Potential Allelopathic Effect of Species of the Asteraceae Family and Its Use in Agriculture

Abstract

Some species are capable of producing substances that affect seed germination, stimulating, or retarding this process, and can also suppress the development of other plants, acting as an antagonistic plant. This can occur naturally, through the release of exudates, or through the action of essential oil, extracts obtained from different parts of the plant, or plant residues with potential allelopathic action. The aim of this chapter is to present the main plant genera of the Asteraceae family with potential phytotoxic or allelopathic activity, with a suppressive effect on the growth of herbicide-tolerant weeds. The genus defined were Acmella, Artemisia, and Bidens, highlighting the form of use—plant extract, essential oil, or plant residues. The Asteraceae family is considered a repository of species to be explored for allelopathy with several associated secondary metabolites such as terpenes, saponins, alkaloids, alkamides, cinnamic acid derivatives, and flavonoids. In addition to these, for the genus Bidens, the presence of the acetylenic compound phenylheptatriine (PHT) is considered an important allelochemical with potent allelopathic action. The presence of this compound is associated with the cytotoxic activity of representatives of this genus, which can be a source of prospecting for new molecules to be used as bioherbicides.

1. Introduction

The term allelopathy has been referenced and conceptualized in different ways and from different perspectives. It can be understood as the ability of plants to interfere with other organisms in the environment [1]; as a process where chemical compounds are released into the environment by an organism and, once released into the environment, interact and can influence the growth and development of biological systems, including inhibition or stimulation effects [2, 3]. Older definitions, such as that of Hans Molisch in 1937, consider allelopathy as the direct or indirect result of the transfer of chemical substances from one plant to another [4]. Thus, in 1996, the International Allelopathy Society (IAS) defined allelopathy as the science that studies any process, essentially involving secondary metabolites produced by plants, algae, bacteria, and fungi, that influence the growth and development of agriculture and biological systems, including positive (stimulation) or negative (inhibitory) effects [5, 6]. It can be seen that, despite the different definitions, these refer, in short, to the central role of secondary metabolites in allelopathy [1], which are involved in defining the characteristics of natural ecosystems and agroecosystems [7].

The compounds identified with potential allelopathic activity are known as allelochemicals and, since their discovery, research has been carried out with the objective of isolating and identifying the substances responsible for this phenomenon and grouping them [8]. Allelochemicals can range from simple hydrocarbons to complex compounds of high molecular weight and can be classified into 10 categories according to their structures and properties: (1) water-soluble organic acids, straight chain alcohols, aliphatic aldehydes, and ketones; (2) simple lactones; (3) long-chain fatty acids and polyacetylenes; (4) quinones (benzoquinone, anthraquinone, and quinone complex); (5) phenolics; (6) cinnamic acid and its derivatives; (7) coumarins; (8) flavonoids; (9) tannins; and (10) steroids and terpenoids (sesquiterpene lactones, diterpenes, and triterpenoids) [9].

The performance of bioassays makes it possible to identify the phytotoxicity of different species, explained by the delay in seed germination, inhibition of plant growth, or any adverse effect caused by specific substances (phytotoxins) or growth conditions [10]. Unlike herbicides, allelochemicals act at low, but constant concentrations, over a long period of time. This fact, associated with the increase in global demand for organic products in the last two decades [11], makes it urgent to invest in studies on the use of allelochemicals as natural pesticides, in order to promote more sustainable agriculture, minimize the effects of pesticides on the environment and human health [12]. Among the benefits associated with the use of allelopathic compounds for the development of new agrochemicals, the fact that most of them are biodegradable and less polluting than traditional pesticides is highlighted due to their shorter half-lives [13].

Allelochemicals can be found in different parts of the plant, including flowers, leaves, stems, roots, or fruits of different species [2, 14]. Secondary metabolites present in medicinal and weed plants have been reported as potent growth inhibitory agents, indicating that such plants act as a depot for allelopathic compounds [15]. Many of these species are representatives of the Asteraceae family, which is composed of approximately 1000 genera, comprising more than 25,000 species of flowering plants [16], representing the largest family among flowering plants in the world, with distribution on all continents, except for Antarctica [17, 18]. This family includes food crops such as lettuce (Lactuca sativa L.), endive (Cichorium endivia L.), edible safflower seeds (Carthamus tinctorius L.), and sunflower (Helianthus annuus L.), species used in oil production (Enclyclopaedia Britannica 2015), medicinal species like Achillea millefolium L. [19], Vernonia spp. [20], and Matricaria chamomilla L. [21]; and species used in the bioremoval of pollutants in urban areas, such as metals and xenobiotics (Solidago, Tanacetum, and Rudbeckia) [22, 23]. Several secondary metabolites are present in the Asteraceae family such as terpenes [24], including sesquiterpene lactones [25, 26], saponins [27], alkaloids [28], alkamides [29], cinnamic acid derivatives, and flavonoids [30].

In this sense, this review intends to highlight, as a target for the search for natural alternatives in crop protection, genera of the Asteraceae family that grow spontaneously in different environments [31], some of which are capable of influencing the development of other species by allelopathy. Therefore, three genera of plants belonging to the Asteraceae family with recognized allelopathic activity were selected: Acmella, Artemisia, and Bidens. Artemisia and Bidens along with Ambrosia, Bellis, Helianthus, and Tagetes are the main genera of the Asteraceae family with allelopathic or phytotoxic activity. Furthermore, the phytotoxic potential of Acmella oleracea was recently described, for the first time, against the weeds Calopogonium mucunoides and Ipomoea purpurea [8], confirming allelopathy of its extract.

