Biochemical aspects of growth-stimulating effects of steroid phytohormones on lupine plants

Presowing treatment of seeds of lupine of various species and cultivars with brassinosteroids—homobrassinolide and epibrassinolide—caused an increase in protein content and a change in the proportion of some amino acids. Both hormones increased heterogeneity of high-and medium-molecular-weight nonhistone chromatin proteins but had no effect on the polypeptide profile of histones. Using epibrassinolide as an example, we showed that the brassinosteroid-induced increase in protein content in lupine seeds was due primarily to accumulation of the low-molecular-weight components of β-conglutin. The content of some amino acids in both α-and β-conglutins changed. These changes in protein metabolism correlated with an increase in the content of indoleacetic acid and a decrease in the content of abscisic acid.     

Regulation by sucrose of storage compounds breakdown in germinating seeds of yellow lupine (Lupinus luteus L.), white lupine (Lupinus albus L.) and Andean lupine (Lupinus mutabilis Sweet). II. Mobilization of storage lipid

Research of the regulatory function of sucrose in storage lipid breakdown was conducted on isolated embryo axes, excised cotyledons and whole seedlings of three lupine species grown in vitro on medium with 60 mM sucrose or without the sugar. Lack of sucrose in the medium caused significant increase in total lipid content in yellow, white and Andean lupine isolated embryo axes but in Andean lupine seedling cotyledons and excised cotyledons, lipid level was clearly lower in carbohydrates deficient conditions. Sucrose caused no significant effect on fatty acids spectra. The main fatty acid in yellow lupine seeds was linoleic acid, in white lupine oleic acid and in Andean lupine both oleic and linoleic acids. The main phospholipid in organs of three lupine species was phosphatidylcholine. In sugar-deficient conditions, content of phosphatidylcholine and some others phospholipids was decreased. The peculiar features of regulation by sugars of storage lipid breakdown in germinating lupine seeds and induction of autophagy in young carbohydrate starved embryo axes is discussed.     

The pivotal role of glutamate dehydrogenase (GDH) in the mobilization of N and C from storage material to asparagine in germinating seeds of yellow lupine

In germinating seeds of legumes, amino acids liberated during mobilization of storage proteins are partially used for synthesis of storage proteins of the developing axis, but some of them are respired. The amino acids are catabolized by both glutamate dehydrogenase (GDH) and transaminases. Ammonium is reassimilated by glutamine synthetase (GS) and, through the action of asparagine synthetase (AS), is stored in asparagine (Asn).This review presents the ways in which amino acids are converted into Asn and their regulation, mostly in germinating seeds of yellow lupine, where Asn can make up to 30% of dry matter. The energy balance of the synthesis of Asn from glutamate, the most common amino acid in lupine storage proteins, also shows an adaptation of lupine for oxidation of amino acids in early stages of germination.Regulation of the pathway of Asn synthesis is described with regard to the role of GDH and AS, as well as compartmentation of particular metabolites. The regulatory effect of sugar on major links of the pathway (mobilization of storage proteins, induction of genes and activity of GDH and AS) is discussed with respect to recent genetic and molecular studies. Moreover, the effect of glutamate and phytohormones is presented at various stages of Asn biosynthesis.     

Chemical composition of different sorts of soybeans and changes under the influence of weather and soil-climatic conditions]

The biochemical composition (the content of protein and lipids, the content of essential amino acids in the protein complex, the fatty acid composition of lipids) of seeds of eighteen soyabean varieties cultivated in different soil-climatic zones under various weather conditions was examined. Changes in the biochemical parameters were primarily associated with the genotype of the variety. The effect of weather, soil and climatic conditions was less marked. With an increase of the protein content the concentration of lysine and methionine in proteins tended to decrease. As long as the oil content in seeds grew, the concentration of oleic acid increased and that of linoleic and linolenic acids decreased.     

