Evaluation of Putative Allelochemicals in Rice Root Exudates for Their Role in the Suppression of Arrowhead Root Growth

In previous studies, 15 putative allelopathic compounds detected in rice root exudates were quantified by GC/MS/MS. In this study, multiple regression analysis on these compounds determined that five selected phenolics, namely caffeic, p-hydroxybenzoic, vanillic, syringic, and p-coumaric acids, from rice exudates were best correlated with the observed allelopathic effect on arrowhead (Sagittaria montevidensis) root growth. Despite this positive association, determination of the phenolic acid dose-response curve established that the amount quantified in the exudates was much lower than the required threshold concentration for arrowhead inhibition. A similar dose-response curve resulted from a combination of all 15 quantified compounds. Significant differences between the amounts of trans-ferulic acid, abietic acid, and an indole also existed between allelopathic and non-allelopathic rice cultivars. The potential roles of these three compounds in rice allelopathy were examined by chemoassay. Overall, neither the addition of trans-ferulic acid nor 5-hydroxyindole-3-acetic acid to the phenolic mix significantly contributed to phytotoxicity, although at higher concentrations, trans-ferulic acid appeared to act antagonistically to the phytotoxic effects of the phenolic mix. The addition of abietic acid also decreased the inhibitory effect of the phenolic mix. These studies indicate that the compounds quantified are not directly responsible for the observed allelopathic response. It is possible that the amount of phenolic acids may be indirectly related to the chemicals finally responsible for the observed allelopathic effect.     

Correlation Between Phytotoxicity on Annual Ryegrass (Lolium rigidum) and Production Dynamics of Allelochemicals Within Root Exudates of an Allelopathic Wheat

An improved allelopathic correlation between phytotoxicity measured in root growth bioassay upon annual ryegrass (Lolium rigidum Gaud.) and the concentrations of a selection of dynamically produced allelochemicals quantified in the root exudates of cv. Khapli wheat (Triticum turgidum ssp. durum (Desf.) Husn.) monitored during the first 15 days of wheat seedling growth in a sterile, agar–water medium, has been established. Changes over the 15-day growth period in the quantities of five exuded benzoxazinones and seven phenolic acids were measured simultaneously using GC/MS/MS. Substantiating pure compound dose–response measurements were conducted over a range of concentrations for the putative allelochemicals within the wheat exudates. One synergism-based proposal using the monitored compounds to explain the observed low-exudate-concentration phytotoxicity was explored, but was found to be experimentally inadequate.     

Allelochemicals in wheat (Triticum aestivum L.): production and exudation of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one

An analytical technique employing gas chromatography and tandem mass spectrometry (GC/MS/MS) was employed to systematically screen fifty-eight wheat accessions for their differential production of 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) from three consecutive sources, i.e., the shoots, roots, and in the associated agar growth medium (collected as root exudates) of 17-day-old wheat seedlings. DIMBOA content differed significantly in the shoots, roots, or in the agar growth medium between accessions. DIMBOA accumulated differentially within the plant, with roots containing more DIMBOA than the shoots. Only 19% of accessions were able to exude DIMBOA from living roots into their growth medium, indicating the exudation of DIMBOA is accession-specific. DIMBOA level in root tissues is expected to be high when a high level of DIMBOA content is detected in the shoots. Wheat seedlings did not release detectable amounts of DIMBOA when the DIMBOA level was low in the root tissues. The valuable genetic material with high levels of DIMBOA in the shoots or roots identified in the present research could be used to breed for wheat cultivars with elevated allelopathic activity.     

The Role of Momilactones in Rice Allelopathy

Large field screening programs and laboratory experiments in many countries have indicated that rice is allelopathic and releases allelochemical(s) into its environment. A number of compounds, such as phenolic acids, fatty acids, phenylalkanoic acids, hydroxamic acids, terpenes, and indoles, have been identified as potential rice allelochemicals. However, the studies reviewed here demonstrate that the labdane-related diterpenoid momilactones are the most important, with momilactone B playing a particularly critical role. Rice plants secrete momilactone B from their roots into the neighboring environments over their entire life cycle at phytotoxic levels, and momilactone B seems to account for the majority of the observed rice allelopathy. In addition, genetic studies have shown that selective removal of the momilactones only from the complex mixture found in rice root exudates significantly reduces allelopathy, demonstrating that these serve as allelochemicals, the importance of which is reflected in the presence of a dedicated momilactone biosynthetic gene cluster in the rice genome.     

