Effects of bilberry (Vaccinium myrtillus L.) litter on seed germination and early seedling growth of four boreal tree species

Laboratory and greenhouse bioassays were used to test for inhibitory effects of senescent and decomposed leaves and aqueous extract from bilberry (Vaccinium myrtillus L.) against seed germination and seedling growth of aspen (Populus tremula L.), birch (Betula pendula Roth.), Scots pine (Pinus sylvestris L.), and Norway spruce [Picea abies (L.) Karst.]. Aqueous extracts from bilberry leaves were inhibitory to aspen seed germination and seedling growth and also induced root damage and growth abnormalities. Addition of activated carbon removed the inhibitory effects of extracts. Senescent leaves reduced pine and spruce seed germination, but rinsing of seeds reversed this inhibition. Senescent leaves were more inhibitory than decomposed leaf litter, suggesting that the inhibitory compounds in bilberry leaves are relatively soluble and released at early stages during decomposition. Spruce was generally less negatively affected by litter and aqueous extracts than the other tested species. This study indicates that chemical effects of bilberry litter have the potential to inhibit tree seedling recruitment, but these effects were not consistently strong. Phytotoxicity is unlikely to be of critical importance in determining success for spruce seedling establishment.     

Potential allelochemicals fromRuta graveolens L. and their action on radish seeds

An aqueous extract ofRuta graveolens L. (250 g/liter) was tested for its allelopathic activity in vitro on radish germination and radicle growth in light and darkness. It caused a delay in the onset and a decrease in the rate of germination (40%) in the light. The photoinhibition of germination was accompanied by an inhibition of water uptake into the seed. Furthermore, the inhibition of radicle growth was slightly higher in the light than in darkness. Three potential allelochemicals, biologically active in the light, were isolated from the extract: 5-methoxypsoralen (5-MOP), 8-methoxypsoralen (8-MOP), and 4-hydroxy-coumarin at concentrations of 10^–4 M, 2×10^–4 M, and 0.4 ×10^–5 M respectively. At a concentration of 2×10^–4 M, 5-MOP was the most potent inhibitor, decreasing radish germination to 32% and radicle growth to 17% with respect to control. Microscopic observations of radish seeds treated with 5-MOP suggest that this substance changes the swelling of the seed coat and aleurone layer, which precedes radicle protrusion.     

Phytotoxicity of <Emphasis Type="Italic">Phytolacca americana</Emphasis> Leaf Extracts on the Growth,and Physiological Response of <Emphasis Type="Italic">Cassia mimosoides</Emphasis>

We examined the allelochemical effects of control soil, native soil (treated soil), and leaf extracts of Phytolacca americana (pokeweed) on the germination rate and seedling growth of Cassia mimosoides var. nomame. We also studied the resulting changes in root-tip ultrastructure and peroxidase isozyme biochemistry. P. americana leaf extract inhibited seed germination, seedling growth, and biomass when compared to control and treated soil. Root and shoot growth in treated soil was stimulated relative to control soil, but root growth was inhibited by 50% in the leaf extract treatment. Biomass of C. mimosoides seedlings grown on leaf extract was reduced sevenfold when compared to the control seedlings. The amounts of total phenolic compounds in the leaf extract, treated soil, and control soil were 0.77, 0.14, and 0.03 mg l⁻¹, respectively. The root tips of C. mimsoides treated with leaf extracts of P. americana showed amyloplasts and large central vacuoles with electron-dense deposits inside them when compared to control root tips. The activity of guaiacol peroxidase (GuPOX) in whole plant, roots, and shoots of C. mimosoides increased as leaf extract increased; maximum activity was observed in extract concentrations of 75% and higher. Root GuPOX activity was three times higher than in shoots. Therefore, we conclude that inhibition of C. mimosoides growth is related to the phenolic compounds in the P. americana leaf extract and the ultrastructure changes in root-tip cells and increased GuPOX activity is a response to these allelochemicals.     