Amino Acid composition of germinating cotton seeds

Total and free amino acid composition of germinating cotton seeds (Gossypium hirsutum L.) was determined. The germinating seeds were separated into cotyledon and developing axis fractions daily and the composition of each tissue was summed to get the whole seed composition. By separating the developing seeds into these two tissue fractions, and determining total and free amino acids, a balance sheet was developed for each amino acid. This technique allowed changes in distribution with time of each amino acid to be followed in each tissue. Data for total content and amount in protein of each amino acid are presented. Asparagine increased in the whole seed, and most of this increase was found in the free pool of the developing axis. Other amino acids (e.g. arginine, glutamic acid) increased in the free pool but showed an over-all decrease, indicating that they were being metabolized. Amino acid contents of storage and nonstorage protein isolates were determined.     

Water uptake and distribution in germinating lupine seeds studied by magnetic resonance imaging and NMR spectroscopy

Magnetic resonance imaging (MRI) was used to study temporal and spatial water uptake and distribution in germinating lupine ( Lupinus luteus L.) seeds. During 24 h of imbibition, water was unevenly distributed within the seed and some anatomical parts were more hydrated than others. Water entered the seed through the hilum and micropyle. The embryonic axis was the first to show hydration followed by seed coat and later cotyledons. The changes in water status were characterized by NMR spectroscopy. Analyses of T₂ relaxation times revealed a three-component water proton system (structural, intracellular and extracellular water) in germinating lupine seeds. The data on the components of transverse relaxation time studies indicated the complex exchange processes taking place between water components inside lupine seed over first 2.5 h of hydration, with a distinguished increase in structural water and decrease in other components. This speaks in favor of the high water-absorbing capacity of lupine seeds as related to high protein content. Germination was accompanied by swelling of protein bodies and changes in the organization of stored reserves with gradual disappearance of protein from the cells.     

Effect of laser light treatment on some biochemical and physiological processes in seeds and seedlings of white lupine and faba bean

The aim of present study was to determine the changes of some biochemical and physiological processes, which occurred in seeds and seedlings of white lupine and faba bean after pre-sowing treatment with laser beams. It was found this treatment of seeds considerably increased the activity of amylolytic enzymes in seeds of both plants. The greatest differentiation of the enzymatic activity was noticed after 120?h from the time of sowing but the activity of these enzymes in the seeds of both tested plants was similar and it had the same course in time. The irradiated seeds of white lupine and faba bean had higher fresh weight at the time of imbibition than the seeds which were not treated with laser beams. It resulted in earlier and more uniform germination. The concentration of free radicals increased considerably in the seeds pre-treated with laser beams and the largest increase in seeds of both plant species was noticed after five exposures to laser beams. Treating seeds with laser beams considerably increased the amount of indole-3-acetic acid (IAA) in the germinating seeds. Three exposures of seeds caused the largest increase of this plant hormone content in the seeds. The activity of IAA in faba bean was slightly higher than in white lupine seeds. Pre-sowing stimulation with laser had a positive influence on the growth and development of seedlings, which had longer hypocotyl and roots in comparison to seedlings which grew from non irradiated seeds.     

Amino acids in seeds and seedlings of the genus Lens

The amino acid content of seeds and 4-day-old seedlings were studied in five species of lentil: Lens culinaris, L. orientalis, L. ervoides, L. nigricans and L. odemensis. Free amino acid and also total protein amino acid content after HCl hydrolysis were determined by HPLC. The nonprotein UV-absorbing amino acids were determined by capillary zone electrophoresis (CZE). The content of free protein amino acids in seeds varied among species and increased dramatically after germination. Asparagine is quantitatively most important in both seed and seedling. The content of free nonprotein amino acids is variable in seeds and seedlings. gamma-Hydroxyarginine, gamma-hydroxyornithine, alpha-aminobutyric acid and taurine were found in both seeds and seedlings. Homoarginine was found in four species but not in L. orientalis while gamma-aminobutyric acid (GABA), alpha-aminoadipic acid (alpha-aaa) and three isoxazolinone derivatives: beta-(isoxazolin-5-on-2-yl)-alanine (BIA), gamma-glutamyl-BIA (gamma-glu-BIA) and 2-carboxymethyl-isoxazolin-5-one (CMI) were found exclusively in the seedlings. CMI was identified for the first time in lentil species. Lathyrine, beta-(2-amino-pyrimidine-4-yl)-alanine, which was reported to be in the seeds of some Lathyrus species was confirmed to be present also in the seedling of L. culinaris (trace amount), L. nigricans and L. odemensis. Trigonelline (N-methyl-nicotinic acid), a plant hormone, is present both in seeds and seedlings in different concentrations except in L. ervoides. The different combination of nonprotein amino acids among the species gives indication of their genetic relationship and might partly explain the varying compatibility for interspecies crossing.     

