Volatile allelopathy in velvet bean (Mucuna pruriens [correction of pruiens] L.) and gravity.

Allelopathy caused by volatile compounds was studied with velvet bean (Mucuna pruriens [correction of prueiens] L.). Preparatory experiment was conducted to identify the relationship between action of volatiles and gravity. The experimental protocol was defined and tested to study on volatile allelopathy of velvet bean. Root growth of lettuce (Lactuca sativa L.) seedlings was inhibited by volatile gas from velvet bean seedlings planted in Agripot. In this study, it was shown that ten-day velvet bean seedling could exhibit the volatile allelopathic ability. Growth chambers for volatile experiment were designed. Velvet bean seed can germinate in a 3-ml small vial.     

Plant growth promotion and suppression of charcoal‐rot fungus (Macrophomina phaseolina) in velvet bean (Mucuna pruriens L.) by root nodule bacteria

Root‐nodulating bacteria are intimate associates of legumes. From a pool of rhizobia isolated from root nodules of Mucuna pruriens (Velvet bean/Kaunch), RMP66 and BMP17 were found to be capable of promoting siderophore and IAA production and phosphate solubilization (insoluble tri‐calcium). Both symbionts were studied further to determine their abilities to promote plant growth and to control root‐rot in Mucuna pruriens caused by the pathogenic plant fungus Macrophomina phaseolina. RMP66 and BMP17 were selected based on their excellent inhibitory activities against M. phaseolina (by 78% and 71%, respectively) in dual culture and in agar‐well assays using cell‐free culture filtrate (CFCF) (by 76% and 62%, respectively). Both strains inhibited fungal growth to a greater extent in iron‐deficient medium (51% and 69%) than in iron‐supplemented medium (37% and 0%), respectively. CFCFs of RMP66 and BMP17 obtained from Pikovskaya's broth and tryptophan‐amended YEM broth inhibited fungal growth by 80%‐55% and 70%‐43%, respectively, and were identified as Sinorhizobium meliloti RMP66 and Bradyrhizobium diazoefficiens BMP17 by 16S rDNA sequencing. Centrifuged and pelleted cells harvested from exponentially grown cultures of S. meliloti RMP66 and B. diazoefficiens BMP17 were used to bacterize seeds of M. pruriens, which then showed enhanced seed germination (by up to 17% and 12%, respectively), and subsequent increases in other plant growth parameters in field trials. Considerable increases in seedling vigour indices (62%: 53% and 110%: 130%) and biomass (8%: 13% and 25%: 28%) were also observed for bacterial treatments. Tn5‐mediated antibiotic‐resistant marker strains showed enhanced nodule occupancy by up to 72% and 68%, respectively. This study describes a multifunctional legume nodule rhizobia that could be utilized in multicropping systems under different agroclimatic conditions as a bioinoculant and alternative to fertilizers.     

Biotic elicitation mediated in vitro production of L-DOPA from Mucuna pruriens (L.) DC. cell cultures

In Vitro Cellular & Developmental Biology - Plant - With the emerging rise in the need for drugs extracted from various plant sources, there also arises the need for the optimum production of...     

Production of L-DOPA from cell suspension culture of Mucuna pruriens f. pruriens

Summary Production of L-DOPA was studied in cell suspension culture of Mucuna pruriens f. pruriens. Suspension culture was established in MSI medium composed of half concentration of Murashige and Skoog's salts and 2% sucrose. A two-stage cell suspension culture was developed for enhanced accumulation of L-DOPA. In the first stage, the culture system was composed of MSI medium without CaCl, which was suitable for cell growth and in the second stage MSI medium containing 42.5 mg.l^–1 KH₂PO₄ and 4% sucrose favoured L-DOPA production. A discernible higher production of L-DOPA was obtained in this two-stage cell suspension culture in comparison to single stage culture.     

