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Possible mechanism of inhibition of 6-methoxy-benzoxazolin-2(3H)-one on germination of cress (Lepidium sativum L.).

6-Methoxy-benzoxazolin-2(3H)-one (MBOA) inhibited the germination of cress (Lepidium sativum L.) seeds at concentrations greater than 0.03 mM. Inhibition was overcome by sucrose, suggesting that MBOA may inhibit sugar metabolism in cress seeds. Induction of alpha-amylase activity in seeds was also inhibited by MBOA at concentrations greater than 0.03 mM. Inhibition of both germination and induction of alpha-amylase activity increased with increasing concentrations of MBOA, and the extent of germination correlated positively with the activity of alpha-amylase in the seeds. MBOA added to a reaction mixture for alpha-amylase assay did not affect enzyme activity, indicating that MBOA does not inhibit in vitro alpha-amylase activity. Cress seeds germinated approximately 16 hr after incubation, and inhibition of alpha-amylase by MBOA occurred within 6 hr after incubation. These results suggest that MBOA may inhibit the germination of cress seeds by inhibiting the induction of alpha-amylase activity, because alpha-amylase plays a key role in the conversion of reserve carbohydrate into soluble sugars, a prerequisite for seed germination.

Benzoxazoles↗

Isoflavone contents in germinated soybean seeds.

The effect of germination on isoflavone contents in two soybean varieties (Hutcheson and Caviness) was investigated. Soybean seeds were soaked at 25 degrees C for 12 h, germinated at 40 degrees C, and freeze-dried. The isoflavone contents of dry, soaked, germinated (hypocotyl length at 0.5, 2.5, and 6.5 mm), and nongerminated seeds were determined by high performance liquid chromatography. The maximum amount of total isoflavone, genistein, and daidzein with their beta-glucoside conjugates was obtained when hypocotyl length of the germinated-seed from var. Hutcheson was 0.5 mm (2.491, 1.500, and 0.671 mg/g), and from var. Caviness was 2.5 mm (2.78, 1.523, and 0.905 mg/g). A dramatic increase in malonylgenistin and malonyldaidzin (1.305 mg/g and 0.476 mg/g in Hutcheson, and 1.308 mg/g and 0.677 mg/g in Caviness, respectively) was observed at these hypocotyl lengths. A decrease was observed after this stage. Genistein and daidzein contents were highest just after soaking. Glycitein and its beta-glucoside conjugates remained almost the same during germination. Controlled germination can be used to enhance isoflavone content in soybean seed.

Chromatography, High Pressure Liquid↗

Bioremediation of HCH-contaminated soil: elimination of inhibitory effects of the insecticide on radish and green gram seed germination.

The effects of technical grade hexachlorocyclohexane (tech-HCH) on the germination of different seeds were tested. Two types of seeds, radish and green gram showed marked reduction in germination percentage and seeding vigour index. The abnormalities and reduction in germination increased with increasing concentration of tech-HCH. At 100 microg HCH level the germination of radish and green gram seeds was inhibited almost completely on moist filter paper and soil. Protease and amylase activities were reduced in seeds grown in soil spiked with tech-HCH. Bioremediation of HCH-spiked soils with a HCH-degrading microbial consortium helped in eliminating the toxic effects of tech-HCH towards seed germination. The degradation of 25 microg tech-HCH g(-1) soil was complete by 120 h. The seed germination and the activities of the assayed enzymes, amylase and protease, were same as before or better in bioremediated soils.

Biodegradation, Environmental↗

Excess copper induced physiological and proteomic changes in germinating rice seeds.