A transfer of carbon atoms from fatty acids to sugars and amino acids in yellow lupine (Lupinus luteus L.) seedlings

The metabolism of 14C-acetate was investigated during the in vitro germination of yellow lupine seeds. Carbon atoms (14C) from the C-2 position of acetate were incorporated mainly into amino acids: aspartate, glutamate, and glutamine and into sugars: glucose, sucrose, and fructose. In contrast to this, 14C from the C-1 position of acetate was released mainly as 14CO2. Incorporation of 1-14C and 2-14C from acetate into amino acids and sugars in seedling axes was more intense when sucrose was added to the medium. However, in cotyledons where lipids are converted to carbohydrates, this process was inhibited by exogenous sucrose. Since acetate is the product of fatty acid beta-oxidation, our results indicate that, at least in lupine, seed storage lipids can be converted not only to sucrose, but mainly to amino acids. Inhibitory effects of sucrose on the incorporation of 14C from acetate into amino acids and sugars in cotyledons of lupine seedlings may be explained as the effect of regulation of the glyoxylate cycle by sugars.     

Protein and free amino acids in field-grown cowpea seeds as affected by water stress at various growth stages

Summary This study was undertaken to evaluate water stress effects during vegetative, flowering, and podfilling stages of cowpea plants (Vigna unguiculata L.) grown under natural field conditions in southern California on seed yield and protein and free amino acid content of the cowpea seeds. The lowest concentration of N was found in the seeds of the control treatment plants while the seed yield from these treatments was the highest as compared with the N concentration and yield of seeds from plants subjected to water stress during flowering and podfilling stages. The concentration of N in the seeds was inversely related to the seed dry weight yield. Protein arginine,-threonine,-serine,-cystine,-valine,-methionine, and-isoleucine were significantly affected by water stress at the three growth stages. There was no consistent pattern in the effect of water stress on the individual amino acids. The sum of protein amino acids in the cowpea seeds was not significantly influenced by the various treatments since some of the protein amino acids increased and others decreased producing an averaging effect on the figures comprising the sums of the amino acids. Water stress during the flowering and pod-filling stages increased the free amino acid pool, and at the same time, inhibited incorporation of the amino acids into the protein chain-thus lowering the protein amino acid fraction simultaneously. With the exception of methionine plus cystine, the essential amino acids in the seeds were present at concentrations equal to or greater than recommended by the World Health Organization and FAO. It is of particular importance to note that the concentration of lysine in the cowpeas was substantially higher than that found in wheat grain. It is also important to note that the amount of essential amino acids per gram of protein was not measurably affected by the water stress treatments during any of the growth stages.     

Chemical differences between seeds and elaiosomes indicate an adaptation to nutritional needs of ants

Ant-dispersed plants usually produce seeds with appendages (elaiosomes) as reward for ants. Plants that produce high-quality elaiosomes benefit because ants preferentially disperse their diaspores. We therefore hypothesized that seeds and elaiosomes differ in chemical composition in ways that make elaiosomes of high nutritional quality for ants, capable of providing essential dietary components that explain the increased fitness and higher gyne production documented for colonies with elaiosome consumption. To test the hypothesis we analysed the content and composition of lipids, amino acids, soluble carbohydrates, proteins and starch in seeds and elaiosomes of 15 central European ant-dispersed plants. After separating the different fractions, total lipids were determined gravimetrically, fatty acids and soluble carbohydrates were detected by gas chromatography (GC) and GC–mass spectrometry, free amino acids by an amino acid analyser while starch and protein were analysed photometrically. Seeds accumulated high molecular weight compounds such as proteins and starch, whereas elaiosomes accumulated more easily digestible low molecular weight compounds such as amino acids and monosaccharides. Analysis of similarities and similarity percentages analysis demonstrated that the composition of fatty acids, free amino acids and carbohydrates differed markedly between elaiosomes and seeds. The most important difference was in total amino acid content, which was on average 7.5 times higher in elaiosomes than in seeds. The difference was especially marked for the nitrogen-rich amino acid histidine. The availability of essential nutrients and, in some species, the higher nitrogen content in elaiosomes suggest that their nutritional value for larvae plays a key role in this interaction. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Activities of protein-inhibitors of proteases in seeds of different varieties of yellow and white lupine plants