An efficient plant regeneration system for Mucuna pruriens L. (DC.) using cotyledonary node explants

Summary The purpose of this study was to develop an efficient micropropagation system for Mucuna pruriens, an important medicinal plant in India. A range of cytokinins was investigated for multiple shoot regeneration with cotyledonary node explants from 7-d-old aseptic seedlings. Of all the cytokinins, 6-benzyladenine (BA), kinetin (KIN) and 2-isopentenyl adenine (2-iP) tested in Murashige and Skoog medium (MS), BA was the most effective and 5.0 μM was found to be optimum for inducing maximum shoots. Medium types, medium strength and pH were also investigated for induction and proliferation of shoots. The highest efficiency of shoot proliferation was observed in 5.0 μM BA and 0.5 μM α-naphthalene acetic acid (NAA) in half-strength MS medium at pH 5.8. The best condition for rooting was half-strength MS medium solidified with agar and with 2.0 μM indole-3-butyric acid (IBA). After rooting, the plantlets were transferred to plastic pots filled with sterile soilrite where 90% grew and all exhibited normal development.     

Reduced allelopathic inhibition of lettuce (Lactuca sativa) growth caused by velvet bean (Mucuna pruriens) under 3D-clinorotation.

Allelopathy between Mucuna pruriens (velvet bean) and Lactuca sativa (lettuce) was studied under 3D-clinorotation. Growth of both roots and shoots of lettuce seedlings was suppressed by the presence of velvet bean. The degree of suppression was less on the clinostat compared to the normal static earth gravity. L-DOPA (L-3, 4-dihydroxyphenylalanine) is known to be a major substance in allelopathy of velvet bean. Amount of L-DOPA diffused out from a sintered filter paper into agar medium was compared between clinorotation and control group, and found no significant difference. It was concluded that some factors related to release, transport, and sensing phenomena of allelopathic substances may be responsible to the new findings in this study.     

Tolerance to acid soil conditions of the velvet beans Mucuna pruriens var. utilis and M. deeringiana

Velvet beans, fast growing leguminous cover crops used in the humid tropics, are shallow rooted on acid soils. This might be due to an inherent branching pattern, to an intrinsic toxicity of the acid subsoil or to a relative preference for root development in the topsoil. Such preference could be based on soil chemical factors in the subsoil or on physical factors such as penetration resistance or aeration. In a field experiment with two species of velvet bean (Mucuna pruriens var. utilis and M. deeringiana) all topsoil was removed and plants were sown directly into the acid subsoil. Root development was neither affected by this treatment nor by P fertilization or liming. In the absence of topsoil good root development in the exposed upper layer of subsoil was possible, so the hypothesis of a toxicity per se of the subsoil could be rejected. To test whether poor root development in the subsoil in the presence of topsoil is due to an inherent branching pattern of the plant or to a relative preference for topsoil, a modified in-growth core technique was used. Local topsoil and subsoil and an acid soil with a higher exchangeable Al content were placed in mesh bags at different depths and at different bulk densities, with and without lime and/or P fertilizer. A comparison of root development in mesh bags placed in the topsoil or subsoil showed that position and thus inherent branching pattern is not important. Root development in the subsoil was poor when this soil was placed in a mesh bag in the topsoil, but in an acid soil of much higher exchangeable Al content and higher percentage Al saturation more roots developed. In a second experiment in mesh bags, bulk density of the repacked soil in the range 1.0–1.5 g cm^-3 had no significant effect on root development. P fertilization and a high rate of liming of the soil placed in the mesh bag had a positive effect on root length density. It is concluded that poor root development in the acid subsoil under field conditions is due to a relative preference for topsoil. Al saturation and bulk density of the soil are not directly involved in this preference, but differences in availability of P and Mg or in Ca/Al ratios might play a role.     

