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The relationship between coleoptile elongation and ethanolic fermentation was investigated in rice (Oryza sativa L.) coleoptiles of four cultivars subjected to a 48-h anoxic stress. The coleoptile elongation of all cultivars was suppressed by anoxic stress; however, the elongation of cvs Yukihikari and Nipponbare was much greater than that of cvs Leulikelash and Asahimochi. The stress did not significantly increase lactate dehydrogenase (LDH) activity or lactate concentration, but increased alcohol dehydrogenase (ADH) and pyruvate decarboxylase (PDC) activities, as well as ethanol concentration in the coleoptiles of all cultivars. The elevated ADH and PDC activities and ethanol concentration in cvs Yukihikari and Nipponbare were much greater than those of cvs Leulikelash and Asahimochi, suggesting that ethanolic fermentation is likely more active in cvs Yukihikari and Nipponbare than in cvs Leulikelash and Asahimochi. ATP concentration in cvs Yukihikari and Nipponbare in anoxia was also greater than that in cvs Leulikelash and Asahimochi in anoxia. The ethanol concentration in the coleoptiles was correlated with anoxia tolerance with respect to the ATP concentration and coleoptile elongation. These results suggest that the ability to increase ethanolic fermentation may be one of the determinants in anoxia tolerance of rice coleoptiles.

Rice seedlings (Oryza sativa L.) were subjected to low temperature pretreatment (LT-PT; 10°C) for various length of time (1, 2, 4, 6, 12, 18, 24 h) followed by a 48-h chilling temperature stress (2°C). Chilling temperature tolerance of rice roots was improved with increasing duration of LT-PT, but LT-PT longer than 12 h gave no additional improvement. Alcohol dehydrogenase activity and ethanol concentration in the roots were increased with increasing duration of LT-PT up to 12 h. Chilling tolerance was also improved by exogenously applied ethanol. These results suggest that LT-PT may increase chilling tolerance in rice roots owing to ethanol accumulation in the roots and LT-PT acclimation to chilling temperature may occur within 12 h.

Donor-receiver bioassay was designed to eliminate the effects of the competitive interference for resources from allelopathic effects. Seeds of cress (Lepidium sativum), lettuce (Lactuca sativa), timothy (Phleum pratense) or ryegrass (Lolium multiflorum) were incubated with 4-day-old buckwheat (Fagopyrum esculentum) seedlings for 3 days in Petri dishes under controlled laboratory condition. The growth of cress, lettuce, timothy and ryegrass seedlings was inhibited by the presence of buckwheat seedlings, and increasing the number of buckwheat seedlings increased the growth inhibition. One inhibiting substance was found in the culture solution in which buckwheat seedlings were hydroponically grown for 10 days. These results suggest that buckwheat seedlings may inhibit the neighboring plant growth due to exudation of allelopathic substance into the neighboring environment. Thus, the inhibitory effect of buckwheat was probably caused by allelopathic chemical reactions.

There is a rotational cropping system of wheat and rice in many areas. Wheat plants contain significant amounts of glucopyranosides of 6-methoxy-benzoxazolin-2(3H)-one (MBOA) and benzoxazolin-2(3H)-one (BOA). After crop harvesting, MBOA and BOA are produced during the degradation process of wheat residues left in the field. The purpose of this study was evaluation for the effect of MBOA and BOA on rice growth and germination. MBOA and BOA significantly inhibited the rice germination and the rice shoot growth at concentrations of 0.1 and 0.3 mM and higher, respectively. MBOA and BOA also inhibited the induction of α-amylase in rice seeds significantly at concentrations of 0.03 and 0.1 mM and higher, respectively, and inhibited gibberellin-induced α-amylase activity in de-embryonated rice. Significant inhibitions in germination, growth, and α-amylase activity were observed as MBOA and BOA concentrations increased. The inhibitions on the germination, growth, and α-amylase by MBOA were greater than those by BOA, which is the demethoxylated analogue of MBOA. These results suggest that MBOA and BOA may inhibit the germination and subsequent seedling growth of rice by inhibiting the gibberellin-induced α-amylase production. Therefore, wheat straw and roots left in the field after crop harvesting may suppress rice germination and growth, because of MBOA and BOA in the soil originated from wheat residues. Further studies are necessary for the determination of MBOA and BOA levels in field condition after the crop harvesting.

Background As a means of biologically controlling Mikania micrantha H.B.K. in Yunnan, China, the influence of sweet potato [Ipomoea batatas (L.) Lam.] on its reproductive characteristics was studied. The trial utilized a de Wit replacement series incorporating six ratios of sweet potato and M. micrantha plants in 25 m2 plots over 2 years. Results Budding of M. micrantha occurred at the end of September; flowering and fruiting occurred from October to February. Flowering phenology of M. micrantha was delayed (P

The labdane-related diterpenoid, momilactone B has potent growth inhibitory activity and was demonstrated to play a particularly critical role in the allelopathy of rice ( Oryza sativa L.). However, there is limited information available about the mode of action of momilactone B on the growth inhibition. The present research describes the effects of momilactone B on protein expression in the early development of Arabidopsis thaliana seedling, which was determined by two-dimensional electrophoresis and MALDI-TOFMS. Momilactone B inhibited the accumulation of subtilisin-like serine protease, amyrin synthase LUP2, β-glucosidase and malate synthase at 1 h after the momilactone application. Those proteins are involved in the metabolic turnover and the production of intermediates needed for cell structures resulting in plant growth and development. Momilactone B also inhibited the breakdown of cruciferin 2, which is essential for seed germination and seedling growth to construct cell structures. Momilactone B induced the accumulation of translationally controlled tumor protein, glutathione S-transferase and 1-cysteine peroxiredoxin 1. These proteins are involved in stress responses and increased stress tolerance. In addition, glutathione S-transferase has the activity of herbicide detoxification and 1-cysteine peroxiredoxin 1 has inhibitory activity for seed germination under unfavorable conditions. The present research suggests that momilactone B may inhibit the seedling growth by the inhibition of the metabolic turnover and the production of intermediates for cell structures. In addition, momilactone induced proteins associated with plant defense responses.