Copper is an essential micronutrient for plants. Present at a high concentration in soil, copper is also regarded as a major toxicant to plant cells due to its potential inhibitory effects against many physiological and biochemical processes. The interference of germination-related proteins by heavy metals has not been well documented at the proteomic level. In the current study, physiological, biochemical and proteomic changes of germinating rice seeds were investigated under copper stress. Germination rate, shoot elongation, plant biomass, and water content were decreased, whereas accumulation of copper and TBARS content in seeds were increased significantly with increasing copper concentrations from 0.2mM to 1.5mM followed by germination. The SDS-PAGE showed the preliminary changes in the polypeptides patterns under copper stress. Protein profiles analyzed by two-dimensional electrophoresis (2-DE) revealed that 25 protein spots were differentially expressed in copper-treated samples. Among them, 18 protein spots were up-regulated and 7 protein spots were down-regulated. These differentially displayed proteins were identified by MALDI-TOF mass spectrometry. The up-regulation of some antioxidant and stress-related proteins such as glyoxalase I, peroxiredoxin, aldose reductase, and some regulatory proteins such as DnaK-type molecular chaperone, UlpI protease, and receptor-like kinase clearly indicated that excess copper generates oxidative stress that might be disruptive to other important metabolic processes. Moreover, down-regulation of key metabolic enzymes like alpha-amylase or enolase revealed that the inhibition of seed germinations after exposure to excess copper not only affects starvation in water uptake by seeds but also results in failure in the reserve mobilization processes. These results indicate a good correlation between the physiological and biochemical changes in germinating rice seeds exposed to excess copper.

Biomass↗

From dormancy to germination: Transcriptome analysis reveals the potential involvement of heat shock proteins in gibberellin-induced seed dormancy release in Solanum torvum.

Solanum torvum, a superior vegetable grafting rootstock and medicinal Solanaceae plant, exhibits strong seed dormancy, which limits its commercial cultivation. Among various strategies explored to improve the germination rate of S. torvum, exogenous application of gibberellin (GA) has been shown to be effective. In this study, a GA concentration of 2.5 mM was established as the optimal for breaking dormancy in S. torvum seeds. Transcriptome analysis of dry, water-soaked, GA-soaked, and GA-induced germinated seeds was conducted to investigate the molecular mechanism of GA-mediated dormancy release. During the soaking period, GA application significantly induced transcriptome changes in processes including protein processing, translation, and peptide biosynthesis. Concurrently, GA treatment promoted plant hormone signal transduction, enhanced DNA-binding transcription factor activity, and activated monocarboxylic acid biosynthetic process, all of which facilitated seed water absorption. Furthermore, the differentially expressed genes (DEGs) induced by GA during soaking primarily functioned in signal transduction or activation. While most of these DEGs returned to their pre-treatment expression levels before subsequent recovery, a subset persisted until seed germination. During radicle protrusion, the persistent DEGs were associated with energy metabolism and cell structure establishment. Notably, heat shock protein (HSP) genes showed dynamic expression across all stages (soaking, germination, and radicle penetration). Furthermore, by adjusting germination conditions, temperature was confirmed to be a necessary but not sufficient condition for GA-induced S. torvum seed germination. However, functional validation (e.g., using HSP inhibitors or genetic approaches) is still required to confirm the causal role of HSPs. Collectively, these findings not only clarify the molecular basis of GA-regulated seed dormancy breaking in S. torvum but also provide practical guidance for optimizing its commercial propagation protocols.

Gibberellins↗

Validation of germination rate and root elongation as indicator to assess phytotoxicity with Cucumis sativus.

Germination rate and root elongation, as a rapid phytotoxicity test method, possess several advantages, such as sensitivity, simplicity, low cost and suitability for unstable chemicals or samples. These advantages made them suitable for developing a large-scale phytotoxicity database and especially applicable for developing quantitative structure-activity relationship (QSAR) to study mechanisms of phytotoxicity. In this paper, the comparative inhibition of germination rate and root elongation of Cucumis sativus by selected halogen-substituted phenols and anilines were determined. The suitability of germination rate and root elongation as phytotoxicity endpoints was evaluated. Excellent reproducibility and stability of germination rate and root elongation in the control test, relatively greater sensitivity and similar dose-response relations for all tested compounds were observed. These results together with those of a 2-day test were used to demonstrate the suitability of this phytotoxicity test method. A QSAR was developed for the phytotoxicity mode of action of the tested compounds to C sativus seeds. Models that combined the logarithm of 1-octanol/water partition coefficient (log Kow) and the energy of the lowest unoccupied molecular orbital (Elumo) were developed for both germination rate inhibition and root elongation inhibition. The results of these studies indicate that phytotoxicity of substituted phenols and anilines to C. sativus seeds could be explained by a polar narcosis mechanism. This paper will promote the application of germination rate and root elongation method and the development of large-scale phytotoxicity database, which will provide the fundamental data for QSAR and ecological risk assessment of organic pollutants.