Tolerance to acid soil conditions of the velvet beans Mucuna pruriens var. utilis and M. deeringiana

Fast growing, climbing leguminous cover crops such as the velvet beans can be used to reclaim weed-infested, degraded soils in the humid tropics, especially land covered by the grass Imperata cylindrica; they climb over the grass leaves and shade the grass out if their cover lasts long enough. Tolerance of two species of velvet bean to eroded soils was investigated by removing topsoil and directly sowing into the subsoil; plots where topsoil was not removed were used as a control. The response to small amounts of P fertilizer and lime was also tested. Removal of the topsoil resulted in retarded growth of both species, in increased dry matter content of the shoot, in decreased specific leaf area and in increased leaf weight ratio, due to shorter internodes. Six weeks after planting the leaf area index (LAI) was about 1.2 where topsoil was retained, sufficient for a shading effect on Imperata. Where topsoil had been removed, the LAI was only 0.6. Mucuna pruriens var. utilis showed a faster aboveground growth than M. deeringiana; the species did not differ in tolerance to eroded soil. Small amounts of P fertilizer had no significant effect on the growth of both Mucuna species. Shoot: root ratios, on a dry weight basis, were much lower when topsoil had been removed, about 3.7 and 2.4 for M. p. utilis and M. deeringiana respectively, compared to 6.2 and 3.3 where topsoil was retained. Removal of topsoil led to reduced Mg and to increased Al concentrations in roots, and to increased levels of Mn and Al in shoots. In the second year no effect of lime or residual effect of P application was found on growth of Mucuna or Imperata. Removal of the topsoil had little effect on the growth of weeds after the cover crop had been harvested. Due to the high Al tolerance of Imperata, reclamation by Mucuna will be less effective if the topsoil has been lost by erosion.     

Ultrastructural and biochemical investigations of protein mobilization of Mucuna pruriens (L.) DC. cotyledons and embryo axis

The mobilization of storage reserves, with particular emphasis on storage proteins of Mucuna pruriens (L.) DC., cotyledons, and embryo was investigated from the ultrastructural and biochemical points of view. Proteins and starch were the two main storage substances in cotyledons, and proteins and lipids were the main ones in the embryo. Embryo protein bodies were smaller and fewer in number than those of cotyledons. Structural and ultrastructural data determined between 24 and 48 h after imbibition and between 48 and 72 h after imbibition, the end of significant embryo and cotyledon protein mobilization, respectively, indicating more precocious storage protein mobilization in the axis than cotyledons. Moreover, storage protein mobilization in embryo and cotyledons occurred before the end of germination. Water soluble proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis, producing 29 bands with molecular weights from 14 to 90 KDa. Embryo extract contained more proteins than cotyledon extract, contained seven characteristic bands, and showed a higher variability of the optical density trend than cotyledon.     

Induction of phenoloxidase in cell suspension cultures of Mucuna pruriens L.

The effects of limitating nitrogen-containing compounds in the medium and of adding the amino-acid analogues p-fluorophenylalanine and ethionine on both phenoloxidase activity and the accumulation of L-3,4-dihydroxyphenylalanine (L-DOPA) are reported for cell suspension cultures of Mucuna pruriens. Nitrogen limitation of the cultures, or the addition of p-fluorophenylalanine or ethionine to the culture medium resulted in an increased phenoloxidase activity. There appeared to be an inverse relationship between phenoloxidase activity and the acccumulation of L-tyrosine into L-DOPA by alginate-entrapped cells occurred at a higher rate when phenoloxidase activity was increased.Abbreviations pFPA p-fluorophenylalanine - L-DOPA L-3,4-dihydroxyphenylalanine     

Isolation and characterization of a Forssman antigen-binding lectin from velvet bean (Mucuna derringiana) seeds