Activities of protein-inhibitors of proteases and their correlation with the total content of protein and alkaloids in yellow and white lupine seeds were studied. Yellow lupine seeds showed lowe activities of protein-inhibitors. They also displayed negative correlation between activities of protein-inhibitors and the total content of protein and alkaloids.     

Limits to sulfur accumulation in transgenic lupin seeds expressing a foreign sulfur-rich protein

The low sulfur amino acid content of legume seeds restricts their nutritive value for animals. We have investigated the limitations to the accumulation of sulfur amino acids in the storage proteins of narrow leaf lupin (Lupinus angustifolius) seeds. Variation in sulfur supply to lupin plants affected the sulfur amino acid accumulation in the mature seed. However, when sulfur was in abundant supply, it accumulated to a large extent in oxidized form, rather than reduced form, in the seeds. At all but severely limiting sulfur supply, addition of a transgenic (Tg) sink for organic sulfur resulted in an increase in seed sulfur amino acid content. We hypothesize that demand, or sink strength for organic sulfur, which is itself responsive to environmental sulfur supply, was the first limit to the methionine (Met) and cysteine (Cys) content of wild-type lupin seed protein under most growing conditions. In Tg, soil-grown seeds expressing a foreign Met- and Cys-rich protein, decreased pools of free Met, free Cys, and glutathione indicated that the rate of synthesis of sulfur amino acids in the cotyledon had become limiting. Homeostatic mechanisms similar to those mediating the responses of plants to environmental sulfur stress resulted in an adjustment of endogenous protein composition in Tg seeds, even when grown at adequate sulfur supply. Uptake of sulfur by lupin cotyledons, as indicated by total seed sulfur at maturity, responded positively to increased sulfur supply, but not to increased demand in the Tg seeds.     

Comparative study of storage compound breakdown in germinating seeds of three lupine species

Storage lipid and protein breakdown in germinating seeds of yellow (Lupinus luteus L.), white (L. albus L.), and Andean lupine (L. mutabilis Sweet) and regulatory function of sucrose were investigated. Less oil bodies were detected in organs of yellow lupine seeds, whereas the highest content of oil bodies was noticed in the Andean lupine seeds. Mature, air-dried yellow, white and Andean lupine seeds do not contain starch. Starch grains appear the earliest in white lupine seeds during imbibition. Sucrose deficiency in tissues enhances breakdown of storage lipid, protein and temporary starch in cotyledons. In sucrose starved embryo axes of all investigated lupine species, an increased level of vacuolization was noted. Interconnections between catabolism of storage protein and storage lipid in germinating lupine seeds were identified by applying ¹⁴C-acetate. To assess the importance of key processes in storage lipid breakdown NaF (inhibitor of glycolysis and gluconeogenesis), KCN, NaN₃ and SHAM (inhibitors of mitochondrial electron transport chain) and MSO (inhibitor of glutamine synthetase) were used. Radioactivity coming from ¹⁴C-acetate was released as ¹⁴CO₂ but mostly was incorporated into ethanol-soluble fraction of embryo axes and cotyledons. Respiratory inhibitors caused a significant decrease in ¹⁴CO₂ and ethanol fractions in all three lupine species studied. MSO stimulated release of ¹⁴CO₂ and radioactivity of ethanol fractions in yellow lupine organs fed with sucrose, but in Andean lupine MSO enhanced the production of ¹⁴CO₂ and radioactivity of ethanol fractions both in organs fed and not fed with sucrose. Different strategies of storage compound breakdown are proposed, depending on relative proportion in storage protein and lipid content in lupine seeds.     