Why Phenolic Acids Are Unlikely Primary Allelochemicals in Rice

Allelopathy in rice (Oryza sativa, L.) effective against weeds has been found in about 3.5% of tested rice germplasm in both laboratory and field experimentation. However, the allelochemicals responsible for growth inhibition of rice-associated weeds have not yet been identified. In the literature, phenolic acids are often mentioned as putative allelochemicals. If phenolic acids commonly reach growth inhibitory concentrations in rice ecosystems, it must be expected that the degree of tolerance to phenolic acids will vary among traditional rice cultivars or plant species adapted to rice environments having inherently different phenolic acid concentrations. Phenolic acids concentrations are normally greater in submerged than in aerobic soils. A dose–response study, however, showed that seedlings of rice cultivars adapted to submerged anaerobic soils did not have higher level of tolerance against p-hydroxybenzoic acid than did seedlings of varieties adapted to aerobic upland soils. Moreover, traditional rice cultivars had no greater tolerance than did improved cultivars that were recently bred for traits other than tolerance of phenolic acids. Similarly, there were no differences in tolerance of p-hydroxybenzoic acid between two Echinochloa weed species adapted to either anaerobic or aerobic growth conditions. Thus, neither the rice cultivars nor weed species had evolved different tolerance levels against the phenolic acid. However, all rice cultivars had significantly greater tolerance of p-hydroxybenzoic acid than did either weed species. In a second experiment, the rates at which rice plants released phenolic acids into solution cultures were measured for at least one month, the time period of greatest allelopathic activity following planting under field conditions. The maximum release rate of phenolic acids during the first month of growth was approximately 10 g/plant/day. At a conventional plant density, the release rate of phenolic acids would be approximately 1 mg/m²day. This order of release rate cannot provide concentrations remotely close to phytotoxic levels determined for these rice cultivars and weed species. The results presented in this paper do not preclude the possibility that phenolic acids might be one component in a mixture of chemicals that, when present simultaneously, are allelopathic.     

Difference in hydroxamic acid content in roots and root exudates of wheat (Triticum aestivum L.) and rye (Secale cereale L.): Possible role in allelopathy

Hydroxamic acids (Hx) produced by some cereal crops have been associated with allelopathy. However, the release of Hx to the soil by the producing plant-an essential condition for a compound to be involved in allelopathy-has not been shown. GC and HPLC analysis of roots and root exudates of wheat (Triticum aestivum L.) and rye (Secale cereale L.) cultivars, with high Hx levels in their leaves, demonstrated the presence of these compounds in the roots of all cultivars analyzed and in root exudates of rye. Moreover, bioassays employing root exudates collected from wheat and rye seedlings demonstrated that only rye exudates inhibited root growth of wild oats,Avena fatua L., a weed whose root growth is inhibited by Hx. These results suggest that rye could potentially interfere with the growth ofAvena fatua in nature and that this interference could be due to the release of Hx to the soil by way of roots.     

Distribution and Exudation of Allelochemicals in Wheat Triticum aestivum

Wheat allelopathy has potential for weed suppression. Allelochemicals were identified in wheat seedlings, and they were exuded from seedlings into agar growth medium. p-Hydroxybenzoic, trans-p-coumaric, cis-p-coumaric, syringic, vanillic, trans-ferulic, and cis-ferulic acids and 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one (DIMBOA) were identified in both the shoots and roots of 17-day-old wheat seedlings and their associated agar growth medium. Wheat accessions with previously identified allelopathic activity tended to contain higher levels of allelochemicals than poorly allelopathic ones. The allelopathic compounds present in the shoots generally also were identified in the roots and in the agar medium. Allelochemicals were distributed differentially in wheat, with roots normally containing higher levels of allelochemicals than the shoots. When the eight allelochemicals were grouped into benzoic acid and cinnamic acid derivatives, DIMBOA, total coumaric, and total ferulic acids, the amount of each group of allelochemicals was correlated between the roots and the shoots. Most of the allelochemicals identified in the shoots and roots could be exuded by the living roots of wheat seedling into the agar growth medium. However, the amounts of allelochemicals in the agar growth medium were not proportional to those in the roots. Results suggest that wheat plants may retain allelochemicals once synthesized. The presence of allelochemicals in the agar growth medium demonstrated that wheat seedlings were able to synthesize and to exude phytotoxic compounds through their root system that could inhibit the root growth of annual ryegrass.     