Biomarkers↗

Influence of diesel fuel on seed germination.

The use of plant-based systems to remediate contaminated soils has become an area of intense scientific study in recent years and it is apparent that plants which grow well in contaminated soils need to be identified and screened for use in phytoremediation technologies. This study investigated the effect of diesel fuel on germination of selected plant species. Germination response varied greatly with plant species and was species specific, as members of the same plant family showed differential sensitivity to diesel fuel contamination. Differences were also seen within plant subspecies. At relatively low levels of diesel fuel contamination, delayed seed emergence and reduced percentage germination was observed for the majority of plant species investigated. Results suggest the volatile fraction of diesel fuel played an influential role in delaying seed emergence and reducing percentage germination. In addition, the remaining diesel fuel in the soil added to this inhibitory effect on germination by physically impeding water and oxygen transfer between the seed and the surrounding soil environment, thus hindering the germination response.

Biodegradation, Environmental↗

Identification of quantitative trait loci for ABA sensitivity at seed germination and seedling stages in rice.

Abscisic acid (ABA) is one of the important plant hormones, which plays a critical role in seed development and adaptation to abiotic stresses. The sensitivity of rice (Oryza sativa L.) to exogenous ABA at seed germination and seedling stages was investigated in the recombinant inbred line (RIL) population derived from a cross between irrigated rice Zhenshan 97 and upland rice IRAT109, using relative germination vigor (RGV), relative germination rate (RGR) and leaf rolling scores of spraying (LRS) or culturing (LRC) with ABA as sensitivity indexes. The phenotypic correlation analysis revealed that only RGV at germination stage was positively correlated to ABA sensitivity at seedling stage. QTL detection using composite interval mapping (CIM) and mixed linear model was conducted to dissect the genetic basis of ABA sensitivity, and the single-locus QTLs detected by both methods are in good agreement with each other. Five single QTLs and six pairs of epistatic QTLs were detected for ABA sensitivity at germination stage. Eight single QTLs and five pairs of epistatic QTLs were detected for ABA sensitivity at seedling stage. Two QTLs were common between LRS and LRC; and one common QTL was detected for RGV, LRS and LRC simultaneously. These results indicated that both single and epistatic loci were involved in the ABA sensitivity in rice, and the genetic basis of ABA sensitivity at seed germination and seedling stage was largely different.

Abscisic Acid↗

The role of GA-mediated signalling in the control of seed germination.

Seed germination is promoted by gibberellin (GA) in many plant species. Several GA signalling factors are known to induce the expression of genes encoding enzymes that mobilise food reserves, including starches, proteins and lipids, stored in the endosperm during seed germination. However, these factors do not control seed germination. Two recent reports have indicated that RGL1 and RGL2, both homologous to the GA-response height-regulating factors GAI/RGA/RHT/d8/SLR1/SLN1, are repressors of seed germination in Arabidopsis. These reports provide new clues as to how GA controls seed germination. The induction of RGL2 expression by imbibition and its repression by GA are of particular interest because they imply that RGL2 acts as an integrator of environmental and endogenous cues for germination.

Arabidopsis↗

Effect of several germination conditions on total P, phytate P, phytase, and acid phosphatase activities and inositol phosphate esters in rye and barley.