A. oleracea (L.) R. K. Jansen (synonymies Spilanthes oleracea L., S. oleracea Jacq., and Spilanthes Acmella auct. Non (L.) Murr.) popularly known as “jambu” is a plant native to the regions of Asia and South America (especially in the northern region of Brazil), where it is widely used in regional cuisine [32, 33, 34]. It is commonly used in folk medicine with proven healing, antispasmodic, anti-inflammatory, antimalarial activity, in the treatment of rheumatism, as a tonic [35, 36], antioxidant, antinociceptive, anti-inflammatory, diuretic, and anesthetic [37]. It also has larvicidal [38], insecticidal [39], acaricide [40], and anthelmintic [41] effects. Acidic amino acids, triterpenes, stigmasterol, and alkaloids predominate in the phytochemical profile of A. oleracea, however, the biological activity seems to be related to the abundant presence of N-alkylamides, especially spilanthol [37, 42]. Studies on allelopathic activity and the metabolites involved, however, are still poorly explored, except for the work of [43] who evaluated the inhibition effect, phytotoxicity, and metabolites present in the aqueous methanolic extract of A. oleracea (L.) R. K. Jansen on the growth of Lolium multiflorum Lam. and Echinochloa crus-galli (L.) P. Beauv.

The genus Artemisia has more than 350 species and is considered a promising source of biologically active compounds with the potential to provide new herbicides and growth regulators. The different species that make up the genus have phytotoxic compounds for monocots, dicots, photosynthetic bacteria, and endomycorrhizal fungi [44], especially Artemisia annua L., which center of origin is Asia [45] but after domestication it began to be cultivated in different countries such as Austria, Brazil, Spain, the United States, France, Poland, and Romania [46]. A. annua stands out in the Asteraceae family both for the variety of natural products characterized (almost 600 in total, including around 50 amorphane and cadinane sesquiterpenes), and by the highly oxygenated nature of secondary metabolites of the terpenoid class [47]. There are several studies that report the allelopathic activity of artemisinin and its synthetic derivatives, which can act as inhibitors and promoters of complex signals in response to biotic and abiotic factors [48, 49, 50, 51, 52], from the aqueous extract, essential oil, or biomass. Plant of its own species (Figure 1).

Figure 1.

Graphic summary referring to the main genera of plants of the Asteraceae family used how potential allelopathic in agriculture.

Bidens pilosa L. is an annual plant native to tropical America and widely distributed in tropical and subtropical regions of the world. The genus has about 280 species and is widespread in both cultivated and uncultivated areas, being considered one of the most harmful weeds in agriculture, promoting crop losses in more than 40 countries [53]. B. pilosa shows rapid growth exhibiting allelopathic effect on various cultures [53, 54, 55, 56, 57]. It is also used as a medicinal plant, cover plant, and source of nectar for bees. Its roots, leaves, and seeds have antibacterial, antidysenteric, anti-inflammatory, antimalarial, antiseptic, anticancer, antipyretic, hepatoprotective, hypotensive, hypoglycemic, diuretic, and antidiabetic activity [58, 59].

Xuan and Khanh [60] systematized a literature review based on 218 literary sources reported over 40 years highlighting chemical constituents, nutraceuticals, ethnomedical, biological, and pharmacological uses, and the effects and toxicity of B. pilosa. In this survey, the authors reported that the main compounds (301 compounds) belong to the group of polyacetylenes, polyacetylene glycosides, flavonoids, flavone glycosides, aurones, chalcones, okanine glycosides, phenolic acids, terpenes, pheophytins, fatty acids, and phytosterols, the which were identified and isolated from different parts of this plant and considered as bioactive compounds potentially responsible for the pharmacological action, biological and allelopathic properties of the species, which will be described in more detail below.

It is important to highlight, however, that most of the articles that suggest the allelopathic effect of the crude extract of a plant species do so through bioassays, useful tools for previous studies on the allelopathic potential of a species or compound, however, not suitable for sufficient to relate the results obtained in vitro, in the laboratory, with field conditions [3, 61, 62]. For this reason, results from bioassays developed under natural conditions were included in this review [63, 64]. Under these conditions, the biosynthesis of allelochemicals and their release can be influenced by temperature, luminosity, humidity, interaction with soil biota, and nutrient availability [3, 65, 66]; in addition to the fact that plants are evaluated at the initial stage of development, considered the most sensitive stage to allelochemical activity [67, 68].

2. Genus in Asteraceae family with potential allelophatic

2.1 Genus: Acmella

2.1.1 Extract

Among the species belonging to the genus Acmella, Acmella oleracea (L.) R.K. Jansen stands out, popularly known as “jambú” [32, 69]. The allelopathic activity of this plant is still poorly explored. Kato-Noguchi [43] evaluated the activity of methanolic extracts of A. oleracea, from the whole dried plant, at concentrations of 3, 10, 30, 100, and 300 mg/ml. The biological activity was tested against Lepidum sativum, Lactuca sativa, Lolium multiflorum, and Echinochloa crus-galli, for this, 10 pre-germinated seeds were used, using Tween 20 as control. After 48 hours, the length of the roots and shoots of the seedlings were measured and the concentrations necessary to inhibit 50% of the growth (IC50) of the roots and shoots were determined. The A. oleracea extract showed dose-dependent action. At a concentration of 30 mg/ml the extract inhibited root growth in 4.7, 0, 21.3, and 57.1%, and for the shoot in 7.8, 0, 43.4, and 98.9% for Lepidium sativum, L. sativa, L. multiflorum, and E. crus-galli, respectively, when compared to the control.