Allelopathic effect of alfalfa (Medicago sativa) on bladygrass (Imperata cylindrica)

Greenhouse and laboratory experiments were conducted at the Agricultural and Water Resources Research Center Station, Baghdad, in 1985 and 1986 to investigate the possible allelopathic potential of alfalfa (Medicago saliva L.) and its decomposed residues on bladygrass (Imperata cylin-drica L. Beauv.), a noxious weed in Iraq, and to isolate, characterize, and quantify possible allelopathic agents in alfalfa residues and root exudates. Results indicated that decomposed alfalfa roots and their associated soil produced a 51–56% reduction in bladygrass seed germination. Root and shoot length of bladygrass seedlings were reduced by an average of 88%. Decayed and undecayed mixtures of alfalfa roots and soil at 0.0151 (w/w) inhibited bladygrass seedlings reproduced from rhizomes by 30 and 42%. It was found that root exudates of alfalfa seedlings caused significant reduction in shoot and root dry weights of bladygrass seedlings when alfalfa and bladygrass were grown together in nutrient culture. Caffeic, chlorogenic, isochloro-genic,p-coumaric,p-OH-benzoic, and ferulic acids were detected in alfalfa root exudates and residues. The highest amount (126 fig phenolic acids/g soil) of these compounds was found in alfalfa root residues after six months of decomposition in soil.     

Changes in Rice Allelopathy and Rhizosphere Microflora by Inhibiting Rice Phenylalanine Ammonia-lyase Gene Expression

Gene expression of phenylalanine ammonia-lyase (PAL) in allelopathic rice PI312777 was inhibited by RNA interference (RNAi). Transgenic rice showed lower levels of PAL gene expression and PAL activity than wild type rice (WT). The concentrations of phenolic compounds were lower in the root tissues and root exudates of transgenic rice than in those of wild type plants. When barndyardgrass (BYG) was used as the receiver plant, the allelopathic potential of transgenic rice was reduced. The sizes of the bacterial and fungal populations in rice rhizospheric soil at the 3-, 5-, and 7-leaf stages were estimated by using quantitative PCR (qPCR), which showed a decrease in both populations at all stages of leaf development analyzed. However, PI312777 had a larger microbial population than transgenic rice. In addition, in T-RFLP studies, 14 different groups of bacteria were detected in WT and only 6 were detected in transgenic rice. This indicates that there was less rhizospheric bacterial diversity associated with transgenic rice than with WT. These findings collectively suggest that PAL functions as a positive regulator of rice allelopathic potential.     

Root Distribution and Potential Interactions Between Allelopathic Rice,Sprangletop (<Emphasis Type="Italic">Leptochloa</Emphasis> spp.), and Barnyardgrass (<Emphasis Type="Italic">Echinochloa crus-galli</Emphasis>) based on <Superscript>13</Superscript>C Isotope Discrimination Analysis