Two assays were conducted to study the evolution of rye and barley phosphatases (phytase and acid phosphatase) and the degradation of its substrates (inositol phosphate esters) during seed germination. In this manner we could obtain a low-phytate, endogenous phosphatase rich ingredient to be used in animal nutrition. In the first assay, the seeds were soaked for 1 and 14 h and germinated for 3 and 5 days with and without the addition of gibberellic acid (GA3). In the second assay, the seeds were soaked for 1 h and germinated for 1, 3, and 5 days with GA3. Phytase (up to 5739 and 3151 U x kg(-1)) and acid phosphatase (up to 18288 and 3151 U x g(-1)) activities, and IP6 (6.09 and 6.01 mg x g(-1)), IP5 (0.48 and 0.48 mg x g(-1)), and IP4 (0.13 and 0.06 mg x g(-1)) were detected in ungerminated rye and barley, respectively. The germination process caused a significant increase of Phy and AcPh activities in rye (up to 112 and 213%) and barley (up to 212 and 634%) and a reduction in the phytate phosphorus content (up to 84 and 58%, respectively). Phytate phosphorus content was affected only by soaking time in the case of rye. Finally, during the course of germination, IP6 and IP5 were rapidly degraded in rye (88 and 79%) and barley (67 and 52%), and IP4 was only a short-living intermediate, which was increased during hydrolysis and degraded to IP3. In conclusion, a marked increase of Phy and AcPh activities in rye and barley with a concomitant decrease in phytate phosphorus content and an increase in the content of lower inositol phosphates were observed during the rye and barley germination.

6-Phytase↗

Essential oils from Mediterranean lamiaceae as weed germination inhibitors.

The essential oils obtained from rosemary (Rosmarinus officinalis L.), thyme (Thymus vulgaris L.), and savory (Satureja montana L.) and the four monoterpenes that are their major constituents have been analyzed by GC and GC-MS and tested for their allelopathic properties on the seeds of three different annual weeds (Chenopodium album, Portulaca oleracea, and Echinochloa crus-galli) and three crops (Raphanus sativus, Capsicum annuum, and Lactuca sativa), with the aim to evaluate in vitro their potential as germination inhibitors. The essential oil composition varied with the species, thymol being the main constituent (44%) of thyme and carvacrol (57%) that of savory oil. Differences in essential oil composition were observed within two different rosemary ecotypes, type A, with alpha-pinene (37%) and 1,8-cineole (23%), and type B, characterized by a 2-fold content of 1,8-cineole (47%). This latest essential oil inhibited completely the germination of weeds while concurrently displaying little effect on pepper. The other two oils showed less selective action. S. montana essential oil, with 57% carvacrol, is the most active compound, completely inhibiting germination both of crops and weeds. Borneol, one of the main constituents of the oil of rosemary type B, showed an activity comparable to that of the whole oil. Crop and weed seeds treated with 1,8-cineole showed germination values that were not significantly different from controls, even if a slowing of the germination process expressed in terms of a significant increase in mean germination time was observed. Monoterpene compounds also present in the essential oils mainly represented the volatile fraction released from the crops and their residues into the soil.

Chromatography, Gas↗

Analysis of phenolic compounds in white rice, brown rice, and germinated brown rice.

Two hydroxycinnamate sucrose esters, 6'-O-(E)-feruloylsucrose and 6'-O-(E)-sinapoylsucrose, were isolated from methanol extracts of rice bran. Soluble and insoluble phenolic compounds as well as 6'-O-(E)-feruloylsucrose and 6'-O-(E)-sinapoylsucrose from white rice, brown rice, and germinated brown rice were analyzed using HPLC. The results demonstrated that the content of insoluble phenolic compounds was significantly higher than that of soluble phenolics in rice, whereas almost all compounds identified in germinated brown rice and brown rice were more abundant than those in white rice. 6'-O-(E)-Feruloylsucrose (1.09 mg/100 g of flour) and 6'-O-(E)-sinapoylsucrose (0.41 mg/100 g of flour) were found to be the major soluble phenolic compounds in brown rice. During germination, an approximately 70% decrease was observed in the content of the two hydroxycinnamate sucrose esters, whereas free phenolic acid content increased significantly; the ferulic acid content of brown rice (0.32 mg/100 g of flour) increased to 0.48 mg/100 g of flour and became the most abundant phenolic compound in germinated brown rice. The content of sinapinic acid increased to 0.21 mg/100 g of flour, which is nearly 10 times as much as that in brown rice (0.02 mg/100 g of flour). In addition, the total content of insoluble phenolic compounds increased from 18.47 mg/100 g of flour in brown rice to 24.78 mg/100 g of flour in germinated brown rice. These data suggest that appropriate germination of brown rice may be a method to improve health-related benefits.