In this study, an allelopathic substance was isolated from an aqueous methanol extract of Hibiscus sabdariffa L. by column chromatography and reverse phase HPLC. The chemical structure of the substance was determined by 1H NMR spectroscopy and mass spectrometry as trimethyl allo-hydroxycitrate. Trimethyl allo-hydroxycitrate inhibited the growth of cress hypocotyls and roots at concentrations greater than 10 mM. The concentrations required for 50% growth inhibition of the hypocotyls and roots of cress were 20.3 and 14.4 mM, respectively. The inhibitory activity of trimethyl allo-hydroxycitrate suggests that the substance may act as an allelopathic substance of H. sabdariffa.

The grass Brachiaria decumbens becomes naturalized and quickly dominant in non-native areas. It was hypothesized that phytotoxic substances of plants may contribute to the domination and invasion of the plants. However, no potent phytotoxic substance has been reported in B decumbens. Therefore, we searched for phytotoxic substances with allelopathic activity in this species. An acpeous methanol extract of B. decumbens inhibited the growth of roots and shoots of cress (Lepidium sativum), lettuce (Lactuca sativa), timothy (Phleum pratense) and ryegrass ilolium multiflorum) seedlings. The extract was then purified using chromatographic methods and a phytotoxic substance with allelopathic activity was isolated and identified by spectral analysis as ( 6R,9S)-3-oxo-α-ionol. These results suggest that this compound may contribute to the allelopathic effect caused by the B. decumbens extract and may be in part responsible for the invasion and domination of B. decumbens. Two other Brachiaria species, B. brizantha and a Bractearia hybrid were also confirmed to contain ( 6R,9S)-3-oxo-α-ionol. Therefore, this compound may play an important role in the phytotoxicity of the Brachiaria species.

Although Plantago major and Plantago asiatica have similar life-history characteristics, P. major populations expand much more rapidly than P. asiatica in habitats where both species occur. One potential factor contributing to this disparity is the production of phytotoxic compounds. However, the phytotoxic characteristics of P. major and P. asiatica have not yet reported. Therefore, we conducted an investigation of possible phytotoxic compounds with allelopathic activity in both species. Aqueous methanol extracts of both plant species inhibited root and hypocotyl growth of garden cress (Lepidium sativum) in a concentration dependent manner. However, the inhibitory effect of P. major was greater than that of P. asiatica. The extracts were further purified by several chromatographic runs and two main phytotoxic compounds were isolated and identified through spectral analysis as (−)-3-hydroxy-β-ionone and blumenol A. (−)-3-Hydroxy-β-ionone and blumenol A inhibited the growth and germination of garden cress at concentrations greater than 1 and 30 μM, respectively. The level of (−)-3-hydroxy-β-ionone in P. major was 7.2-fold greater than that in P. asiatica, while the level of blumenol A was equal in both plant species. Although these Plantago species produce two main phytotoxic compounds with allelopathic activity, the higher level of (−)-3-hydroxy-β-ionone in P. major may result in it contributing the primary inhibitory effects of the extract of P. major. (−)-3-hydroxy-β-ionone may also serve to distinguish the invasive habit of two species and may be involved in the expansion in habitats of P. major.

Background There are a variety of ways of increasing crop diversity to increase agricultural sustainability and in turn having a positive influence on nearby natural ecosystems. Competitive crops may provide potent management tools against invasive plants. To elucidate the competitive mechanisms between a sweet potato crop (Ipomoea batatas) and an invasive plant, mile-a-minute (Mikania micrantha), field experiments were carried out in Longchuan County of Yunnan Province, Southwest China, utilizing a de Wit replacement series. The trial incorporated seven ratios of sweet potato and mile-a-minute plants in 25 m2 plots. Results In monoculture, the total biomass, biomass of adventitious root, leafstalk length, and leaf area of sweet potato were all higher than those of mile-a-minute, and in mixed culture the plant height, branch, leaf, stem node, adventitious root, flowering and biomass of mile-a-minute were suppressed significantly (P

Aqueousmethanol extracts of Cit rus junos, C. unshiu and C. sudachi fruit peel in hib ited the growth of the roots and hypocotyls of alfalfa (Medicago sativa L.), cress (Lepidiumsativum L.) and lettuce (Lactuca sativa L.) seedlings. Signifi cantreductions in the root and hypocotyl growth were observed as the extract concentration in creased in all bioassays. The in hibitory activity of C. junos ex tract on the growth of test plants was 3.3-to 17.9-fold and 3.6- to 20.6-fold greater than that of C. unshiu and C. sudachi extracts, respectively. Theconcent rationin C. junos was 3.5- and 4.9-fold greater than that in C. unshiu and C. sudachi, re spectively. Thus, there was a good correlation-between abscisic acid-b-D-glucopyranosylester (ABA-GE) concentrationsin C. junos, C. unshiu and C. sudachi fruit peel and theinhibito ryactivities of the irextracts. The concentration of ABA-GE in C. junos fruit peel was in creased with fruit maturityasgrowthinhibit oryactivity of C. junos fruit peel was reported to be in creased with fruit maturity, in dicating that the concentrations of ABA-GEin C. junos fruit peel was correlated withgrowthinhibit oryactivity of C. junos fruit peel in time course of fruit maturation. These findings suggest that ABA-GE may be involved in the growth inhibitory effect of C. junos, C. unshiu and C. sudachi fruit peel.