Weed-suppressive rice cultivars hold promise for improved and more economical weed management in rice. Interactions between roots of rice and weeds are thought to be modulated by the weed-suppressive activity of some rice cultivars, but these phenomena are difficult to measure and not well understood. Thus, above-ground productivity, weed suppression, and root distribution of 11 rice cultivars and two weed species were evaluated in a drill-seeded, flood-irrigated system at Stuttgart, Arkansas, USA in a two-year study. The allelopathic cultivars, PI 312777 and Taichung Native 1 (TN-1), three other weed-suppressive cultivars, three indica-derived breeding selections, and three non-suppressive commercial cultivars were evaluated in field plots infested with barnyardgrass (Echinochloa crus-galli (L.) Beauv.) or bearded sprangletop (sprangletop, Leptochloa fusca (L.) Kunth var. fascicularis (Lam.) N. Snow). The allelopathic cultivars produced more tillers and suppressed both weed species to a greater extent than did the breeding selections or the non-suppressive cultivars. ¹³C isotope discrimination analysis of mixed root samples to a depth of 15 cm revealed that the allelopathic cultivars typically produced a greater fraction of their total root mass in the surface 0–5 cm of soil depth compared to the breeding selections or the non-suppressive cultivars, which tended to distribute their roots more evenly throughout the soil profile. These trends in root mass distribution were apparent at both early (pre-flood) and late-season stages in weed-free and weed-infested plots. Cultivar productivity and root distribution generally responded similarly to competition with the two weed species, but barnyardgrass reduced rice yield and root mass more than did sprangletop. These findings demonstrate for the first time that roots of the allelopathic cultivars PI 312777 and TN-1 explore the upper soil profile more thoroughly than do non-suppressive cultivars under weed-infested and weed-free conditions in flood-irrigated U.S. rice production systems. They raise the interesting prospect that root proliferation near the soil surface might enhance the weed-suppressive activity of allelochemical exudates released from roots. Plant architectural design for weed suppressive activity should take these traits into consideration along with other proven agronomic traits such as high tillering and yield.     

Evaluation of Root Exudates of Seven Sorghum Accessions

Seven sorghum accessions were evaluated quantitatively and qualitatively for the composition of their root exudates. Utilizing a unique capillary mat growing system, root exudates were collected from all sorghum accessions. Exudates were subjected to TLC and HPLC analysis to evaluate their chemical composition. Within each sorghum accession, variation existed in the amount of exudate produced and the chemical constituents of each exudate. Sorgoleone was the predominant constituent identified in each accession's exudate. Other closely related compounds, including 5-ethoxysorgoleone, 2,5-dimethoxysorgoleone, three other minor components (MW = 364, 388, and 402), and one unidentified component comprised the minor constituents of the root exudate. Our past work has shown that several of these compounds have potent phytotoxic activity as photosystem II inhibitors, thereby lending further support to the concept that Sorghum spp. are allelopathic and weed suppressive.     

Identification of Phytotoxic Substances from Early Growth of Barnyard Grass (Echinochloa crusgalli) Root Exudates

Barnyard grass is a problematic weed worldwide. It competes with crops and causes reduction in crop yields. In this study, barnyard grass suppressed rice emergence, and the degree of rice inhibition was proportional to the density of barnyard grass. Root exudates of barnyard grass reduced germination and growth of lettuce, rice, and monochoria. Fifteen compounds potentially involved in the phytotoxic activities of barnyard grass were isolated and identified, including phenolics, long-chain fatty acids, lactones, diethyl phthalate, acenaphthene, and derivatives of phthalic acids, benzoic acid, and decane. Quantities of diethyl phthalate, decanoic acid, myristic acid, stearic acid, 7,8-dihydro-5,6-dehydrokavain, and 7,8-dihydrokavain were 2.7, 11.1, 19.6, 35.5, 10.3, and 15.5 μg/ml of barnyard grass root exudates, respectively. The two lactones exhibited the greatest inhibition, followed by the phenolics and the derivatives of phthalic acids. Fatty acids had stronger suppression than diethyl phthalate and ethyl ester-4-ethoxy-benzoic acid. The acenaphthene and decane derivatives were the least phytotoxic. The phytotoxins released by barnyard grass roots showed strong inhibition on growth of broadleaf indicator plants and paddy weeds, but were less effective on barnyard grass itself and rice. Our study revealed that in addition to competition, barnyard grass also interferes with rice and other plants in its surroundings by chemical means.T. D. Khanh and I. M. Chung are members of the research team of Friendly Environmental Low Input Natural Herbicide New Material Study of the Konkuk University.     