Chromatography, High Pressure Liquid↗

Effect of germination and thermal treatments on folates in rye.

Effects of germination conditions and thermal processes on folate contents of rye were investigated. Total folate contents were determined microbiologically with Lactobacillus rhamnosus (ATCC 7469) as the growth indicator organism, and individual folates were determined by high-performance liquid chromatography after affinity chromatographic purification. Germination increased the folate content by 1.7-3.8-fold, depending on germination temperature, with a maximum content of 250 micro g/100 g dry matter. Hypocotylar roots with their notably high folate concentrations (600-1180 micro g/100 g dry matter) contributed 30-50% of the folate contents of germinated grains. Germination altered the proportions of folates, increasing the proportion of 5-methyltetrahydrofolate and decreasing the proportion of formylated folate compounds. Thermal treatments (extrusion, autoclaving and puffing, and IR and toasting) resulted in significant folate losses. However, folate levels in grains that were germinated and then were heat processed were higher than for native (nongerminated) grains. Opportunities to optimize rye processing to enhance folate levels in rye-based foods are discussed.

Folic Acid↗

Changes in oil, sugars and nitrogenous components during germination of sunflower seeds, Helianthus annuus.

Since germination has been found to improve the nutritional quality of grains, sunflower seeds Helianthus annuus were germinated for up to five days, and the cotyledons were analyzed for oil, sugar, free amino acid, non-protein nitrogen, lysine, tryptophan and methionine contents. Protein was fractionated. Soaking and germination increased the non-protein nitrogen, total free amino acid, lysine and tryptophan contents. Protein content and dry weight decreased. The oil content decreased significantly after 72 hours of germination. The reducing sugars increased gradually until day 'five' of germination. The saline soluble albumin and globulin fractions decreased while the glutelin content increased during germination. No changes were noticed in the prolamin and methionine contents.

Amino Acids↗

Nutritional composition and antinutritional factors of chickpeas (Cicer arietinum L.) undergoing different cooking methods and germination.

The effects of cooking treatments (boiling, autoclaving and microwave cooking) and germination on the nutritional composition and antinutritional factors of chickpeas were studied. Cooking treatments and/or germination caused significant (p < 0.05) decreases in fat, total ash, carbohydrate fractions, antinutritional factors, minerals and B-vitamins. Germination was less effective than cooking treatments in reducing trypsin inhibitor, hemagglutinin activity, tannins and saponins; it was more effective in reducing phytic acid, stachyose and raffinose. Cooking treatments and germination decreased the concentrations of lysine, tryptophan, total aromatic and sulfur-containing amino acids. However, cooked and germinated chickpeas were still higher in lysine, isoleucine and total aromatic amino acid contents than the FAO/WHO reference. The losses in B-vitamins and minerals in chickpeas cooked by microwaving were smaller than in those cooked by boiling and autoclaving. Germination resulted in greater retention of all minerals and B-vitamins compared to cooking treatments. In vitro protein digestibility, protein efficiency ratio and essential amino acid index were improved by all treatments. The chemical score and limiting amino acid of chickpeas subjected to the various treatments varied considerably, depending on the type of treatment. Based on these results, microwave cooking appears to be the best alternative for legume preparation in households and restaurants.