When the IC50 values in the roots were analyzed, they were 5.9, 0.4, 25.9, and 7.8 mg/ml and in the shoot 4.3, 0.8, 126.0, and 15.1 mg/ml, for L. sativum, L. sativa, L. multiflorum, and E. crus-galli, respectively. The extracts were purified and two compounds were isolated, identified as (E,E)-2,4-undecadien-8,10-diinoic acid isobutylamide and nona-(2Z)-en-6,8-diinoic-2-phenylethylamide. The first compound inhibited the growth of roots and shoots of L. sativum and root of E. crus-galli at concentrations greater than 0.3 mM. The second compound inhibited the growth of L. sativum shoots and E. crus-galli roots at concentrations greater than 1 mM. Thus, these two compounds may be associated with the allelopathic activity of A. oleracea [43].

Araújo [8] analyzed the allelopathic activity of the hydroalcoholic extract, hexane, dichloromethane, and ethyl acetate fraction from the dry leaf of A. oleracea. The phytotoxic activity was tested on L. sativa using concentrations between 0.1 and 1 mg/ml of extracts and fractions. Calopogonium mucunoides was also used as a test species for the fractions at a concentration of 0.2 mg/ml. Ten seeds of L. sativa and five seeds of C. mucunoides were sown in Petri dishes containing a filter paper. Water and DMSO were used as negative controls and menadione as positive. Germination was recorded after 24 hours by root protrusion and root length was measured after 5 days. For L. sativa, both the hydroalcoholic extract and the fractions affected its germination, and the dichloromethane and ethyl acetate fractions were more harmful. At the concentration of 1 mg/ml, the inhibition was 76 and 60% of root growth for the dichloromethane and ethyl acetate fractions, respectively. The IC50 for the dichloromethane fraction was 0.48 mg/ml, which is similar to the results of Kato-Noguchi [43]. All fractions inhibited the germination of C. mucunoides, around 60%, at the concentration studied, and compared to lettuce seeds, the latter seems to be more resistant to A. oleracea allelochemicals. The dichloromethane fraction presented as major compounds: p-methoxy-cinnamic and 3,4-dimethoxy-cinnamic acids, 3,4-dimethoxy- and 3,4,5-trimethoxybenzoic acids, and palmitic acid. The presence of fatty acids and phenolic acids are related to the allelopathic activity presented by this fraction. The IC50 values found were 1.13, 0.94, 0.36, 0.37, and 0.19 mg/ml for hydroalcoholic extract, hexane, dichloromethane, ethyl acetate, and aqueous residue, respectively.

2.1.2 Vegetable residue

Suwitchayanon, Kunasakdakul and Kato-Noguchi [70] evaluated the allelopathic activity of 14 plants, including Acmella oleracea. L. sativa was used as a test plant. A. oleracea plant material was dried and ground and added to the Petri dish together with agar and finally, a new layer of agar was added. As a control, only agar was used. Each plate received five L. sativa seeds and incubated for 48 hours. All the plants studied showed a greater inhibitory effect on root development than on the hypocotyl. A. oleracea at a concentration of 50 mg dry plant weight inhibited radicle appearance in 71.8% and hypocotyl development in 47.4%. There is a need for further studies using this technique with different concentrations of the plant drug.

2.2 Genus: artemisia

2.2.1 Extract

Many researches have evaluated the allelopathic effects of aqueous extracts, or even volatile organic compounds, released by different parts of Artemisia plants [71, 72, 73]. Water stress conditions [74] and drought conditions [75] can intensify the allelopathic effect of the aqueous extract of these plants. Seeking answers to these statements, [71] developed an experiment of laboratory simulations of stress in relation to artemisinin production, obtained by the aqueous extract of A. trifida in environments with abiotic stress conditions. In view of the reported results, Guo [71] organized their experiment through tests of growth and development of rye subjected to four treatments, one characterized by the application of Hoagland’s nutrient solution, a second with artemisinin solution at a concentration of 20 ml/l and, the others characterized by the combinations of the previous treatments, being one with cultivation of rye under the addition of extract of A. trifida at 5% (mass/volume) and Hoagland solution and, finally, cultivation with 20 mg/l of artemisinin solution together with A trifida extract. In Guo’s research [71], abiotic stress was characterized by the submission of rye plants to freeze/thaw cycles and, throughout development, biomass accumulation, photosynthetic parameters, relative water content, malondialdehyde acid (MDA) and activity of superoxide dismutase, catalase, peroxidase, and ascorbate peroxidase enzymes. By applying the methodology and collecting the results, the authors concluded that the allelopathic effect of the A. trifida extract can be intensified by the abiotic stress studied, promoting inhibition of growth, biomass, photosynthesis, in addition to triggering MDA, and osmotic regulatory substances on rye seedlings.