Change in acceptability of barley plants to aphids after exposure to allelochemicals from couch-grass (Elytrigia repens)

The response of the bird cherry-oat aphid, Rhopalosiphum padi, to barley plants was investigated following exposure of the plants to root allelochemicals from the aggressive weed couch-grass, Elytrigia (Agropyron) repens. Plants were treated either with root exudates from living couch-grass plants or with previously identified couch-grass root compounds [5-hydroxyindole-3-acetic acid, DL-5-hydroxytryptophan, L-5-hydroxytryptophan hydrate, and 6-hydroxy-1,2,3,4-tetrahydro--carboline-3-carboxylic acid (carboline)] either separately or in mixtures. In choice and no-choice settling tests, aphid acceptance of barley plants was significantly reduced following treatment with root exudates, and the carboline when tested alone or in combination with the other compounds. In contrast, the other compounds without the carboline were less active in reducing aphid acceptance. In a probing bioassay, individual substances were either neutral or stimulatory to aphids, indicating that the reduced settling was probably not due to direct effects on aphids, but rather due to effects on the plant. This was confirmed in olfactometer assays, in which aphids were repelled by odors from barley plants following treatment with a mixture containing all four chemicals.     

Effects of Temperature and Photoperiod on Phytotoxic Root Exudates of Cucumber (Cucumis sativus) in Hydroponic Culture

In order to elucidate the effects of temperature and photoperiod on the quality and quantity of plant root exudates, a Japanese cucumber (Cucumis sativus, cv. Shougoin-Aonaga-Fushinari) was grown hydroponically in growth chambers under controlled temperature and photoperiod conditions with or without the addition of activated charcoal (AC) to the nutrient solutions. Fresh AC was used to trap the organic compounds exuded from cucumber roots every two weeks. Cucumber plants without AC were severely retarded in root growth and in the accumulation of dry matter, especially at high temperature and long photoperiod, compared to those with AC. The growth inhibitors, adsorbed on the AC or accumulated in the nutrient solution without AC, were extracted by organic solvents and analyzed by GC-MS. Benzoic acid and its derivatives, cinnamic acid derivatives, and fatty acids were identified. The rate of root exudation in vegetative and reproductive stages for some of these organic acids increased with the elevation of temperature and the elongation of photoperiod, and the mean rate was two or more times higher than the minimum exudation at low temperature with short photoperiod. Some of the identified compounds significantly inhibited the germination and/or root growth of lettuce and cucumber.     

新鲜椰子果肉脂肪酸成分的GC-MS分析

以海南产新鲜高种椰为原料,参照国家标准GB/T 14772-93和GB5009.6-85,分别采用索氏抽提法和酸水解法提取新鲜椰肉脂肪酸成分,再经过氢氧化钾碱催化法进行脂肪酸的甲酯化处理后,通过气相色谱/质谱联用技术分析测定脂肪酸的组成。共检出9种脂肪酸成分,主要成分为以月桂酸为主的C6至C18系列饱和脂肪酸。     

Differences Among Rice Cultivars in Root Exudation, Methane Oxidation, and Populations of Methanogenic and Methanotrophic Bacteria in Relation to Methane Emission

Greenhouse experiments were conducted under subtropical conditions to understand the mechanism of rice cultivar differences in methane (CH₄) emission. Three rice cultivars were studied. Differences in CH₄ emission rates among the three rice cultivars became evident in the middle and late growth stages. Rice root exudates per plant measured as total released C were significantly different among rice cultivars. The effect of root exudates on CH₄ production in soil slurry differed accordingly. The amount of root exudates was not significantly different among rice cultivars when computed on a dry matter basis, indicating that it is positively correlated to root dry matter production. The root CH₄-oxidizing activity differed among rice cultivars. IR65598 had a higher oxidative activity than IR72 and Chiyonishiki. Root air space was not significantly different among rice cultivars at the late growth stage, indicating that it is probably not a factor contributing to cultivar differences in CH₄ emission. The population level of methanogenic bacteria differed significantly in soil grown to different rice cultivars, but not in roots, at booting stage and ripening stage. Methanotrophic bacteria population differed significantly in roots among rice cultivars at ripening. Rice cultivars with few unproductive tillers, small root system, high root oxidative activity, and high harvest index are ideal for mitigating CH₄ emission in rice fields.     