Amino Acids↗

Production of two highly active bacterial phytases with broad pH optima in germinated transgenic rice seeds.

Phytate is the main storage form of phosphorus in many plant seeds, but phosphate bound in this form is not available to monogastric animals. Phytase, an enzyme that hydrolyzes phosphate from phytate, has the potential to enhance phosphorus availability in animal diets when engineered in rice seeds as a feed additive. Two genes, derived from a ruminal bacterium Selenomonas ruminantium (SrPf6) and Escherichia coli (appA), encoding highly active phytases were expressed in germinated transgenic rice seeds. Phytase expression was controlled by a germination inducible alpha-amylase gene (alphaAmy8) promoter, and extracellular phytase secretion directed by an betaAmy8 signal peptide sequence. The two phytases were expressed in germinated transgenic rice seeds transiently and in a temporally controlled and tissue-specific manner. No adverse effect on plant development or seed formation was observed. Up to 0.6 and 1.4 U of phytase activity per mg of total extracted cellular proteins were obtained in germinated transgenic rice seeds expressing appA and SrPf6 phytases, respectively, which represent 46-60 times of phytase activities compared to the non-transformant. The appA and SrPf6 phytases produced in germinated transgenic rice seeds had high activity over broad pH ranges of 3.0-5.5 and 2.0-6.0, respectively. Phytase levels and inheritance of transgenes in one highly expressing plant were stable over four generations. Germinated transgenic rice seeds, which produce a highly active recombinant phytase and are rich in hydrolytic enzymes, nutrients and minerals, could potentially be an ideal feed additive for improving the phytate-phosphorus digestibility in monogastric animals.

6-Phytase↗

Studies on germination conditions and antioxidant contents of wheat grain.

Germination time and conditions for wheat grain were studied to determine optimum conditions that would maximize the production of antioxidants. Wheat grains were first steeped in water for 24 or 48 h, followed by incubation in the dark for 9 d at 98% RH and 16.5 degrees C. The changes in the concentration of vitamins C and E, beta-carotene, ferulic acid and vanillic acid were monitored over the germination period. Vitamins C and E and beta-carotene were barely detectable in the dry grains. However, upon germination the concentrations of these antioxidant vitamins steadily increased with increasing germination time, reaching their peaks after 7 d at 550 mu/g for vitamin C, 10.92 micrograms/g for alpha-tocopherol, and 3.1 micrograms/g for beta-carotene. Concentrations of ferulic and vanillic acids were also increased, reaching their maxima after 7 d at 932.4 micrograms/g and 12.9 micrograms/g, respectively. The grains steeped for 48 h before germination became wet, sticky, yellow-brown color with acidic smell after 7 d. These results suggested that wheat grains steeped for 24 h and germinated for 7 d would produce the most desirable sprouts with respect to antioxidant concentrations and sensory properties.

Antioxidants↗

Germinal centers without T cells.

Germinal centers are critical for affinity maturation of antibody (Ab) responses. This process allows the production of high-efficiency neutralizing Ab that protects against virus infection and bacterial exotoxins. In germinal centers, responding B cells selectively mutate the genes that encode their receptors for antigen. This process can change Ab affinity and specificity. The mutated cells that produce high-affinity Ab are selected to become Ab-forming or memory B cells, whereas cells that have lost affinity or acquired autoreactivity are eliminated. Normally, T cells are critical for germinal center formation and subsequent B cell selection. Both processes involve engagement of CD40 on B cells by T cells. This report describes how high-affinity B cells can be induced to form large germinal centers in response to (4-hydroxy-3-nitrophenyl) acetyl (NP)-Ficoll in the absence of T cells or signaling through CD40 or CD28. This requires extensive cross-linking of the B cell receptors, and a frequency of antigen-specific B cells of at least 1 in 1,000. These germinal centers abort dramatically at the time when mutated high-affinity B cells are normally selected by T cells. Thus, there is a fail-safe mechanism against autoreactivity, even in the event of thymus-independent germinal center formation.

Animals↗