Relative importance of ethanolic, lactate and alanine fermentation pathways was estimated in coleoptiles of rice seedlings (Oryza sativa L.) subjected to anoxic stress. The in vitro activities of alcohol dehydrogenase (ADH, EC 1.1.1.1), pyruvate decarboxylase (PDC, EC 4.1.1.1) and alanine aminotransferase (AlaAT, EC 2.6.1.2) in the coleoptiles increased in anoxia, whereas no significant increase was measured in lactate dehydrogenase (LDH, EC 1.1.1.27) activity. At 48 h, the ADH, PDC and AlaAT activities in anoxic coleoptiles were 62-, 15- and 7.6-fold greater, respectively, than those in the presence of oxygen. Ethanol and alanine in the coleoptiles accumulated rapidly under anoxia, increasing by 48 h, 57- and 5.6-fold compared with those in the presence of oxygen, respectively. However, lactate concentration did not increase and no initial burst of lactate production was detected. The relative ratio of carbon flux from pyruvate to ethanol, lactate and alanine in anoxic coleoptiles was estimated to be 92, 1 and 7% of the total carbon flux, respectively. These results suggest that the potential carbon flux from pyruvate to ethanol may be much greater than the potential flux from pyruvate to lactate and alanine in rice coleoptiles during anoxia.

Effects of anoxia on the levels of free-amino acids were investigated in the coleoptiles of rice (Oryza sativa L.) seedlings. Rice coleoptiles are able to grow in extremely low oxygen conditions. Anoxic stress increased the concentration of total free-amino acids in the coleoptiles. Alanine (Ala) and γ-aminobutyric acid (Gaba) were the main amino acids accumulated. After 48 h, Ala and Gaba concentrations in anoxic coleoptiles were 3.4- and 11.2-fold greater than those in non-stressed coleoptiles, respectively. Ala and Gaba represented 19 and 23 % of the amino acid pool in anoxic coleoptiles, respectively. Submergence stress also increased Ala and Gaba concentrations in rice coleoptiles. Since Ala and Gaba are bio-compatible solutes and their accumulation is known to stabilize osmotic potential and/or cytoplasmic pH in plant cells, these stress-induced amino acids may allow rice coleoptiles to make biochemical adjustment that enable them to cope with the stress conditions. Therefore, the ability to increase the concentrations of Ala and Gaba may be important for anoxic and submergence stress tolerance of rice seedlings.

The allelopathic potential of an aqueous methanol extract of Citrus junos leaves was investigated under laboratory conditions. The extract inhibited the growth of roots and shoots of cress (Lepidium sativum L.), lettuce (Lactuca sativa L.) and alfalfa (Medicago sativa L.) seedlings. Significant reductions in the root and shoot growth were observed as the extract concentration increased. Since abscisic acid -β-D-glucopyranosyl ester (ABA-GE) was found to be a major inhibitor in fruit peel of C. junos, ABA-GE in C. junos leaves was determined and the concentration was 540 μg kg-1 fresh weight in the leaves harvested on September 26, 2002. These results suggest that C. junos leaves possess allelopathic activity and could work as a weed inhibiting agent, which should be investigated further in the field for practical application.

Germination of lettuce seeds was inhibited by 6-methoxy-2-benzoxazolinone (MBOA) at concentrations greater than 0.03 mmol/L. MBOA also inhibited the induction of alpha-amylase activity in the lettuce seeds at concentrations greater than 0.03 mmol/L. These two concentration-response curves for the germination and alpha-amylase indicate that the percentage of the germination was positively correlated with the activity of alpha-amylase in the seeds. Lettuce seeds germinated around 18h after incubation and inhibition of alpha-amylase by MBOA occurred within 6h after seed incubation. These results show that MBOA may inhibit the germination of lettuce seeds by inhibiting the induction of alpha-amylase activity.

Fructose analog, psicose, and glucose analog, mannose, inhibited root growth of lettuce seedlings. Psicose is phosphorylated by hexokinase and fructokinase (EC 2.7.1.4) to psicose-6-phosphate with no known capacity for further metabolism. Mannose is phosphorylated by hexokinase (EC 2.7.1.1) to mannose-6-phosphate which is further metabolized very slowly. Hexokinase is known to have a sugar-sensing function and possibly triggers a signal cascade resulting in changes of several gene expressions. It was determined, compared with the behaviour of mannose, whether psicose inhibits the root growth through this system. The addition of phosphate into the growth medium of lettuce seedlings did not affect the inhibition by psicose and mannose, and both sugars did not reduce adenosine triphosphate (ATP) level in the roots, suggesting that the inhibition is not due to phosphate starvation and ATP depletion. The inhibiting effects of psicose and mannose were overcome by adding sucrose into the medium, which suggests that the inhibition is not caused by accumulation of psicose-6-phosphate or mannose-6-phosphate in the seedlings. Mannoheptulose, a specific competitive inhibitor of hexokinase, defeated the mannose-induced inhibiting but was not able to relieve the psicose-induced inhibition. Thus, the phosphorylation of mannose by hexokinase may trigger a signal cascade resulting in the growth inhibition of lettuce roots, which is consistent with the hypothesis established in Arabidopsis. However, psicose cannot inhibit the growth of lettuce roots via a hexokinase-mediated pathway, and the phosphorylation of psicose by fructokinase might trigger a hexokinase-independent signal cascade resulting in the growth inhibition.

Effects of anoxia on the levels of free-amino acids were investigated in the coleoptiles of rice (Oryza sativa L.) seedlings. Rice coleoptiles are able to grow in extremely low oxygen condition. Anoxic stress increased the concentration of total free-amino acids in the coleoptiles. After 48 h, alanine (Ala) and γ-aminobutyric acid (Gaba) in anoxic coleoptiles were 2.7- and 10.6-fold greater than those in non-stressed coleoptiles, respectively. Ala and Gaba represented 40% of the amino acid pool in anoxic coleoptiles. These anoxic-induced amino acids may allow rice coleoptiles to make biochemical adjustment that enable them to cope with the stress condition.