Composition of phenolic acids in sea buckthorn (<Emphasis Type="Italic">Hippophae rhamnoides</Emphasis> L.) berries

The composition of phenolic acids in several varieties of sea buckthorn berries was determined by GC and MS. In six cultivars the total content of phenolic acids ranged from 3570±282 to 4439±405 mg per kg of berries, on a dry basis. Seventeen phenolic acids were tentatively identified in the berries. Salicylic acid was the principal phenolic acid in sea buckthorn berries, accounting for 55 to 74.3% of the total phenolic acids present. The phenolic acids liberated from esters and glycosidic bonds were the major fractions of phenolic acids in the berries, whereas free phenolic acids constituted only up to 2.3% of total phenolic acids present.     

Allelopathic effects ofChrysanthemum morifolium on germination and growth of several herbaceous plants

Aqueous extracts obtained from young green tops ofChrysanthemum morifolium inhibited the germination of six flowering plants, including chrysanthemum itself, provided for experiments. The same phenomenon was also clearly observed when powder made from young green tops and old leaves of chrysanthemum was used. Moreover, the growth of seedlings planted again in garden soil which was once used for the culture of chrysanthemum was greatly interrupted. Chrysanthemum cultured in used garden soil showed far less dry weight than that cultured in fresh garden soil. The weight of chrysanthemum cultured using its root exudates was also less than that cultured with water leachate of fresh garden soil, and therefore these results may be considered to indicate allelopathic effects. In order to find the allelochemicals related to this phenomenon, benzoic acid and phenolic acids such as salicylic,p-hydroxybenzoic, vanillic, gentisic, protocatechuic, syringic, gallic, ferulic, and caffeic acids were identified by gas chromatography.     

Biomimetic measurement of allelochemical dynamics in the rhizosphere

Polydimethylsiloxane (PDMS) materials were used to quantify levels of the photosynthesis inhibitor sorgoleone in the undisturbed rhizosphere of sorghum plants. The materials used included stir bars coated with PDMS (stir bar sorptive extraction), technical grade optical fiber coated with a thin film of PDMS (matrix-solid phase microextraction), and PDMS tubing. PDMS tubing retained the most sorgoleone. As analyzed by high performance liquid chromatography, amounts of sorgoleone retained on the PDMS materials increased with time. Other materials tested (polyurethane foam plugs, C18 and Tenax disks, and resin capsules) proved less suitable, as they were subject to sometimes extensive penetration by fine root hairs. These results demonstrate the potential for PDMS-based materials to monitor the release of allelochemicals in the undisturbed rhizosphere of allelopathic plants. Unlike extraction procedures that recover all available compounds present in the soil, PDMS functions in a manner more analogous to plant roots in sorbing compounds from soil solution or root exudates. Information on chemical dynamics in the rhizosphere is crucial for evaluating specific hypotheses of allelopathic effects, understanding allelopathic mechanisms, and assessing the importance of allelopathic processes in plant communities.     

Defoliation Affects Chemical Defenses in All Plant Parts of Rye Seedlings

The effect of defoliation and consecutive defoliation (condefoliation) of rye seedlings on the allocation patterns of biomass and hydroxamic acids (Hx) was evaluated five days after treatment. Growth of condefoliated seedlings was lower than that of defoliated and nondefoliated ones. Concentration of Hx decreased in shoots of condefoliated seedlings compared to nondefoliated ones, while concentration of Hx in root exudates increased. Allocation of Hx to roots and root exudates increased at the expense of allocation to the shoots in condefoliated seedlings. The ratio of Hx-aglucone to Hx-glucoside was higher in shoots of defoliated and condefoliated seedlings. The decrease in quantity of defense in shoots was accompanied by an increase in its quality, given that aglucones are more toxic than glucosides. The increase in concentration of Hx—an allelopathic compound also involved in mineral uptake by roots—in root exudates of condefoliated seedlings was suggested to lead to an advantage in the acquisition of resources for the regeneration of lost biomass.