Synergistic inhibitory effects of vanillic andp-hydroxybenzoic acids on radish and grain sorghum

Radish and grain sorghum germination and sorghum growth were inhibited in a synergistic manner by combinations of vanillic andp-hydroxybenzoic acids. At threshold inhibition levels, 2.5 × 10^–3 M vanillic acid-treated radish seeds had 71 % of control germination after 24 hr and 2.5 × 10^–3 Mp-hydroxybenzoic acid-treated radish yielded 95% germination. A mixture of 2.5 × 10^–3 M of each of these two phytotoxins showed 52% germination after 24 hr. Equimolar mixtures of 5 × 10^–3 M vanillic andp-hydroxybenzoic acids allowed sorghum germination of 60% of untreated seeds after 24 hr, whereas separate treatments of individual phenols had 93% and 96% of control seed germination. Sorghum root and shoot elongation and total seedling growth were more sensitive than germination to vanillic andp-hydroxybenzoic acid treatments, and synergistic effects also were apparent. A combination of 5 × 10^–3 M vanillic with 5 × 10^–3 Mp-hydroxybenzoic reduced root length more than either did individually, and a mixture of 5 × 10^–4 M vanillic with 5 × 10^–4 Mp-hydroxybenzoic acid reduced sorghum seedling growth to approximately that resulting from a 10^–3 M concentration of either phenol alone. Phytotoxin levels inhibitory to sorghum growth caused small increases in lower leaf surface diffusive resistance, but did not close stomates, and this effect was not judged to be the cause of reduced sorghum growth.     

Allelopathy of oats. I. Assessment of allelopathic potential of extract of oat shoots and identification of an allelochemical

The allelopathic potential of oat (Avena sativa L.) extracts was investigated under laboratory conditions. The ethyl ether-, acetone-, and water-soluble fractions obtained from the extract of oat shoots inhibited the germination and growth of roots and hypocotyls of lettuce (Lactuca sativa L.). The inhibitory activity of the water-soluble fraction was maximum, followed by that of ethyl ether-soluble and acetone-soluble fraction. An active principle of the water-soluble fraction was isolated and its structure was determined by spectral data asl-tryptophan.l-Tryptophan inhibited the growth of hypocotyls and roots of lettuce seedlings at concentrations greater than 0.03 and 0.1 mM, respectively. These results suggested thatl-tryptophan may be an allelochemical which affects the growth or germination of different plant species.     

Methyl jasmonate as an allelopathic agent: sagebrush inhibits germination of a neighboring tobacco,Nicotiana attenuata

Artemisia tridentata ssp. tridentata is the dominant and defining shrub in the Great Basin Desert, with well-documented allelopathic tendencies that have generally been ascribed to its most abundantly released secondary metabolites. However, as a minor component, sagebrush releases a highly biologically active substance, methyl jasmonate (MeJA), which is known to function as both a germination inhibitor and promoter in laboratory studies. Nicotiana attenuata is a tobacco species native to the Great Basin Desert and grows in newly burned juniper–sagebrush habitats for 2–3 yr following a fire. With a combination of field and laboratory studies, we examined the role of MeJA release from sagebrush by both air and water transport in inhibiting N. attenuata seed germination. We demonstrated that sagebrush interacts allelopathically with the seed bank of N. attenuata through its release of MeJA. In the field, seeds buried 0–40 cm from sagebrush plants for 4 months in net bags had significantly reduced germination compared to seeds buried similarly but protected in plastic bags. Moreover, germination on soils collected from underneath sagebrush plants was reduced by 60% compared to seeds placed on soils collected between sagebrush plants or outside of the sagebrush population. Exposure to A. tridentata seeds and seedlings did not affect N. attenuata germination, suggesting that established sagebrush plants only influence the tobacco's seed bank. In the laboratory, exposure of seeds to sagebrush emissions resulted in germination delays of up to 6 d. Exposure to volatile and aqueous MeJA also inhibited germination of N. attenuata seeds at quantities that are released naturally by sagebrush: 3.5 g/hr and 1.12 g/seed cup (56 ng/seed), respectively. A. tridentata seeds were significantly more resistant to MeJA, being inhibited at 336 g MeJA (16.8 g/seed), 300 times greater than the level of aqueous MeJA required to inhibit N. attenuata seeds. MeJA inhibited N. attenuata germination regardless of the seed's dormancy state and the specific epimer (trans- or cis-) of MeJA. Germination on sagebrush chaff that had been heated to reduce MeJA content was negatively correlated with the amount of MeJA remaining in the chaff. Germination of a nondormant, conspecific tobacco, N. trigonophylla, which grows in the same area but is not associated with fire, is less sensitive than N. attenuata to the extracts of sagebrush litter, but similarly sensitive to MeJA. Additionally, four of five other tobacco species that are not known to be associated with sagebrush are less sensitive to MeJA, suggesting an evolved sensitivity to MeJA. To determine the proportion of germination inhibition of a sagebrush extract that could be attributed to MeJA, we serially diluted sagebrush extracts with water and restored the quantity of MeJA of the original extract by adding appropriate quantities of synthetic MeJA; 16–60% of the inhibitory activity of the original extract could be attributed to the MeJA. We conclude that MeJA release from sagebrush plays an allelopathic role for N. attenuata seed banks, but other unidentified compounds are also involved.     