Pastures are the food base for the herds that drive the world economy, but most of them are degraded by either anthropogenic actions or agricultural activities. Leaving this comfort zone for a widely accepted response of grassland degradation, Wang [72] went further and asked himself: is the allelopathic effect of A. frigida enough to explain, in part, the degradation of grasslands in northern China? And they added: does the allelopathic effect of A. frigida intensify as pasture degradation progresses? To answer the hypotheses raised in the study, the authors collected samples of A. frigida, randomly distributed in the experimental plots, from pastures with different grazing intensities for 180 days, considering such intensities as low, medium, and high. As a control, an area without animal grazing was kept next to the other areas. From the samples collected, Wang [72] cut 3 mm sections of the aerial part of A. frigida plants and obtained the aqueous extract by adding the plant tissues in an Erlenmeyer flask and placing them in an incubator at 20°C for 48 hours. After the time, the material was filtered through a sieve with a diameter of 0.5 mm. Concentrations of 0.25 g/ml (0.25 g of A. frigida aerial part per ml of distilled water) of the aqueous extract from each grazing intensity plot were kept at 4°C for a period of 10 days.

The allelopathic effect was evaluated in two different ways by Wang [72]. At first, the authors verified the effect of seed germination of Melilotus officinalis, M. sativa, and A. splendens (plants found in grazing areas) when submitted to aqueous extracts of A. frigida. In a second moment, the authors cultivated A. frigida plants in pots for 2 months to verify the allelopathic potential on their own species. In this experiment, the authors used activated carbon mixed with soil to absorb the allelochemicals released by mugwort plants. As a first result, the authors verified that 1,8-cineole and β-terpineol may be the main components contained in A. frigida induced by the grazing gradient. It is also important to report that the authors concluded that as grazing is intensified, the allelopathic effect of A. frigida increases significantly, since the concentration of secondary metabolites increases in the chemical composition of the extract. This variation in the concentration of chemical components verified by the authors in the aqueous extracts significantly inhibited seed germination and seedling growth of all grass species tested in the experiment.

Zhou [76] studied the phytotoxic effect of the aqueous extract of Artemisia ordosica leaves against two soil microalgae (Chlorella vulgaris and Nostoc sp.). The algae cells were grown in pyrex flasks with the addition of 0, 1, 5, 10, or 25 g/l of A. ordosica extract, with a light:dark cycle of 12 hours:12 hours. The cultivation without the addition of the extract served as a control. The concentration of chlorophyll a after 48 hours of incubation was also evaluated. The aqueous extract of A. ordosica showed 23 main compounds including alcohols (d-pinitol, diethylene glycol, 2,3-butanediol, inositol, glycerol, and tetriite), organic acids (glycolic acid, palmitic acid, octadecanoic acid, acid, and pentaric acid), phenolic derivatives (3,4-dihydroxyphenylglycol, 3-vinylcatechol, 1-(5-hydroxy-2-methoxyphenyl)-1,2-ethanediol, and 2-(2-hydroxyethyl)phenol), and sugars (inositol, d-glucopyranose, sucrose, and l-mannitol). The growth rates of C. vulgaris increased significantly at the concentration of 1 g/l and the chlorophyll a content increased with the time of culture under 5 g/l of the extract of A. ordosica. The highest concentrations (10 and 25 g/l) inhibited the growth of C. vulgaris and the concentration of chlorophyll a decreased. The growth rates of Nostoc sp. were not affected in the presence of 1 g/l of extract, and the other treatment concentrations inhibited the microalgae growth.

Luo [77] evaluated the allelopathic activity of aqueous extracts of leaves, stems, and roots of Artemisia halodendron against Agriophyllum squarrosum, Setaria viridis, Artemisia scoparia, Lespedeza davurica, Chenopodium acuminatum, and Corispermum macrocarpum. The extracts were used at concentrations of 10, 20, 40, 60, 80, and 100 g/l, and distilled water was used as a control. The analyzes were performed in Petri dishes adding the extracts and 30 seeds of the test plants. Afterward, the plates were incubated for 21 days with a photoperiod of 14 hours with light and 10 hours in the dark. At the end of the experiment, the seeds that showed growth of both the radicle and the bud were analyzed. The germination rate of seeds varied significantly between the six species under treatment and between the concentrations evaluated. The present study suggests a significant negative allelopathic effect of A. halodendron in other species. The extracts significantly reduced the germination of A. squarrosum seeds, and this suppression increased with increasing concentration, with the leaf extract being more effective when compared to the stems and roots. For Centrosema macrocarpum the extracts did not differ in their effects on seed germination.

Li [78] analyzed the biological activity of Artemisia argyi against Brassica pekinensis, L. sativa, and Oryza sativa. The extracts were prepared with dried leaves of A. argyi having water, 50% alcohol, and 100% alcohol as solvents. The concentrations of the extracts used were 50, 100, and 150 mg/ml. Twenty seeds of the tested species were added to Petri dishes with two layers of filter paper and then treated with extracts prepared from A. argyi at the different study concentrations. Ultrapure water was used as a control. The plates were grown at 25°C, 85% humidity, and a controlled cycle of 12 hours light/12 hours dark. The number of germinated seeds was counted from the second day after treatment and the counting lasted 1 week. The root length, stem length, and biomass of each treatment were also analyzed.