The allelopathic potential of Citrus junos fruit waste after juice extraction was investigated. Aqueous methanol extracts of peel, inside and seeds separated from the fruit waste inhibited the growth of the roots and shoots of alfalfa (Medicago sativa L.), cress (Lepidium sativum L.), crabgrass (Digitaria sanguinalis L.), lettuce (Lactuca sativa L.), timothy (Pheleum pratense L.), and ryegrass (Lolium multiflorum Lam.). The inhibitory activity of the peel extract was greatest and followed by that of the inside and seed extracts in all bioassays. Significant reductions in the root and shoot growth were observed as the extract concentration was increased. The concentrations of abscisic acid-β- d -glucopyranosyl ester (ABA-GE) in peel, inside and seeds separated from the C. junos fruit waste were determined, since ABA-GE was found to be one of the main growth inhibitors in C. junos fruit. The concentration was greatest in the peel, followed by the inside and seeds; there was a good correspondence between these concentrations and the inhibitory activities of the extracts. This suggests that ABA-GE may also be involved in the growth inhibitory effect of C. junos waste. These results suggested that C. junos waste may possess allelopathic potential, and the waste may be potentially useful for weed management.

Although most cereal roots cannot elongate under anoxic conditions, primary roots of three-day-old rice (Oryza sativa L.) seedlings were able to elongate during a 24-h period of anoxia. Hypoxic pretreatment (H-PT) increased the elongation of their roots. Sucrose synthase (EC 2.4.1.13), glucokinase (EC 2.7.1.2), fructokinase (EC 2.7.1.4), pyruvate decarboxylase (EC 4.1.1.1) and alcohol dehydrogenase (EC 1.1.1.1) activities were increased by anoxia in both H-PT and non-pretreated (N-PT) roots. However, these activities were greater in the H-PT roots than in the N-PT roots. The average rate of production of ethanol for the initial 6h after the onset of anoxia was 3.7 and 1.4 micromolg(-1) fresh weight h(-1) for the H-PT and N-PT roots, respectively, suggesting that ethanolic fermentation may increase more quickly in the H-PT roots than in the N-PT roots. Roots of the seedlings lost ATP and total adenine nucleotides in anoxia, however, the H-PT roots maintained higher levels of ATP and total adenine nucleotides compared to the N-PT roots. These results show that rice roots are able to utilize the set of enzymes involved in the metabolism of soluble sugars under anoxia. The ability to maintain an active fermentative metabolism for production of ATP by fueling the glycolytic pathway with fermentable carbohydrate is probably greater in H-PT than in N-PT roots.

The growth of the first three internodes of tall (cv. Alaska) and dwarf (cv. Progress No. 9) cultivars of 6-day-old pea (Pisum sativum L.) seedlings was similar in darkness. Red light inhibited the growth of Progress No. 9 to a greater extent than Alaska, which indicates that the dwarf habit of Progress No. 9 is manifest only in red light. Gene expression of gibberellin (GA) 3β-hydroxylase, which converts inactive GA20 to biologically active GA1, in the internodes of both pea cultivars was higher in darkness than in red light. Changes in the transcript levels of the GA 3β-hydroxylase gene in dark-grown seedlings of Alaska and Progress No. 9 were monitored after being transferred to red light. The transcript levels in apical buds of both peas increased during the first 3 h and then decreased, which may relate to the morphological changes in the apical buds. Red light gradually decreased the transcript levels in the 3rd internodes of both cultivars and the levels at 24 h were 46% and 44% of the initial levels for Alaska and Progress No. 9, respectively. The differences in transcript levels cannot, therefore, account for the growth habit differences. These results indicate that the growth habit of Progress No. 9 in red light is unlikely to be due to a light-induced blockage in transcription of the GA 3β-hydroxylase gene.

Ability of metabolic adaptation in upland and lowland rice (Oryza sativa L.) seedlings to flooding stress was compared. Flooding stress increased alcohol dehydrogenase (ADH) activity and ethanol concentration in shoots and roots of the upland and lowland rice seedlings. The difference in ADH activity and ethanol concentration in shoots between the upland and lowland rice was not apparent. However, both ADH activity and ethanol concentration in roots of the lowland rice were 2-fold greater than those in roots of the upland rice, suggesting that flooding-induction of ethanolic fermentation in lowland rice roots may be significantly greater than that in the upland rice roots. Since flooding often causes the anaerobic conditions in rooting zone than aerial part of plants and ethanolic fermentation is essential to survive in the anaerobic conditions, the ability of metabolic adaptation in lowland rice seedlings to flooding stress may be greater than that in upland rice seedlings.

The residue of peas (Pisum sativum L.) has allelopathic activity and the putative compound causing this inhibitory effect was isolated from a methanol extract of pea shoots. Chemical structure of this compound was determined by high-resolution MS, IR and 1H NMR spectral data as pisatin. Pisatin inhibited growth of cress (Lepidium sativum L.) and lettuce (Lactuca sativa L.) seedlings at concentrations greater than 10 and 30 microM, respectively. The doses required for 50% growth inhibition of roots and hypocotyls of cress were 61 and 91 microM, respectively, and those of lettuce were 78 and 115 microM, respectively. The concentration of pisatin in the pea shoots was 32.7 nmol x g(-1) fresh weight. The effectiveness of pisatin on growth inhibition in cress and lettuce, and its occurrence in pea shoots suggest that it may contribute to the growth inhibitory effect of pea residue, and may play an important role in pea allelopathy.

The allelopathic potential of lemon balm (Melissa officinalis) was investigated under laboratory conditions. The shoot powder of 30-day-old plants inhibited the germination and the growth of roots and shoots of Amaranthus caudatus, Digitaria sanguinalis and lettuce (Lactuca sativa). Significant reductions in the germination and growth of roots and shoots were observed as the powder concentration increased. The concentration-dependent responses of the test plants to the powder of lemon balm suggest that the powder might contain allelochemicals, which should be investigated further in the laboratory and the field for practical application of the powder as weed inhibiting agent.

Rice (Oryza sativa L. cv. Soja-Akamai) seedlings accelerated the growth of roots of alfalfa (Medicago sativa L.) seedlings when both seedlings were grown together. Growth promoting activity was found in the culture solution in which the rice seedlings were hydroponically grown for 15 d. These findings suggest that this cultivar of rice may produce and release a growth promoting substance into the culture solution or neighboring environment.