Allelopathy as a factor in the success ofHelianthus mollis Lam.

A method to rapidly screen species suspected of producing allelochemics, using results from simple bioassay tests, is presented. By measuring the osmotic potential ofH. mollis extracts and using mannitol solutions of comparable osmotic potential, the influence of osmotic potential in the bioassay was eliminated. Nested analysis of variance was used to examine the separate influences of (1) extract concentration, (2) source of plants used in extract preparation (edge or center of clones) (3) osmotic potential of the extract, and (4) the differential development of radicles and shoots of species used in the bioassay tests. Bioassay tests for allelopathy showed that extracts made of wholeH. mollis plants significantly inhibited both radicle and shoot development of radish and wheat, but only the radicle of little bluestem. There was a significant increase in the inhibition of radish shoots and wheat radicles at high concentration of the extract, but the radicle of little bluestem was inhibited more at the lower concentration. Extracts prepared from plants collected from the clone center inhibited radish radicle development significantly more than extracts made of plants growing at the clone edge.     

Chemical inhibition of fire-prone grasses by fire-sensitive shrub,Conradina canescens

In an investigation of potential chemical activity of fire-sensitive shrubs in Florida's sand pine scrub community, bioassays of foliar washes ofConradina canescens showed significant inhibitory activity on three native grasses that are known to fuel frequent surface fires; inhibition was concentrated seasonally in spring and summer. Application of runoff fromConradina leaves to one of the grasses caused a 50% reduction in growth over a 20-week period. Isolation of the biologically active fractions from the fresh leaves ofC. canescens yielded numerous monoterpenes, a number of which were identified from a GC-MS reference library and/or MS comparison to authentic compounds: 11 from the diethyl ether extract, 11 from steam distillation, and four from the foliar leaf wash. Numerous other monoterpenes present in the extractions were unknown. The terpenoid fraction completely inhibited seed germination of one of the native grasses and of lettuce. Saturated aqueous solutions of nine of the monoterpenes inhibited germination and radicle growth of two native grasses. SEM views of the leaf surfaces ofConradina reveal secretory trichomes that appear to be the source of the monoterpenes as well as the triterpene, ursolic acid. The biological activity ofC. canescens as a fire-sensitive component of the scrub community is reviewed in light of the chemical evidence.     