A Forssman antigen (GalNAc1-3GalNAc1-3Gal1-4Gal1-4Glc1-1Cer)-binding lectin has been purified from velvet bean (Mucuna derringiana) seeds by a combination of affinity chromatography and reversed phase HPLC. This lectin agglutinates both native and trypsin-treated sheep erythrocytes as well as trypsinized rabbit erythrocytes, but neither native rabbit nor human erythrocytes, irrespective of blood group type. SDS-PAGE and gel filtration chromatography reveal the lectin to be a homodimer consisting of two 54 kDa subunits linked by non-covalent bonds. The results obtained by quantitative precipitation, haemagglutination inhibition and TLC overlay assays indicate that theMucuna lectin specifically recognizes Forssman antigen and Forssman disaccharide (GalNAc1-3GalNAc)-related structures. Abbreviations: The abbreviations and the trivial names used are: AH, 6-aminohexyl; BSA, bovine serum albumin; Cer, ceramide; HPLC, high performance liquid chromatography; PAGE, polyacrylamide gel electrophoresis; PBS, 10mm phosphate-buffered saline, pH 7,2, containing 0.15m NaCl; PMSF, phenyl methyl sulfonyl fluoride; SDS, sodium dodecyl sulphate; TFA, trifluoroacetic acid; TBS, 20mm tris-buffered saline, pH 7.2; TLC, thin-layer chromatography; A disaccharide, GalNAc1-3Gal; A trisaccharide, GalNAc1-3[Fuc1-2]Gal; Forssman disaccharide, GalNAc1-3GalNAc; CDH (ceramide dihexoside or lactosyl ceramide) Gal1-4Glc1-1Cer (LacCer); CTH (ceramide trihexoside or globotriosyl ceramide), Gal1-4Gal1-4Glc1-1Cer (GbOse₃Cer or Gb₃); globoside (globotetraosyl ceramide), GalNAc1-3Gal1-4Gal1-4Glc1-1Cer (GbOse₄Cer or Gb₄); Forssman antigen (globopentaosyl ceramide), GalNAc1-3GalNAc1-3Gal1-4Gal1-4Glc1-1Cer (GbOse₅Cer).     

The effect of some environmental factors on the production of L-DOPA by alginate-entrapped cells of Mucuna pruriens

In-vitro-grown cells of Mucuna pruriens, immobilized in calcium-alginate gels, were able to transform the precursor L-tyrosine into L-dihydroxyphenylalanine (L-DOPA). After the immobilization in alginate the plant cells released 90% of the produced L-DOPA into the medium; supplementation of the medium with calcium inhibited both the transformation of L-tyrosine into L-DOPA and the release of L-DOPA into the medium. Continuous illumination of the beads had a slight beneficial effect on the synthesis of L-DOPA. A simple production medium for the transformation of L-tyrosine into L-DOPA was designed. This medium contained only sucrose and sodium chloride as osmotic stabilizers, a low concentration of calcium chloride for stabilization of the alginate beads, and L-tyrosine as the precursor.     

Stimulatory effect of copper and zinc sulphate on plant regeneration,glutathione-S-transferase analysis and assessment of antioxidant activities in Mucuna pruriens L. (DC)

Plant Cell, Tissue and Organ Culture (PCTOC) - The effect of heavy metals, CuSO4 and ZnSO4 on morphogenic response of cotyledonary node explants of Mucuna pruriens (L.), a fast growing tropical...     

Physiological and genetic analysis of root responsiveness to auxin-producing plant growth-promoting bacteria in common bean (Phaseolus vulgaris L.)

Plant root development can be largely affected through the association of roots with plant growth-promoting rhizobacteria (PGPR). However, little is known about the identity of plant genes enabling such PGPR-plant root associations. Differences in the responsiveness to PGPR among cultivars suggest genetic variation for this trait within germplasm. In this study, two genotypes of common bean (Phaseolus vulgaris L.), BAT477 and DOR364, were identified showing contrasting responsiveness in root development to inoculation with the PGPR Azospirillum brasilense Sp245. Inoculation with an A. brasilense Sp245 mutant strain strongly reduced in auxin biosynthesis or addition of increasing concentrations of exogenous auxin to the plant growth medium, indicated that the differential response to A. brasilense Sp245 among the bean genotypes is related to a differential response to the bacterial produced auxin. To further assess the role of the plant host in root responsiveness, a population of Recombinant Inbred Lines (RILs) of the DOR364×BAT477 cross was used to evaluate the efficacy of exogenous auxin on root development. We detected significant phenotypic variation among the RILs for basal root formation during germination upon addition of auxin to the growth medium. Genetic analysis revealed two quantitative trait loci (QTLs) associated with basal root responsiveness to auxin of which one explained 36% of the phenotypic variation among the RILs. This latter QTL mapped to the same location as a QTL for root tip formation at low P, suggesting that the host effect on root responsiveness to IAA interacts with specific root development. Also, significant correlations between basal root responsiveness to auxin and growth, root tips and root dry weight at low P were identified. To our knowledge, this is the first report on QTL detection for root responsiveness to auxin.     

