seed soaking with soybean isoflavones on drought tolerance of Glycine max and Glycine soja

In this study, the Glycine max Jackson cultivar (the relatively drought-sensitive) and the Glycine soja BB52 accession (the drought-tolerant one) were used as the experimental materials. Effects of seed soaking with exogenous soybean isoflavones (daidzin or genistin) on seed germination, and seedlings photosynthesis, relative electrolytic leakage, content of thiobarbituric acid reactive substances , and anti-oxidative activities were investigated under drought stress conditions. The results showed that, treatments of seed soaking with daidzin or genistin could improve seed germination, and alleviate cell damage, enhance anti-oxidative activities and photosynthesis in drought-stressed seedlings, therefore displayed mitigated effects on soybean drought injury, especially for the drought-sensitive G. max Jackson cultivar. Thus seed soaking with exogenous soybean isoflavones may be a usable approach to enhance drought tolerance of cultivated soybean in practice.     

A film study of growth activity of hypocotyl in the early phases of the germinating pea seed (Pisum sativum L.)

Using the time-lapse cinematography technique, activity of the growing centres of the pea seed hypocotyl (Pisum sativum L.) was followed in the course of the early phases between 15 and 60 hours of germination under standard experimental conditions. The resulting data were plotted in a summary graph with conjugates scales (Fig. 4) indicating time-space limits for further biochemical analysis and the interpretation of mechanism of hypocotyl elongation concerning the dependence of the IAA and RNA content, growth rate, localization of growing zones, transport of substances from the swollen cotyledons into the embryo etc. The most important results are as follows: Between 15 and 20 hours of seed swelling, activity is manifested in two growing centres of the hypocotyl (Fig. 5); between 30 and 36 hours only one zone becomes localized, with a maximum elongation in the region which was originally at a distance of 2·5 mm. from the hypocotyl base. The greatest elongation of the hypocotyl—0·77 mm./hour— was observed between 30 and 48 hours in the zone which was originally at a distance of 2·75 mm. from the hypocotyl base. The zone of maximum growth did not become localized by the termination of the experiment (60 hours) in the presumed zone close to the apex, but was displaced further to the zone which was originally at a distance of 4·0 mm. from the hypocotyl base.     

芡个体发育早期的研究

本文研究了芡个体发育的早期, 即心形胚至种苗。心形胚至成熟胚表现为:苗端先发育, 根端弱育;胚芽叶节上的节生根原基先发育, 根端无明显分化。种子胚至种苗表现为:种子萌发时, 下胚轴末端产生多细胞分枝下胚轴毛;种苗形成中, 节生根先发育, 胚根后发育, 且长达1mm左右即停止生长。这些器官发育顺序上的特点在被子植物中是很特殊的, 应该是系统发生上的原始性状。下胚轴毛是水生或湿生被子植物比较普遍的性状, 是区分下胚轴与胚根的指示性状。     

The 7B-1 mutant in tomato shows blue-light-specific resistance to osmotic stress and abscisic acid

Germination of wild-type (WT) tomato ( Lycopersicon esculentum Mill.) seed is inhibited by mannitol (100-140 mM) in light, but not in darkness, suggesting that light amplifies the responsiveness of the seed to osmotic stress (M. Fellner, V.K. Sawhney (2001) Theor Appl Genet 102:215-221). Here we report that white light (W) and especially blue light (B) strongly enhance the mannitol-induced inhibition of seed germination, and that the effect of red light (R) is weak or nil. The inhibitory effect of mannitol could be completely overcome by fluridone, an inhibitor of abscisic acid (ABA) biosynthesis, indicating that mannitol inhibits seed germination via ABA accumulation in seeds. The inhibition of WT seed germination by exogenous ABA was also amplified by W or B, but not by R. In a recessive, ABA-overproducing, 7B-1 mutant of tomato, seed germination and hypocotyl growth were resistant to inhibition by mannitol or exogenous ABA, both in W or B. Experiments with fluridone suggested that inhibition of hypocotyl growth by W or B is also partially via ABA accumulation. De-etiolation in the mutant was especially less in B compared to the WT, and there was no difference in hypocotyl growth between the two genotypes in R. Our data suggest that B amplifies the responsiveness of tomato seeds and hypocotyls to mannitol and ABA, and that W- or B-specific resistance of the 7B-1 mutant to osmotic stress or ABA is a consequence of a defect in B perception or signal transduction.     

桃儿七种子解剖结构及其萌发生长期形态特征

为探明种皮和胚乳是否是限制桃儿七种子萌发的主要因素,利用组织切片和显微技术,对桃儿七种子及其不同萌发期（1、7、14、21、28 d）解剖结构和播种后一定时期内（7~210 d）的植株生长形态进行观察。桃儿七种子由种皮、胚乳和胚构成。种皮包括外种皮和内种皮,外种皮致密规整,由外至内分别为栅状石细胞和表皮层细胞,内种皮由5~6层海绵细胞组成。胚乳占种子体积的绝大部分,包括珠孔胚乳和外胚乳。胚由胚根、胚轴和子叶组成,被致密种皮、多层珠孔胚乳和外胚乳包围。萌发期1~7 d胚根和胚轴开始伸长,7~14 d两片子叶分离,14~21 d胚根突破珠孔胚乳和种皮,21~28 d胚根、胚轴和子叶继续扩张伸长。种子播种210 d后可平均形成3片功能真叶和5条不定根。致密种皮（物理休眠）和多层胚乳（机械休眠）是限制桃儿七种子萌发的两个主要因素。