Fungistatic Activity of Ruta graveolens Extract and Its Allelochemicals

The biological activity of an extract of Ruta graveolens and its allelochemicals [8-methoxypsoralen (8-MOP), 5-methoxypsoralen (5-MOP) and 4-hydroxycoumarin] (previously isolated) was evaluated gainst pathogenic fungi: Fusarium solani, Pyrenochaeta lycopersici, and Trichoderma viride. In addition a rue lyophilized extract was also tested against Penicillium sp., Thielaviopsis basicola, and Verticillium dahliae. We determined that 5 g rue lyophilized extract/liter, the lowest concentration tested, inhibited the mycelial growth of P. lycopersici and V. dahliae by 63.6% and 47%, respectively, with IC₅₀ (the concentration required to inhibit growth 50%) values of 4.16 and 6.5 g/liter, respectively. Moreover, the rue lyophilized extract was shown to reduce radial growth of all six fungal species significantly at concentrations ranging from 5.0 to 40.0 g/liter. Mycelial growth of Penicillium sp. was promoted at 5.0 g lyophilized rue extract/liter and inhibited at higher concentrations. 4-OH coumarin had no effect on the growth of these fungi at concentrations ranging from 0.32 to 2.60 mg/liter, whereas 8-MOP and 5-MOP inhibited the mycelial growth of F. solani and T. viride with IC₅₀ values of 6 mg/liter (5-MOP) and 27 mg/liter (8-MOP) for both fungi, respectively. A mixture of 5-MOP and 8-MOP, tested at 64.8 mg/liter (as they naturally occur in the crude rue extract) significantly inhibited the growth of all tested fungal species. These results suggest potential role for the rue extract and its allelochemicals in controlling pathogenic fungal infections.     

Isolation,characterization and activity of phytotoxic compounds from quackgrass [Agropyron repens (L.)Beauv.]

Previous experiments showed that legumes grown in the presence of living or herbicidally treated quackgrass residues or extracts exhibited reduced seedling root and shoot growth and decreased nodulation and nitrogen fixation. Aqueous extracts of quackgrass shoots were most inhibitory to plant growth. Upon sequential partitioning of an aqueous extract of quackgrass shoots, the ether extract possessed the most activity and caused 50% reductions in radicle elongation of eight crop and weed species at concentrations of less than 240 g/ml (small-seeded species) and 1000 g/ml (large-seeded species). Snapbeans (Phaseolus vulgaris L. Bush Blue Lake) grown aseptically in agar containing an ether extract at 100 and 200 g/ml exhibited severe root browning, lack of root hair formation, and a two- to three-fold reduction in root and shoot dry weights. The ether extract of quackgrass shoots had no inhibitory effect on the growth of fourRhizobium species in Petri dishes or two species in broth culture. Inhibitors present in the ether extract may influence the legumeRhizobium symbiosis indirectly by reducing legume root growth and root hair formation. The ether extract of quackgrass shoots was separated using high-pressure liquid, thin-layer, and liquid column chromatography in an attempt to isolate and identify the inhibitors responsible for the inhibition of seedling growth. Two closely related flavonoid inhibitors were isolated from the ether extract. One was identified as 5,7,4-trihydroxy-3,5-dimethoxyflavone (tricin). Both flavonoids caused 50% inhibition of radicle elongation in cress (Lepidium sativum L. Burpee curly) seeds at concentrations of less than 125 g/ml. Both flavonoids were found in ether extracts of quackgrass shoots and rhizomes, but the largest amounts of both compounds occurred in quackgrass shoots collected from the field.Journal article No. 11887 of the Michigan Agricultural Experiment Station.     

Allelopathy of yellow fieldcress (Rorippa sylvestris): Identification and characterization of phytotoxic constituents

Both the neutral and acidic fractions of the acetone extract of yellow fieldcress (Kireha-inugarashi,Rorippa sylvestris Besser) inhibited lettuce seed germination. Salicylic,p-hydroxybenzoic, vanillic, and syringic acid were identified in the acidic fraction. In the neutral fraction, hirsutin (8-methylsulfinyloctyl isothiocyanate), 4-methoxyindole-3-acetonitrile, and pyrocatechol were identified. Bioassay using a root exudate recirculating system showedR. sylvestris during flowering inhibited the lettuce seedling growth. Hirsutin (13g/plant/day) and pyrocatechol (9.3g/plant/day) were the major compounds released into the rhizosphere. Several combinations of pyrocatechol,p-hydroxybenzoic acid, vanillic acid, and hirsutin reduced lettuce seedling growth. These compounds seemed to be allelochemicals.     