Aqueous methanol extracts of Citrus junos, C. unshiu and C. hassaku fruit peel inhibited the growth of the roots and hypocotyls of lettuce (Lactuca sativa L.) seedlings. Significant reductions in the root and hypocotyl growth were observed as the extract concentration increased in all bioassays. The inhibitory activity of C. junos extract on the growth of lettuce roots and hypocotyls was about 13- and 24-fold greater than that of C. unshiu and C. hassaku extracts, respectively. The concentration of abscisic acid-β-D-glucopyranosyl ester (ABA-GE) in fruit peel of C. junos, C. unshiu and C. hassaku was determined since ABA-GE had been found to be a possible cause of the growth inhibitory effect of C. junos. The concentration was 132, 10.6 and 5.0 µg g−1 dry weight in C. junos, C. unshiu and C. hassaku fruit peel, respectively. Thus, there was a good correlation between ABA-GE concentrations in C. junos, C. unshiu and C. hassaku fruit peel and the inhibitory activities of their aqueous extracts, which suggests that ABA-GE may be involved in the growth inhibitory effect of C. junos, C. unshiu and C. hassaku fruit peel.

The catalytic direction of pyrophosphate:fructose-6-phosphate 1-phosphotransferase (PFP; EC 2.7.1.90) in coleoptiles of rice (Oryza sativa L.) seedlings subjected to anoxia stress is discussed. The stress greatly induced ethanol synthesis and increased activities of alcohol dehydrogenase (ADH; EC 1.1.1.1) and pyruvate decarboxylase (PDC; EC 4.1.1.1) in the coleoptiles, whereas the elevated PDC activity was much lower than the elevated ADH activity, suggesting that PDC may be one of the limiting factors for ethanolic fermentation in rice coleoptiles. Anoxic stress decreased concentrations of fructose 6-phosphate (Fru-6-P) and glucose 6-phosphate, and increased concentration of fructose 1,6-bisphosphate (Fru-1,6-bisP) in the coleoptiles. PFP activity in rice coleoptiles was low in an aerobic condition and increased during the stress, whereas no significant increase was found in ATP:fructose-6-phosphate 1-phosphotransferase (PFK; EC 2.7.1.11) activity in stressed coleoptiles. Fructose 2,6-bisphosphate concentration in rice coleoptiles was increased by the stress and pyrophosphate concentration was above the Km for the forward direction of PFP and was sufficient to inhibit the reverse direction of PFP. Under stress conditions the potential of carbon flux from Fru-6-P toward ethanol through PFK may be much lower than the potential of carbon flux from pyruvate toward ethanol through PDC. These results suggest that PFP may play an important role in maintaining active glycolysis and ethanolic fermentation in rice coleoptiles in anoxia.

A search for growth inhibitors in rice root exudates was undertaken in order to clarify the allelopathic system in rice (Oryza sativa L.). Rice seedlings inhibited the growth of cress (Lepidium sativum L.) and lettuce (Lactuca sativa L.) seedlings when the cress and lettuce were grown with rice seedlings. The putative compound causing the inhibitory effect of rice seedlings was isolated from their culture solution, and the chemical structure of the inhibitor was determined by spectral data as momilactone B. Momilactone B inhibited the growth of cress and lettuce seedlings at concentrations greater than 3 and 30 microM, respectively. The concentration of momilactone B was 3.4 and 1.1 nmol per seedling in the culture solutions of husked and non-husked rice seedlings, respectively. These results suggest that rice seedlings may release momilactone B into the environment and the stress caused by the husk-treatment may increase the amount of momilactone B released. Thus, momilactone B may play an important role in rice allelopathy.

To clarify the allelopathic system of lemon balm (Melissa officinalisL.), purification and isolation of allelopathic substances was undertaken in acetone extract of the shoots of the 30-d-old plants. Three growth-inhibiting substances were found in the neutral fraction of the extract after silica gel column chromatography, and most active substance was further purified and isolated by thin layer chromatography, C18Sep Pack cartridge and reversephase high performance liquid chromatography. This substance inhibited the growth of cress (Lepidium sativunL.) at concentrations greater than 0.3 μg mL-1and the concentration required for 50% growth inhibition was 2.9μg mL-1. These results suggest that lemon balm shoots may contain at least three growth-inhibiting substances, which may be released into the environment, either as exudates from living tissues or leachates from residues of the plant, and may act as allelochemicals to neighboring plants.

Summary The changes in growth and gene expression of gibberellin (GA) 3β-hydroxylase were determined in dark-grown, 6-day-old, tall (cv. Alaska) and dwarf (cv. Progress No. 9) pea ( Pisum sativum L.) seedlings after transfer to conditions of red light irradiation. Red light inhibited the shoot elongation of Progress No. 9 much more so than that of Alaska, and the elongation of Progress No. 9 during 48 h was 23 percnt; that of Alaska. The transcript level of the GA 3β-hydroxylase gene was slightly greater in the shoots of dark-grown Alaska than that of dark-grown Progress No. 9. The red light decreased the transcript levels in both cultivars and the levels at 48 h were about 50 percnt; and 60 percnt; compared to the initial levels for Alaska and Progress No. 9, respectively. These results indicate that the difference in the transcript levels in both cultivars is not apparent. The only difference in GA 3β-hydroxylase proteins between Alaska and Progress No. 9 was the replacement of alanine with threonine at position 229 in their amino acid residues, which may affect the activity of the enzyme and may, at least partially, contribute to the growth habits of those cultivars. A possibility of additional function to distinguish the growth habits of those cultivars is also discussed here.