The three types of extracts were analyzed by UPLC-Q-TOF-MS, the aqueous extract presented as main components: caffeic acid, schaftoside, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3,5-dicapheoylquinic acid; 50% ethanol extract: 4,5-dicapheoylquinic acid, 3-caffeoylquinic acid, schaftoside, rutin, kaempferol 3-rutinoside, 3,4-dicapheoylquinic acid, 3,5-caffeoylquinic acid, 3-caffeoyl acid, 1-p-coumaroylquinic acid, 1,3,4-tri-caffeoylquinic acid, and eupatiline, and for the 100% ethanol extract the main metabolites were eupatiline, jaceosidine, and casticin. There was a concentration-dependent increase in inhibition of the extracts used. The aqueous extract exerted a significant inhibitory effect on germination and biomass production of the evaluated plants. The 50% ethanol extract showed inhibition on the germination and biomass index of B. pekinensis and L. sativa, but moderate inhibitory effects on O. sativa. The ethanol extract (100%) showed only inhibitory effects on the germination of B. pekinensis and L. sativa and no effect on O. sativa. However, it exerted inhibitory effects on the biomass of the three plants at high concentrations. Thus, it is possible to affirm that aqueous extract of A. argyi exhibited the strongest allelopathic effect. For example, B. pekinensis can be significantly inhibited by a low concentration of extract (50 mg/ml), showing the order of inhibition efficiency of: germination index > germination speed index > root length > germination rate > stem length > biomass. In summary, according to the comprehensive allelopathy index of the indicators studied, the order in which they were sensitive to the aqueous extract was B. pekinensis > L. sativa > O. sativa. O. sativa was selected as a test plant for RNA isolation and sequencing, transcriptome data and RT-qPCR verification showed that suppression of chlorophyll synthesis and photosynthesis was one of the main mechanisms of the inhibitory effect of A. argyi on test plants.

2.2.2 Essential oil

Researches with aqueous extract of Artemisia are broader and gain prominence in research, however, the essential oil of this plant can also present allelochemicals. Reports in the literature confirm that Artemisia essential oils can exert high phytotoxicity against weeds and other undesirable plants in a crop [79, 80] because they present terpenoids, especially monoterpenes and sesquiterpenes, which are the main components of the essential oil and are often responsible for its plant inhibitory activity [81].

Within this line of research, Yang [82] investigated the chemical composition of allelochemicals present in the essential oil of Artemisia ordosica and the effects that this could have on growth rates, photosynthetic activity, and oxidative damage in plant species present in biological soil crusts. In the work of Yang [83], the experimental units were composed of soil collected, superficially, on which a total of 0, 1, 3, 5, or 10 ml of A. ordosica essential oil were applied. These volumes were dissolved in 50 ml of dimethylsulfoxide. The essential oil in this study was obtained by hydrodistilling, for 3 hours, 100 g of air-dried leaves of A. ordosica, using the steam distillation method as described in [84]. The authors evaluated possible alterations in the plants present in the biological crust of the soil by means of analyses, carried out in triplicate, of chlorophyll a fluorescence, presence of reactive oxygen, in addition to the activities of the enzymes peroxidase, superoxide dismutase (SOD), and malondialdehyde acid (MDA). As highlighted results observed by the authors, mention is made of the 37 chemical components identified in the essential oil of A. ordosica, with emphasis on terpenoids, alcohols, esters, and acetones. The high concentrations of allelochemical compounds present in the essential oil resulted in the inhibition of photosynthetic activity (Fv/Fm), decrease in photosynthetic parameters ET0/ABS, ET0/TR0, and RC/CS0 due to the decrease in electron flow and photon absorption and reduction of the PSII reaction center. There were also increases in POD, SOD activities, and the presence of MDA. The results indicated that the essential oil of A. ordosica inhibited the growth and negatively affected the development of plants present in the biological crust of the soil surface.

The allelopathic potential of Artemisia essential oil was tested on seed germination of nine weeds and two wheat varieties by Benarab [85]. In this study, the essential oil was obtained by hydrodistillation of the aerial part of the plants. After characterization by gas chromatography coupled with mass spectrometry, the essential oil was tested on seed germination and seedling growth of weeds and wheat. The concentrations used were 0.2, 0.4, 0.6, and 0.8 μl/ml, with 1 ml of these added in Petri dishes used for germination tests. The authors found 36 chemical compounds in the essential oil of Artemisia herba-alba, especially camphor (28.58%), cis-thujone (22.03%), eucalyptol (11.65%), and trans-thujone (7.03%). They found that essential oil extracts have a significant effect on weed germination inhibition, with the best results having been observed at a concentration of 0.2 μl/ml. These positive results for the allelopathic potential of the essential oil evaluated, suggest that, according to Benarab [85], Benvenuti [79], Önen [80], the same can be tested as a bio-herbicide.