Symbionts in Mucuna pruriens stimulate plant performance through nitrogen fixation and improved phosphorus acquisition

刺毛黎豆共生体通过固氮和提高磷吸收来提高植株的生长表现南非主要以热带稀树草原和草原生态系统为主,这些生态系统具有酸性和营养缺乏的属性,特别是磷和氮素的缺乏。刺毛黎豆(Mucuna pruriens)是在大多数非洲国家广泛存在的一种本土豆科植物,该 植物可以抵御这些不利的土壤条件。豆科植物在世界上许多国家具有药用价值,也被用于土壤施肥。尽管已有文献记载了在营养胁迫生态系统中刺毛黎豆的生长和建植,但尚未对其在磷缺乏条件下的共生互作、氮源偏好和与生长相关的碳成本进行研究。在本研究中,我们确定了微生物共生体对磷缺乏条件下的刺毛黎豆氮营养和生长相关的碳成本的影响。我们从4个不同的地理位置采集微生物接种土壤,然后将种子在这些天然土壤中发芽,在根瘤发育的早期,将幼苗转移到无菌石英砂中,并提供不同磷 浓度的营养培养基。16S RNA序列结果显示,在不考虑磷浓度的情况下,Burkholderia sp., Paenibacillus sp.和Bacillus 均有分枝。尽管磷缺乏导致总生物量/生长下降,但根系生物量、根瘤数量和碳成本增加。低磷供给的幼树丛枝菌根真菌根系定殖率最高。刺毛黎豆中,大气中的氮与磷水平呈正相关关系,幼树对大气氮素和土壤氮素具有双重依赖,低磷植株对土壤氮素的依赖程度增加。因此,在磷缺乏的情况下,刺毛黎豆表现出不同的形态和微生物共生关系。     

Comparative cytogenetic mapping between the lima bean (Phaseolus lunatus L.) and the common bean (P. vulgaris L.)

The common bean (Phaseolus vulgaris) and lima bean (P. lunatus) are among the most important legumes in terms of direct human consumption. The present work establishes a comparative cytogenetic map of P. lunatus, using previously mapped markers from P. vulgaris, in association with analyses of heterochromatin distribution using the fluorochromes chromomycin A3 (CMA) and 4′,6-diamidino-2-phenylindole (DAPI) and localization of the 5S and 45S ribosomal DNA (rDNA) probes. Seven BACs selected from different common bean chromosomes demonstrated a repetitive pericentromeric pattern corresponding to the heterochromatic regions revealed by CMA/DAPI and could not be mapped. The subtelomeric repetitive pattern observed for BAC 63H6 in most of the chromosome ends of common bean was not detected in lima bean, indicating lack of conservation of this subtelomeric repeat. All chromosomes could be identified and 16 single-copy clones were mapped. These results showed a significant conservation of synteny between species, although change in centromere position suggested the occurrence of pericentric inversions on chromosomes 2, 9 and 10. The low number of structural rearrangements reflects the karyotypic stability of the genus.     

Effect of temperature on aflatoxin production in Mucuna pruriens seeds.

This paper describes the effect of temperature on the level of aflatoxin production in Mucuna pruriens seeds. The highest level of aflatoxin B1 (1.75 micrograms/g) was detected in the samples incubated at 25 degrees C for three weeks. At 20, 30, and 35 degrees C, aflatoxin levels were 0.30 to 0.56, 0.37 to 1.20, and 0.26 to 0.65 micrograms/g, respectively. The lowest concentration of aflatoxin B1 (0.10 to 0.29 microgram/g) was produced at 15 degrees C.     