Summary Seedlings of Avena sativa were subjected to 24 h anoxic stress after 24 h hypoxic-pretreatment (H-PT). The H-PT increased 4.5-fold the survivability of the anoxia in coleoptiles of the seedlings compared to that of non-pretreated (N-PT) seedlings. During pretreatment periods, alcohol dehydrogenase (ADH, EC 1.1.1.1) and pyruvate decarboxylase (PDC, EC 4.1.1.1) activities in the coleoptiles became 4.1- and 3.9-fold greater respectively than those of N-PT seedlings. After the onset of anoxic stress, ADH and PDC activities in all coleoptiles increased, but their activities in the H-PT coleoptiles remained much greater than those in the N-PT coleoptiles, and the average production rate of ethanol for the initial 6 h was 3.1 and 1.4 μmol g −1 fresh weight h −1 for the H-PT and N-PT coleoptiles, respectively. Coleoptiles of the seedlings lost ATP rapidly under anoxic stress; however, the decrease in ATP was much slower in the H-PT coleoptiles than in the N-PT coleoptiles. These results suggest that the H-PT may improve the subsequent tolerance of anoxia in Avena coleoptiles due to maintenance of ATP levels with rapid induction of ethanolic fermentation. The present results are consistent with the results that were found in roots of several plants, suggesting that the mechanism in hypoxic acclimation to anoxia implicated in their roots may act in Avena coleoptiles.

Hypoxic pretreatment (H-PT; incubated in 5% O2 atmospherefor 24 h) greatly improved subsequent tolerance of anoxia in theprimary roots of Avena sativa. Survival of H-PT roots inanoxia increased by 6.4-fold compared to that of non-pretreated (N-PT) roots ofthe seedlings. ATP concentration in the H-PT roots was 4.8-fold greater thanthat in the N-PT roots at 12 h after onset of anoxia. Duringpretreatment periods, alcohol dehydrogenase (EC 1.1.1.1) and pyruvatedecarboxylase (EC 4.1.1.1) activities in the roots became 7.5- and 4.3-foldgreater than those in the N-PT roots, respectively. After the onset of anoxicstress, their activities in the H-PT roots remained greater than those in theN-PT roots, and the average production rate of ethanol was much greater in theH-PT roots than N-PT roots. These results suggest that the H-PT may increaseanoxia tolerance due to maintenance of anaerobic glycolysis with induction ofethanolic fermentation to generate ATP.

A phytotoxic compound was isolated from the root extracts of Cymbopogon nardus by bioassay-guided purification and characterized to be myrislignan. Myrislignan significantly inhibited seedling growth of cress at a concentration greater than 100 μM (p < 0.05). The concentrations required for 50% growth inhibition of myrislignan on hypocotyl and root growth of cress were 429 μM and 517 μM, respectively. The inhibition on seedling growth indicated that myrislignan had a phytotoxic activity and may contribute to the phytotoxic effect of C. nardus.

Cyanotis axillaris, an annual succulent weed in the family Commelinaceae, was taken in this experiment to explore its allelopathic potential as plant allelopathy is considered as a biological option to control weeds. The aqueous methanol extracts of C. axillaris were applied on the growth of four dicotyledonous (cress, lettuce, alfalfa and rapeseed), and four monocotyledonous (barnyard grass, Italian ryegrass, timothy and foxtail fescue) test plant species at six different concentrations. The C. axillaris extracts inhibited the shoot and root growth of all test plant species, and such inhibition increased with increasing extract concentrations. Cress and lettuce showed complete growth inhibition at the concentration of 0.1 g dry weight equivalent extract/ml, whereas alfalfa, rapeseed, barnyard grass, Italian ryegrass, timothy and foxtail fescue showed growth inhibition greater than 50% of the control growth. At a concentration of 0.3 g dry weight equivalent extract/ml of C. axillaris extracts, cress, lettuce, alfalfa and rapeseed showed complete growth inhibition and that inhibition was greater than 30% of the control growth for barnyard grass, Italian ryegrass, timothy and foxtail fescue. Comparing the required concentration for 50% growth inhibition, lettuce was more susceptible and timothy was less susceptible with the extracts. Results indicate that C. axillaris may have allelopathic activity and may contain growth inhibitory substances. Therefore, C. axillaris could be a potential candidate to develop environment-friendly bio-herbicides for weed control.

Acacia concinna (Willd,) DC. (Fabaceae) has been used as a traditional medicine and well-known for many of their pharmacological and toxicological properties. However, the allelopathic property of the plant have not yet been evaluated. Therefore, we investigated the allelopathic activity of Acacia concinna pods. Aqueous methanol extracts of A. concinna pods inhibited the growth of cress (Lepidium sativum L.), lettuce (Lactuca sativa L.), alfalfa (Medicago sativa L.), rapeseed (Brassica napus), barnyard grass (Echinochloa crus-gallis L.), Italian ryegrass (Lolium multiflorum Lum.), foxtail fescue (Vulpia myuros (L.) C. C. Gmel.) and timothy (Phleum pretense L.) at concentrations greater than 0.003 and 0.001 g dry weight equivalent extract/mL for their shoots and roots, respectively. The inhibitory effect was increased with increasing the concentration of the pod extracts. The concentrations required for 50% inhibition of shoot and root growth of the eight test plant species ranged between 1.00 to 5.17 and 0.02 to 2.59 g dry weight equivalent extract/mL, respectively. Root growth of all test plant species were more sensitive to the extract than shoot growth of them. The extract was purified by column chromatographies and reverse phase HPLC, and an active substance (ACP-1) was isolated. The present result indicates that A. concinna has allelopathic activity and ACP-1 may be responsible for the allelopathic effect of A. concinna. The pods and the pod extracts of A. concinna could be useful for one of the weed management options.

The fern Gleichenia japonica often dominates plant communities by forming large monospecific stands throughout the temperate to tropical Asia. The objective of this study was the investigation of allelopathic property and substances of the decomposing litter of the fern to evaluate the possible involvement of its allelopathy in the domination. An aqueous methanol extract of G. japonica litter inhibited the growth of garden cress (Lepidium sativum), lettuce (Lactuca sativa), barnyard grass (Echinochloa crus-galli), and ryegrass (Lolium multiflorum). This result suggests that G. japonica litter contains growth inhibitory substances. The extract was purified by chromatography while monitoring the inhibitory activity, and a growth inhibitory substance was isolated. The chemical structure of the substance was determined by spectral data to be a novel compound, 13-O-β-fucopyranosyl-3β-hydroxymanool. This compound inhibited root and shoot growth of garden cress and barnyard grass at concentrations ranging from 89.7 to 271 μM for 50% inhibition. In addition, the compound had potent growth inhibitory activity with the soil taken from near the colony. The concentration of the compound in soil under a pure colony of G. japonica was 790 μM, suggesting that the compound may contribute to the establishment of monocultural stands by this fern.