Phenolic compounds isolated from bitter lupine seeds and their inhibitory effects on germination and seedling growth of lettuce

The phenolic acids, including 4-hydroxybenzoic and 4-hydroxycinnamic acids, and their derivatives, such as 6,7-dihydroxycoumarin and 1,2-dihydroxybenzene, were isolated from bitter lupine seeds and were identified using gas chromatography-mass spectrometry. These compounds inhibited lettuce seed germination in the first 24 hr after sowing, but after 72 hr germination was comparable with that of the control. However, very strong suppression of seedling growth, especially the roots, was observed for higher concentrations of the lupine seed fractions containing phenolic acids. Effects observed in the lettuce germination bioassays were compared with those produced by nine pure phenolic acids previously identified in the mixture. The ethyl acetate fraction from lupine seed extract inhibited seedling growth as effectively as pure 1,2-dihydroxybenzene, the strongest inhibitor of the pure phenolic compounds studied. The possible reason for this could be the synergistic effect created in the mixture of phenolic compounds isolated from the extract.     

Allelopathic potential ofErica vagans,calluna vulgaris,andDaboecia cantabrica

In laboratory tests the allelopathic potential ofErica vagans, Calluna vulgaris, andDaboecia cantabrica was determined. Aqueous extracts of flowers ofD. cantabrica and leaves ofC. vulgaris inhibit root and hypocotyl growth of red clover, the former causing 51% inhibition of germination. Intact aerial parts of the Ericaceae here studied drastically reduced the growth of red clover and 100% inhibition of germination was caused by flowers ofD. cantabrica. Inhibition of aqueous extracts remains after Chromatographic separation, and two well-defined inhibition zones may be observed. Hydrosoluble organic compounds (phenol-like compounds) could probably be responsible for the inhibitions detected.     

Possible allelopathic constituents ofCoffea arabica

The aqueous extracts of leaves, stems, and roots ofCoffea arabica significantly inhibited the seed germination and radicle growth of rye grass, lettuce, and fescue. When the extracts were diluted to 1% solution, significant suppression of lettuce growth was still found and was particularly pronounced in the extract of young seedlings. The paper chromato-gram of the ether fraction of an aqueous extract of coffee leaves was bioassayed with lettuce seeds and revealed a remarkable inhibition throughout the chromatogram except for the segment ofR _f 0.00–0.12. Paper without spotting extract was used as a standard. The phytotoxins present in coffee tissue were identified by paper and thin-layer chro-matography and mass spectrometry. The compounds include caffeine, theobromine, theophylline, paraxanthine, scopoletin, and chlorogenic, ferulic,p-coumaric,p-hydroxybenzoic, caffeic, and vanillic acids. All compounds except caffeic acid exhibited significant phytotoxicity to lettuce growth at a concentration of 100 ppm.Paper no. 222 of the Scientific Journal Series, Institute of Botany, Academia Sinica, Taiwan, and Journal Article 3582 of the Agricultural Experiment Station, Oklahoma State University, Stillwater, Oklahoma. The abstract was published in the Proceedings of 63rd Annual Meeting of American Societies for Experimental Biology, held on April 1–10, 1979, Dallas, Texas.     

Seed germination and growth inhibitory cadinenes fromEupatorium adenophorum spreng

Chloroform extract of the aerial parts ofEupatorium adenophorum Spreng was fractionated and examined for growth inhibition. Bioassay-directed fractions of the plant materials afforded three known cadinenes and -sitosterol. The effects of different fractions as well as isolated cadinenes were determined usingAllium cepa, Raphanus sativus, andCucumis sativus seeds. Three-day exposure to these cadinenes significantly inhibited germination and seedling growth of all three assay seeds. The degree of inhibition was dependent upon seed species and the concentrations of the compounds tested. Cadinene(1) was found to be more inhibitory to the seeds tested and the activity of the cadinene(3) was less than that of(1) and(4).     

HPLC identification of allelopathic compounds fromLantana camara

Aqueous extracts ofLantana camara L. leaves inhibited ryegrass (Lolium multiflorum Lam.) germination and seedling growth. Phytotoxic compounds were fractionated from crude aqueous extracts and fractions were evaluated for their phytotoxicity. Inhibition was most pronounced with the alkaline and acid hydrolysates. Plant inhibition by the crude extract reflected a complex interaction of numerous individual components of diverse chemical compositions and potencies. Presumptive identification of the individual components was accomplished with high-performance liquid chromatography (HPLC). Thirteen phenolic compounds were identified, and most of these compounds were phytotoxic to ryegrass seedlings. Radicle elongation was more sensitive to the toxins than shoot elongation.Florida Agricultural Experiment Station Journal Series No. 7906.     