AAP1 regulates import of amino acids into developing Arabidopsis embryos

The embryo of Arabidopsis seeds is symplasmically isolated from the surrounding seed coat and endosperm, and uptake of nutrients from the seed apoplast is required for embryo growth and storage reserve accumulation. With the aim of understanding the importance of nitrogen (N) uptake into developing embryos, we analysed two mutants of AAP1 (At1g58360), an amino acid transporter that was localized to Arabidopsis embryos. In mature and desiccated aap1 seeds the total N and carbon content was reduced while the total free amino acid levels were strongly increased. Separately analysed embryos and seed coats/endosperm of mature seeds showed that the elevated amounts in amino acids were caused by an accumulation in the seed coat/endosperm, demonstrating that a decrease in uptake of amino acids by the aap1 embryo affects the N pool in the seed coat/endosperm. Also, the number of protein bodies was increased in the aap1 endosperm, suggesting that the accumulation of free amino acids triggered protein synthesis. Analysis of seed storage compounds revealed that the total fatty acid content was unchanged in aap1 seeds, but storage protein levels were decreased. Expression analysis of genes of seed N transport, metabolism and storage was in agreement with the biochemical data. In addition, seed weight, as well as total silique and seed number, was reduced in the mutants. Together, these results demonstrate that seed protein synthesis and seed weight is dependent on N availability and that AAP1-mediated uptake of amino acids by the embryo is important for storage protein synthesis and seed yield.     

不同产地长柄扁桃种仁成分分析

采用GC-MS、AAS、HPLC等方法对选自陕西神木、内蒙古包头、内蒙古阿拉善盟、河北承德4个不同产地的长柄扁桃种仁的一般成分(粗脂肪、粗蛋白、水分、灰分、总糖、总膳食纤维)、氨基酸、微量元素、脂肪酸及苦杏仁苷含量等进行分析。研究表明,不同产地长柄扁桃种仁各组分含量有一定差别,但均富含粗脂肪(416~478g/kg)和粗蛋白(206~286g/kg);种仁中氨基酸种类齐全,含量丰富;均含Ca、Mg、P、Fe、K等9种对人体有益的矿质元素,未检出Pb、Cd、Hg、As等元素;长柄扁桃油中不饱和脂肪酸含量在96.8%以上,以油酸和亚油酸主;苦杏仁苷含量在3%左右。从测定结果来看,长柄扁桃种仁可以作为开发食用油、蛋白粉以及苦杏仁苷的原材料,具有较好的经济价值,适宜于广大北方荒漠地区推广种植。     

Manipulation of amino acid composition in soybean seeds by the combination of deregulated tryptophan biosynthesis and storage protein deficiency

The ability of genetic manipulation to yield greatly increased concentrations of free amino acids (FAAs) in seeds of soybean was evaluated by introduction of a feedback-insensitive mutant enzyme of tryptophan (Trp) biosynthesis into two transformation-competent breeding lines deficient in major seed storage proteins. The storage protein-deficient lines exhibited increased accumulation of certain other seed proteins as well as of FAAs including arginine (Arg) and asparagine in mature seeds. Introduction of the gene for a feedback-insensitive mutant of an α subunit of rice anthranilate synthase (OASA1D) into the two high-FAA breeding lines by particle bombardment resulted in a >10-fold increase in the level of free Trp in mature seeds compared with that in nontransgenic seeds. The amount of free Trp in these transgenic seeds was similar to that in OASA1D transgenic seeds of the wild-type cultivar Jack. The composition of total amino acids in seeds of the high-FAA breeding lines remained largely unaffected by the expression of OASA1D with the exception of an increase in the total Trp content. Our results therefore indicate that the extra nitrogen resource originating from storage protein deficiency was used exclusively for the synthesis of inherent alternative nitrogen reservoirs such as free Arg and not for deregulated Trp biosynthesis conferred by OASA1D. The intrinsic null mutations responsible for storage protein deficiency and the OASA1D transgene affecting Trp content were thus successfully combined and showed additive effects on the amino acid composition of soybean seeds.     