Allelopathy in the natural and agricultural ecosystems and isolation of potent allelochemicals from Velvet bean (Mucuna pruriens) and Hairy vetch (Vicia villosa).

We have studied on allelopathy of plants and developed methods to identify the effective substances in root exudates, leaf leacheate, and volatile chemicals emitted from plants. We found traditional cover plants that show allelopathic activity are useful for weed control. It could eliminate the use of synthetic chemicals for this purpose. Allelopathy is a natural power of plants to protect themselves by producing natural organic chemicals. Some endemic plants in Asia, already known by farmers in the region, as either cover crops used in intercropping, hedgerow, or agroforestry, were found to possess strong allelopathic abilities. Our group identified several allelochemicals from these plants. These allelopathic cover crops, mostly leguminous plants, provide protein rich food, and grow easily without artificial fertilizers, herbicides, insecticides and fungicides. In this regards, these allelopathic cover crops could save food shortage in rural area, and are useful for environmental conservation. Screenings of allelopathic plants by specific bioassays and field tests have been conducted. Hairy vetch (Vicia villosa) and Velvet bean (Mucuna pruriens) are two promising species for the practical application of allelopathy. An amino acid, L-DOPA, unusual in plants, plays an important role as allelochemical in Velvet bean (Mucuna pruriens). Hairy vetch is the most promising cover plant for the weed control in orchard, vegetable and rice production and even for landscape amendment in abandoned field in Japan. We have isolated "cyanamide", a well known nitrogen fertilizer, from Hairy vetch. This is the first finding of naturally produced cyanamide in the world.     

Nutritional characterization of Mucuna pruriens: 4. Does replacing soybean meal with Mucuna pruriens in lamb diets affect ruminal,blood and tissue l-dopa concentrations?

The harmful effects of the 3,4-dihydroxy-l-phenylalanine (l-dopa) in Mucuna pruriens (Velvet bean) seeds have limited its use as a protein supplement for monogastrics and humans. Little is known about the extent of metabolism of Mucuna l-dopa in ruminants or its accumulation in ruminant tissues consumed as food by humans. This study aimed to determine if replacing soybean meal (SB) with Mucuna increases concentrations of l-dopa in rumen fluid, blood and muscle tissue of lambs. Twenty-seven RM lambs (RM; initial body weight, BW = 33.8 ± 5.44 kg) and 12 Florida Native (FN; initial BW = 24.9 ± 8.63 kg) lambs were assigned to four treatments and fed a basal diet of coastal bermudagrass hay, corn grain, and liquid molasses for 42 (FN) or 49 days (RM) and then slaughtered. Dietary supplements were formulated by substituting 0 (SB), 330 (Lo), 670 (Med) or at least 1000 (Hi) g/kg of SB with rolled Mucuna seeds (l-dopa, 24 g/kg dry matter, DM). Body weight was measured weekly and carcass characteristics and concentrations of l-dopa in rumen fluid, blood and the sterno-mandibularis muscle were measured at slaughter when concentrations of blood urea N, glucose, haptoglobin and cerruloplasmin were also measured. Lambs fed SB had higher (P<0.05) average daily gain than those fed Mucuna (0.20 versus 0.15 kg/day for RM; 0.21 versus 0.14 kg/day for FN). However, concentrate protein source did not affect dressing and concentrations of blood urea N, or blood glucose. Feeding the Hi diet versus the SB diet did not increase concentrations of blood cerruloplasmin, or l-dopa or concentrations of l-dopa metabolites in blood. No l-dopa was found in the ruminal fluid of the lambs and l-dopa concentrations in the sterno-mandibularis muscle were low (i.e., <5 ng l-dopa/g), and unaffected (P>0.05) by diet. Ingested Mucuna l-dopa was extensively metabolized in lambs, and did not accumulate to toxic levels in muscle tissues.