Allelopathic effects of water extracts ofArtemisia princeps var.orientalis on selected plant species

The allelopathic effects of wormwood plants (Artemisia princeps var.orientalis) and their possible phytotoxicity on receptor species were investigated. The aqueous extracts of mature leaf, stem, and root of wormwood plants caused significant inhibition in germination and decreased seedling elongation of receptor plants, whereas germination of some species was not inhibited by extracts of stems and roots. Dry weight growth was slightly increased at lower concentrations of the extract, whereas it was proportionally inhibited at higher concentrations. The calorie value of the organic matter in receptor plants measured by bomb calorimeter was reduced proportionally to the extract concentration. However, results with extracts of juvenile leaf did not correlate with inhibition or promotion of elongation and dry weight.     

Biological activity and tentative identification of flavonoid components in velvetleaf (Abutilon theophrasti Medik.) seed coats

The biological activity of aqueous extracts of velvetleaf (Abutilon theophrasti Medik.) seed coats and their flavonoid components against three plant species and five soil fungi was investigated. Aqueous extracts slightly inhibited germination and significantly inhibited radicle growth of all plant species tested. Fungal growth was inhibited or not affected, depending on species, by aqueous extracts. The aqueous extracts were extracted into methanol and separated using paper chromatography for identification of compounds responsible for inhibition of seedling and fungal growth. Six flavonoid compounds were isolated and tentatively identified as delphinidin, cyanidin, quercetin, myricetin, (+)-catechin, and (–)-epicatechin. Bioassays revealed that the flavonoid compounds significantly inhibited germination and radicle growth of all test species at a concentration of 1.0 mM. Individual flavonoids had variable effects on fungi but appeared to inhibit growth and sporulation of potential seed-decomposing fungi rather than beneficial fungi. These results indicate that the flavonoid complement of velvetleaf seed coats may function in a dual defensive role against competing seedlings and seed-attacking fungi.Journal article No. 9878 of the Missouri Agricultural Experiment Station.     

Allelopathy in agroecosystems: Wheat phytotoxicity and its possible roles in crop rotation

The germination rates of cotton and wheat seeds were significantly affected by various extracts of wheat mulch and soils collected from the wheat field. This toxicity was even more pronounced against seedling growth. Five allelochemics: ferulic,p-coumaric,p-OH benzoic, syringic, and vanillic acids, were identified from the wheat mulch and its associated soil. Quantitatively, ferulic acid was found at higher concentrations thanp-coumaric acid in the soil. Various concentrations of ferulic andp-coumaric acids were toxic to the growth of radish in a bioassay. The functional aspects of allelochemic transfer from decaying residue to soil and the subsequent microbial degradation within agroecosystems are discussed, particularly as they relate to wheat crop rotation, with wheat and cotton, in Pakistan.     

Allelopathy ofSasa cernua

Sasa (Sasa cernua Makino) is a very serious weed pest. Its allelopathy was studied using lettuce, wheat, timothy, and green amaranth as testing species. Cultured in the rhizosphere soil ofSasa cernua, the seedlings were inhibited by 42–80% compared with the controls cultured in normal soil and vermiculite. The phenolic fraction extracted with 1 M NaOH from the rhizosphere soil ofS. cernua caused significant inhibitions on the seed germination and seedling growth of lettuce, timothy, green amaranth, and barnyard grass.p-Coumaric, ferulic, vanillic, andp-hydroxybenzoic acids andp-hydroxybenzaldehyde were identified as the main allelochemicals in sasa soil by HPLC and [¹H]NMR. Their contents in the rhizosphere soil were 5640, 1060, 860, 810 and 630 μg/100 g soil. The neutral fraction inhibited the seed germination and seedling growth of lettuce in the TLC direct bioassay. Volatile compounds released from sasa leaves also inhibited the growth of lettuce, wheat, timothy, and green amaranth grown under light, and the growth of etiolated seedlings of barley and wheat. These results confirm thatS. cernua produces typical allelopathy through its rhizosphere soil and air space.