Background Centrostachys aquatica is a perennial emergent macrophyte in marshy places and in rivers. The species was recorded in Senegal and Nigeria, but widespread in tropical Africa, and South and East Asia. Aqueous methanol extracts C. aquatica was found to be toxic to several plant species. However, no phytotoxic substance has been reported in this species. Therefore, we investigated phytotoxic activity and searched for phytotoxic substances with allelopathic activity in C. aquatica. Results An aqueous methanol extract of C. aquatica inhibited the growth of roots and hypocotyls of cress (Lepidium sativum). The extract was then purified by several chromatographic runs and a phytotoxic substance with allelopathic activity was isolated and identified by spectral analysis as loliolide. Loliolide inhibited cress root and hypocotyl growth at concentrations greater than 0.03 μM. The concentrations required for 50% growth inhibition of cress roots and hypocotyls was 0.18 and 0.15 μM, respectively. Conclusion These results suggest that loliolide is a phytotoxic substance and may contribute to the allelopathic effect caused by C. aquatica. Electronic supplementary material The online version of this article (doi:10.1186/s40529-014-0059-1) contains supplementary material, which is available to authorized users.

Neem (Azadirachta indica) has been widely used as a traditional medicine and several bioactive compounds have been isolated from this species, but to date no potent allelopathic active substance has been reported. Therefore, we investigated possible allelopathic property and phytotoxic substances with allelopathic activity in neem. An aqueous methanol extract of neem leaves inhibited the growth of roots and shoots of cress, lettuce, alfalfa, timothy, crabgrass, ryegrass, barnyard grass and jungle rice. The extracts were then purified by several chromatographic runs while monitoring the inhibitory activity and two phytotoxic substances were isolated. The chemical structures of the two substances were determined by spectral data to correspond to novel compounds, nimbolide B (1) and nimbic acid B (2). Nimbolide B inhibited the growth of cress and barnyard grass at concentrations greater than 0.1‒3.0 μM. Nimbic acid B inhibited the growth of cress and barnyard grass at concentrations greater than 0.3–1.0 μM. These results suggest that nimbolide B and nimbic acid B may contribute to the allelopathic effects caused by neem leaves.

Onopordum acanthium L. (Asteraceae) is a plant native to southern Europe and southwestern Asia, but it is invasive in disturbed areas and agricultural fields around the world, causing many agronomic problems by interfering with crops or preventing animals from grazing on pastures. Allelopathy could be one of the reasons that this plant has spread over different continents. The aim of the present study was to bioprospect O. acanthium leaf extracts through the isolation and purification of allelopathic secondary metabolites with phytotoxicity to explain their invasive behavior. Phytotoxic activity was tested using etiolated wheat coleoptiles. The most active extract was selected to perform a bioassay-guided isolation of two flavonoids, pectolarigenin (1) and scutellarein 4’-methyl ether (2), and two sesquiterpene lactones, elemanolide 11(13)-dehydromelitensin b-hydroxyisobutyrate (3) and acanthiolide (4). All compounds were isolated for the first time from O. acanthium, and acanthiolide (4) is described for the first time. Compound 3 strongly inhibited the growth of wheat coleoptiles and 1 showed an intermediate effect. The results indicate that these compounds could contribute to the invasion of O. acanthium in ecological systems and agricultural fields.

To determine the allelopathic potential of root exudate from early developmental stage of rice (Oryza sativa L), 6-d-old seedlings of eight cultivars were grown with 3-d-old alfalfa (Medicago sativa L.), cress (Lepidium sativum L.) or lettuce (Lactuca sativa L.) seedlings in Petri dishes under controlled condition. All rice cultivars (cv. Norin 8, Kamenoo, Nipponbare, Kinuhikari, Koshihikari, Sasanishiki, Yukihikari and Hinohikari) inhibited growth of roots, shoots and fresh mass of alfalfa, cress and lettuce seedlings. Effectiveness of cv. Koshihikari was the greatest and more than 60% inhibition was recorded in all bioassays, followed by that of cv. Norin 8 of which effectiveness was more than 40%.

The n-hexane-, acetone- and water-soluble fractions obtained from an aqueous acetone extract of lemon balm (Melissa officinalis L.) shoots inhibited the germination and the growth of roots and shoots of cockscomb (Amaranthus caudatus L.), cress (Lepidium sativum L.), crabgrass (Digitaria sanguinalis L.), timothy (Phleum pratense L.), lettuce (Lactuca sativa L.) and ryegrass (Lolium multiflorum Lam.). The inhibitory activity of the water-soluble fraction was the greatest, followed by that of acetone- and n-hexane-soluble fractions in all bioassays. The effectiveness of these fractions on the roots was greater than that of the shoots of the test plants. Significant reductions in the germination and growth of the roots and shoots were observed as the extract concentration increased. Such rate-dependent responses of the test plants to the fractions suggest that each fraction might contain allelochemical(s), but that the greatest potential was in the water-soluble fraction.

The relationship between primary root growth and ethanolic fermentation was investigated in five cultivars of germinating rice (Oryza sativa L.) seeds subjected to 48 h-anoxic stress. The anoxic stress inhibited the growth of all rice cultivars, however, there were significant differences in the growth among the cultivars. The stress increased alcohol dehydrogenase activities and ethanol concentrations in the roots of all rice cultivars but there were differences between cultivars in the activity and the concentration. The ethanol concentrations in the roots were closely correlated with the root growth of the corresponding cultivars. These results suggest that ability to induce ethanolic fermentation may play an important role in the primary root growth of germinating rice in anoxic condition.