With objectives similar to those of the works mentioned above, Benvenuti [79] collected 20 species of Asteraceae, during the spring–summer period, in several places in the Tuscany region. From each of the species, the yield and quality of the essential oils obtained were evaluated, as well as the verification of the inhibition of germination and growth of weeds. The essential oils were obtained by hydrodistilling the dried and ground flower buds in a Clevenger apparatus for 2 hours and their yield was calculated considering the values of the dry biomass of the flower buds collected in an area of 1 m². As some species did not show sufficient oil yields, the authors used only 10 species of Asteraceae in this experimental design, which consisted of Petri dishes lined with filter paper (Whatman no. 1, Whatman, Maidstone, UK) moistened with 7 cm³ of distilled water, on which 50 seeds of each weed species were placed. Essential oils in concentrations of 10 and 100 μl/l were added to the plates. A second experiment was tested by the authors. In this, the essential oils were sprayed at concentrations of 10, 100, and 1000 mg/l, in weeds at two different phenological stages, when they had expanded cotyledons and when the third true leaf appeared. After analyzing the results, Benvenuti [79] found total inhibition of germination of the weed A. retroflexus at a concentration of 10 μl/l, and as the main result of experiment two, a reduction in weed fresh weight (about 20–30% after 10 days) and chlorophyll content (destroyed after the same period of time) was observed after application of the essential oil of Artemisia annua, thus confirming the total and rapid effectiveness of these essential oils and, according to the authors of the work, the essential oil of A. annua can be used as a natural herbicide.

2.2.3 Vegetable residue

Li [78] also evaluated the allelopathic activity of Artemisia argyi leaf powder against B. pekinensis, L. sativa, O. sativa, Portulaca oleracea, Oxalis corniculata, and S. viridis. The experiment was carried out in pots containing sand soil and A. argyi leaf powder for soil preparation, A. argyi powder was mixed with sand soil in a ratio of 100:0, 100:2, 100:4, or 100:8. Fifty seeds of B. pekinensis, L. sativa, O. sativa, P. oleracea, O. corniculata, and S. viridis were sown independently in each pot. Germination rate and plant height were measured on the ninth day for B. pekinensis, L. sativa, and O. sativa and on the thirteenth day for P. oleracea, O. corniculata, and S. viridis. When the soil:powder ratio of A. argyi leaves was 100:2, the germination rate of B. pekinensis and L. sativa was inhibited, as was the height of plants of B. pekinensis, L. sativa, O. sativa, and P. oleracea. The rates analyzed were dependent on the proportion of powder, the higher the concentration, the greater the inhibition. In the ratio of 100:8, the germination inhibition rates of B. pekinensis, L. sativa, O. sativa, P. oleracea, O. corniculata, and S. viridis were 71.82, 93.20, 31, 75, 65.47, 63.60, and 60.78%, respectively. These authors also analyzed the activity of A. argyi leaf powder in a field experiment. Chrysanthemum morifolium seedlings were transplanted to beds divided into plots. The A. argyi powder was uniformly applied to the plots at concentrations of 0, 0.1, and 0.2 kg/m². After 30 days, the biomass of each treatment group, growth, and yield of C. morifolium was evaluated. A. argyi leaf powder inhibited the germination and growth of the test plants. Only one species grew in the group treated with 0.2 kg/m². After the harvest of C. morifolium, there were no significant differences in the number of flowers and in the weight of the flowers between the groups with the addition of A. argyi leaf powder and the control group. Therefore, A. argyi leaf powder did not inhibit the growth of C. morifolium in the field, it only exerted an inhibitory effect on weed seeds. Thus, the powder from the leaves of A. argyi can be used as a herbicide.

2.3 Genus: bidens

2.3.1 Extract

B. pilosa is a weed widely distributed in subtropical and tropical regions with phytotoxic action already described for different species. The allelopathic activity of B. pilosa was evaluated by Deba [86] from the aqueous extract of the acid fraction (100, 200, and 500 ppm) of its leaves, stems, and roots. The species used as indicator plants were Raphanus sativus and E. crus-galli. For this, 10 seeds of each species were placed in Petri dishes lined with filter paper soaked with 8 ml of each solution of B. pilosa extract. As a control treatment, distilled water was used. After 7 days at room temperature (25–28°C) the percentage of germination and the length of the shoot and roots of R. sativus and E. crus-galli.

The extracts of B. pilosa showed a strong reduction in the growth of the indicator plants, however, the inhibition of germination did not exceed 20% in all treatments compared to the control. At all concentrations tested, the inhibitions promoted by stem and root extracts against E. crus-galli and R. sativus ranged from 70 to 90%. The herbicidal activity of B. pilosa was proportional to the applied doses, with almost complete inhibition of the growth of hypocotyls and radicles of E. crus-galli and R. sativus at a concentration of 500 ppm.

Chemical analysis of B. pilosa extracts by GC–MS detected 15 compounds including pyrocatechin, salicylic acid, p-vinylguaiacol, dimethoxyphenol, eugenol, 4-ethyl-1,2-benzenediol, iso-vanillin, 2-hydroxy-6-methylbenzaldehyde, vanillin, vanillic acid, p-hydroxybenzoic acid, protocatechuic acid, p-coumaric acid, ferulic acid, and caffeic acid. All of them were found in root extracts, except vanillin and iso-vanillin. Demethoxyphenol, eugenol, iso-vanillin, and vanillic acid, in turn, were not found in the stems of B. pilosa; and, in the leaves, salicylic acid, dimethoxyphenol, and vanillic acid were not observed. In all parts of the evaluated plant, the phenolics pyrocatechin, p-vinylguaiacol, 4-ethyl-1,2-benzenediol, 2-hydroxy-6 methylbenzaldehyde, p-hydroxybenzoic acid, protocatechuic acid, p-coumaric acid, and ferulic acid were verified, and caffeic acid, present in much greater amounts than the other phenolics (117.4, 298.7, and 350.3 μg/g in leaves, stems, and roots, respectively). In roots, ferulic acid was higher than pyrocatechin; however, it was lower in leaves and stems. In short, the total amount of these phenolic acids in the root.