Changes in accumulation of seed nitrogen compounds in maize under conditions of sulphur deficiency

Maize ( Zea mays L., hybrid INRA 260) was grown in the greenhouse with mineral nutrition of different sulphate concentrations. Mature seeds from these plants were compared for their free amino acid and protein N forms. For the most S-deficient sample, the Asx (asparagine + aspartic acid) content increased by 30% as compared with control, while methionine and cysteine decreased (by 25 and 30%, respectively), as well as glycine, lysine, histidine, arginine and tryptophan. In seeds lowest in S the non-protein N to total N ratio was 77% higher than in the control. Free asparagine dominated in starved seeds (50 mol % of total free amino acids) and was ten-fold more concentrated than in the control, where proline was the predominant free amino acid. Thus the Asx of non-protein N reached 28% of the total mol Asx of the whole starved seed. Altered S nutrition had virtually no effect on the amino acid composition of the main protein fractions, but it significantly changed their ratios. Zeins, which are poor in S-containing amino acids, showed 25% higher level than in seeds supplied with normal S. As a counterbalance, two glutelin subfractions rich in S-containing amino acids, decreased by 36–71% under limiting S nutrition.
It is concluded that the plant reacts against S deficiency by modifying its N metabolism. Significant accumulation occurred of free asparagine, which is the main form of N transportation. The biosynthesis of seed storage protein occurred through the accumulation of the highest possible protein quantity allowed by the available S-containing amino acids, i.e. proteins low in S-containing amino acids were preferentially synthesized.     

Deoxyribonucleic Acid, Ribonucleic Acid, Protein and Uncombined Amino Acid Content of Legume Seeds During Embryogeny

The nucleic acid, protein and uncombined amino acid contentof seeds of soya-bean (Glycine max L. Merr.), garden pea (Pisumsativum L.), kidney bean (Phaseolus vulgaris L.) and peanut(Arachis hypogaea L.) were measured at various times duringseed formation in an effort to understand why the soya-beanhas nearly twice as much protein as the other legume seeds.In all these species the concentration of deoxyribonucleic acid,ribonucleic acid and uncombined amino acids decreased duringseed formation. The protein level of kidney bean was relativelyconstant during development whereas the protein levels of pea,peanut and soya-bean increased during development. The proteincontent of the soya-bean increased throughout development whereasthe protein increase in peanut took place early and that inpea took place later in development. The ratio of protein toribonucleic acid was highest in peanut, less in soya-bean, andlowest in pea and kidney bean. Similarly, the ratio of proteinto deoxyribonucleic acid was higher in kidney bean than in soya-bean.Soya-beans had a lower amino acid content than any of the otherseeds at all stages of development. These results indicate thatneither total deoxyribonucleic acid, ribonucleic acid nor uncombinedamino acid content is responsible for the higher protein contentof soya-beans.     

The nutritional value of narrow-leafed lupine (Lupinus angustifolius) for fattening pigs

The aim of this study was to determine the nutrient digestibility of seeds of four varieties of narrow-leafed lupines (Lupinus angustifolius) and the possibility of soya bean meal (SBM) substitution by lupine seeds alone and in combination with rapeseed meal (RSM) in the diets of pigs. The seeds of the lupine varieties Kalif, Sonet, Zeus and Boruta were analysed. The apparent total tract digestibility (ATTD) was determined on 50 cross-bred pigs using the difference method with titanium dioxide as a marker. The substitution of SBM by lupine seeds alone (at 0 – 100%) was tested on 60 pigs (20–105 kg body weight (BW)) and by a combination of lupine seeds and RSM on 180 fattening pigs (35–80 kg BW). The chemical composition of lupine seeds differed considerably, especially in terms of crude protein and mineral content. All seeds contained less than 0.05% alkaloids and 9.3% oligosaccharides in dry matter. The ATTD of protein ranged from 70% to 74%, those of ether extract from 36% to 55% and those of gross energy from 77% to 84%. The entire replacement of SBM by lupine seeds (var. Sonet) did not have a negative effect on the performance of grower and fattener pigs. The substitution of SBM by a combination of lupines and RSM reduced the performance of growing and finishing pigs significantly.