Effects of ethanol, the end product of ethanolic fermentation, on the growth of rice (Oryza sativa L.) seedlings were determined as a means of evaluating growth responses under anoxia. The ethanol concentrations in roots and coleoptiles of the seedlings subjected to 48 h-anoxia, and in their culture medium were 23 and 32 µmol g−1 fresh weight, and 19 µmol ml−1, respectively. The growth of the roots and coleoptiles of the seedlings was restricted by exogenous ethanol at concentrations above 50 mM and 100 mM, respectively, suggesting that the roots are more sensitive to ethanol than the coleoptiles.

In order to clarify the effect of protein synthesis inhibition on anoxiatolerance, lettuce (Lactuca sativa L.) seedlings weresubjected to anoxic stress in the presence of cycloheximide (CHM). Atconcentrations greater than 3 , CHM decreased thesurvivability of the roots and the survivability decreased with increasing CHMdoses. At 100 CHM, the survivability was 41% of thatof non-CHM-treated seedlings. Alcohol dehydrogenase (EC 1.1.1.1) activity andATP concentration in the roots of the seedlings were also decreased by CHM,which may be one of the causes of the reduced anoxia tolerance of the seedlings.

To understand the effect of wounding stress on alcohol dehydrogenase(ADH, EC 1.1.1.1) in monocotyledonous and dicotyledonous plants, maize(Zea mays L.) and lettuce (Lactucasativa L.) seedlings were subjected to wounding stress and ADHactivity and abscisic acid (ABA) concentration were determined. In response tothe stress, the ADH activity in seedlings of both species increased rapidly asaresult of increased synthesis of the ADH. At 12 h after thestress,the activities in the wounded lettuce and maize seedlings, respectively,increased to 1.7- and 1.5-fold of that in non-stressed seedlings. Woundingstress also increased the concentration of endogenous ABA during the first 6h in both seedlings. The maximum increased levels of ABA in thelettuce and maize seedlings were 4.9- and 4.7-fold of that in the non-stressedseedlings, respectively.

Maize (Zea mays L.) seedlings were exposed to osmotic stress, and alcohol dehydrogenase (ADH) activity and abscisic acid (ABA) concentration were determined. The osmotic stress increased ADH activities in both roots and shoots, whereas the increase was 2-fold greater in roots than the shoots. The stress also increased ABA concentration in both roots and shoots and the increase was greater in the roots than in the shoots.

Alcohol dehydrogenase (ADH), its isozyme profiles and ethanol concentration in lettuce (Lactuca sativa L.) seedlings subjected to flooding stress were determined. Flooding stress caused increases in ADH activity and ethanol concentration. By 48 h, ADH activity and ethanol concentration in the flooded seedlings increased 3.2- and 7.0-fold, respectively, in comparison with those in non-stressed seedlings. Five electrophoretically separable ADH bands were found in extract of the flooded seedlings, whereas only two or three ADH bands were found in extract of non-stressed seedlings. These results indicate that lettuce ADH may have a system of three-gene and six-isozyme, and the increase in ADH activity in the flooded seedlings may be due to increased synthesis of the enzyme.

According to the Davies–Roberts hypothesis, plants primarily respond to oxygen limitation by a burst of lactate production and the resulting pH drop in the cytoplasm activates ethanolic fermentation. To evaluate this system in lettuce (Lactuca sativa L.), seedlings were subjected to anoxia and in vitro activities of alcohol dehydrogenase (ADH, EC 1.1.1.1), pyruvate decarboxylase (PDC, EC 4.1.1.1) and lactate dehydrogenase (LDH, EC 1.1.1.27) and concentrations of ethanol, acetaldehyde and lactate were determined in roots of the seedlings. The in vitro activities of ADH and PDC in the roots increase in anoxia, whereas no significant increase was measured in LDH activity. At 6 h, the ADH and PDC activities in the roots kept in anoxia were 2.8- and 2.9-fold greater than those in air, respectively. Ethanol and acetaldehyde in the roots accumulated rapidly in anoxia and increased 8- and 4-fold compared with those in air by 6 h, respectively. However, lactate concentration did not increase and an initial burst of lactate production was not found. Thus, ethanol and acetaldehyde production occurred without an increase in lactate synthesis. Treatments with antimycin A and salicylhydroxamic acid, which are respiratory inhibitors, to the lettuce seedlings in the presence of oxygen increased the concentrations of ethanol and acetaldehyde but not of lactate. These results suggest that ethanolic fermentation may be activated without preceding activation of lactate fermentation and may be not regulated by oxygen concentration directly.

Allelochemicals were isolated from the germinating maize (zea mays L.) seeds in order to know the allelopathic constituents in the plants at the early developmental stage. Three allelochemicals were found in the acetone extract obtained from the mesocotyls and coleoptiles of the 5-day-old seedlings. By spectral analysis, they were identified as 5-chloro-6-methoxy-2-benzoxazolinone (Cl-MBOA) which is a naturally occurring new benzoxazolinone, 6-methoxy-2-benzoxazolinone (MBOA) and 2,4-dihydroxy-l,4-benzoxazin-3-one (DIBOA). At concentrations greater than 0.03 mM for Cl-MBOA and DIBOA and 0.1 mM for MBOA, respectively, were inhibited the growth of the roots of cress (Lepidium sativum L.) seedlings. Activity of a new benzoxazolinone, Cl-MBOA, was greater than that of its analogue, MBOA. These results suggest that germinating maize seeds have at least three allelochemicals which may affect the growth or germination of other plant species.

In order to know the effect of anaerobiosis on alcohol fermentation in early developmental stage of alfalfa (Medicago sativa L.), 6-d-old seedlings were subjected to anaerobic stress and alcohol dehydrogenase (ADH) activity and ethanol concentration in the seedlings were determined. The anaerobic stress caused increases in ADH activity and ethanol concentration. At 48 h after onset of the stress, ADH activity and ethanol concentration in the seedlings were 4.2- and 20-fold greater than those in non-stressed seedlings, respectively. These results suggest that anaerobic stress accelerates the alcohol fermentation pathway in the alfalfa seedlings.