Khanh [87] reported the allelopathic effect of leaves, stems, and roots of the acidic ethyl acetate fraction of B. pilosa extract on the same species tested (E. crus-galli and R. sativus) by Deba [86]. Root and stem extracts showed inhibitory effects of 70–90% on the emergence of E. crus-galli and R. sativus, for concentrations from 100 to 500 ppm. The potential allelopathic effect of B. pilosa extract on crops and weeds has already been described for other crops, with reports of inhibition of germination and growth of seedlings of soybean, mung bean, rice, corn, radish, cucumber, lettuce, sorghum, peanuts, and vines [87, 88, 89, 90, 91].

Hsu and Kao [54] evaluated the allelopathic effect of the aqueous extract of leaves, stems, and roots of Bidens pilosa var. radiata on the germination and growth of seedlings of the same species and of the sympatric species, Bidens bipinnata and Ageratum conyzoides. The plants of B. pilosa var. radiata were collected in an abandoned area of a farm in southern Taiwan, and then separated into leaves, stems, and roots, which were used to prepare the aqueous extract. The effect of the extract on seedling growth was verified by measuring the radicle and hypocotyl length of the germinated seeds of the tested model plants. The control treatment consisted of the use of distilled water. In this treatment, 99% of B. pilosa var. radiata germinated after 8 days of incubation. The final germination percentage of B. pilosa var. radiata was significantly reduced by the application of root extracts (39%), however, it was not affected by the stem and leaf extracts.

For the species B. bipinnata, after incubation in distilled water for 17 days, 97% of seed germination was verified. In relation to the control, the final percentage of germination reduced by 36% and 15% for treatments with extracts of roots and leaves, respectively. The stem extract also reduced the percentage of seed germination, however, without significant effect. Differently from what was observed for B. pilosa var. radiata and B. bipinnata, the germination percentage of A. conyzoides seeds was not affected by any of the extracts was higher than in the leaves and stem.

Stem and leaf extracts reduced the overall growth of new tissues in B. pilosa var. radiata, but contributed to the elongation of hypocotyls and radicle. Both the stem and leaf extracts significantly stimulated hypocotyl elongation and inhibited radicle elongation. On the other hand, the root extract of B. pilosa did not affect the elongation of hypocotyls or radicles. The seedlings of B. bipinnata did not have the global growth affected by any type of extract, however, the hypocotyl elongation was stimulated by the leaf extract. In contrast, the growth of hypocotyls and roots of A. conyzoides was significantly stimulated by the three types of extracts of B. pilosa var. radiata evaluated. According to [54] such differential responses reveal that the aqueous extract of B. pilosa var. radiata was more harmful for seed germination and seedling development of B. pilosa and B. bipinnata than for A. conyzoides. This pattern indicates a high capacity of B. pilosa to compete with similar taxa, given the results found when the species is sympatric, as is the case of B. bipinnata.

Mao [55] described the allelopathic effect of the aqueous extract of B. pilosa, which, at concentrations of up to 20 mg/ml, had some facilitating effect on the growth of Trifolium repens and Medicago sativa pasture buds, however, at high concentrations (100 mg/ml or more) it showed considerable inhibitory effect on seed germination and seedling growth, noting, therefore, that the allelopathic inhibitory effects generally increase with increasing concentrations.

Lima [56] evaluated the effect of the ethanolic extract (0.5% w/v) of the aerial part of the species B. pilosa and B. alba on the growth of seedlings of L. sativa L. For this, a sheet of Whatman number 6 paper was placed in a 15 cm Petri dish and soaked with a 0.5% solution of extract, resulting in a final concentration of 5 mg of dry extract. Ethanol was used as a control treatment. After drying in an oven at 40°C for a period of 12 hours, and subsequent evaporation of the solvent, the filter papers corresponding to each species were placed in Gerbox boxes, on which 3 ml of distilled water were added and 25 seeds were distributed in each box. The boxes were kept under ideal conditions for germination (17°C, with minimum limits of 16.8°C and maximum of 17.2°C) and after 7 days, the radicle and hypocotyl length were measured and compared to the control treatment. Both species inhibited radicle and hypocotyl growth. The extract of B. pilosa promoted an inhibition of 47.29% on radicle growth and 60.63% on hypocotyl growth (60.63%), while B. alba inhibited radicle growth by 95.94% and hypocotyl in 56.50%.

The allelopathic potential described for B. pilosa is associated with the fact that this species presents a variety of secondary metabolites, among them, phenolic compounds, saponins, flavonoids, flavones glycosides, polyacetylenes, terpenes chalcone glycosides, phenylpropanoids glycosides, terpenoids [92, 93, 94], with emphasis on the group of polyacetylenes, phenolics, and terpenoids. Polyacetylenes are derived from natural hydrocarbons characterized by one or more acetylene groups in their structures and are produced predominantly in the roots of plants of the Asteraceae family [92].

Phytochemical studies on B. pilosa have reported the presence of alkaloids, flavonoids, triterpenes, acyl chalcones, polyacetylenes, 1-phenyl-1,3,5-heptatriino [95], phenolic compounds such as quercitin 3-O-rutinoside, phenolic acids such as chlorogenic acid and 3,4-di-O-caffeoylquinic, 3,5-di-O-caffeoylquinic, and 4,5-di-O-caffeoylquinic acids [96]. In this work, the authors attribute the allelopathic activity presented by the ethanolic extract of B. pilosa to the presence of polyacetylene and phenolic compounds. Some acetylenic compounds are toxic to a variety of organisms and harmful to plants [97], and the acetylenic compound phenylheptatriine (PHT) is considered an important allelochemical of B. pilosa [98], with potent allelopathic action. This is the most studied compound and its activity was first verified in leaves of B. pilosa [92]. Deba [86] and Priestap and Bennett [100] also identified and isolated PHT from B. pilosa oils using GC, GC–MS, and HPLC techniques, confirming that this is the main constituent in all parts of B. pilosa. Among the identified compounds, PHT showed the highest concentration (30–48%) in B. pilosa, followed by 1-phenylhept-5-ene-1,3-diyno (0.2–37.1%) and 7-phenylhept-2-ene-4,6-diinyl acetate (1.3–22.5%) [100] which may be involved in phytotoxic action. PHT was also verified in leaves of B. alba with a varying concentration in nature and in response to the photoenvironment [97]. Other polyacetylenes such as [cisdehydromatricariaester (cis-DMK), lachnophylum ester (LE), matricaria ester (ME), dehydromatricaria lactone, α-tertienyl, and thiophene polyacetylenes] isolated from higher plants are reported to have allelopathic action [98, 100].

Phenolics represent secondary metabolites with an allelopathic effect associated with the inhibition of seed germination and the establishment of plants in plant communities. The allelopathic activity of simple phenols (benzoic and cinnamic acid derivatives, flavonoids, and tannins) is well documented in the literature. GC–MS and HPLC analyzes indicated the presence of 15 phenolic compounds (salicylic acid, vanillin, phydroxybenzoic acid, caffeic acid, p-coumaric acid, and ferulic acid) in the leaves, stem, and root of B. pilosa [101], the main allelochemicals in nature according to Blum [102]. Deba [101] verified for all parts of the plant that the caffeic acid content was higher (117.4, 298.7, and 350.3 μg/g in leaves, stems, and roots) than pyrocatechin (18.5, 32.9, and 29.6 μg/g) and ferulic acid. Caffeic acid is one of the main allelochemicals present in the genus Leonurus [103] with proven allelopathic activity. Compounds such as p-hydrobenzoic acid, vanillic acid, ferulic acid, p-coumaric acid, and syringic acid are also considered the main allelochemicals in nature and can be found in both invasive weeds and species [102, 104, 105].

Substances such as phenolic acids, polyphenols, and flavonoids have allelochemical characteristics and can act as photosynthesis inhibitor herbicides, altering electron transport, and phosphorylation in photosystems. Furthermore, phenolic acids induce an increase in the activity of oxidative enzymes, having as a final consequence the modification of membrane permeability and the formation of lignin that contributes to the reduction of root elongation [106]. The results described by Lima [56] indicate that the two species analyzed can be used in the search for new herbicide molecules with less impact on agroecosystems and human health.

2.3.2 Essential oil

Studies point to the potential use of B. pilosa essential oil as an allelopathic, because it is in these secondary metabolites that terpenoids are present, whose natural functions are variable, including signal molecules, allelochemicals, phytoalexins, visual pheromones, pigments, photoprotective agents, member constituents, and reproductive hormones [4]. Terpenoids, including volatile terpenes, are the main components of essential oils, causing an allelopathic interaction between plants that have volatile allelochemicals [107].

Deba [101] and Priestap and Bennett [99] evaluated all the plant structures and the EOs (leaves, stems, roots, flowers, and oils), they reported that the oils contain from 60 to 114.6% of the total components detected. Most of the identified compounds (terpenes, thiophenes, and polyacetylene constituents) are referred to as allelochemicals, such as β-caryophyllene (0.10–7.50%) found in all parts of B. pilosa, limonene (0.2–2.12%), 4-terpineol (0.14–0.41%), β-linalool (0.09–0.43%), β-pinene (0.07–0.39%), α-pinene (0.2–5.97%), linalool (0.1–0.14%), sabinene (0.2–0.6%), and eugenol isobutyrate (0.5% at the root).

The studies by Cantonwine and Downum [97] and Zeng and Luo [108] compared the chemical composition of the essential oil of B. pilosa obtained from plants in Japan and Argentina and found that the main total components of β-copanene (11.2%), germacrene D (39.5%), 1-phenylhept-5-ene-1,3-diyno (27.0%), α-humulene (3.3%), and 1-phenylhepta-1,3,5-triine (78.9%) were not found in essential oil from Japan. On the contrary, the main components of B. pilosa from Japan, β-Bourbonene (2.09%), megastigmatrienone (7.39%), and diphenylenemethane (3.71%) were not detected in plants from Argentina. Very similar compounds of essential oils were found in plants from both areas, however, the percentage composition of essential oils (%) of plants from Japan was lower than that of plants from Argentina. According to the authors, it should be taken into account that the phytotoxic components of B. pilosa increase in drought conditions and the PHT compound vary significantly with geographic and seasonal factors.

The results found agree with those found by Ni [109] and Chen [110] when confirming the variations in the chemical components of Mikania micrantha, which is justified by the fact that the production and release of secondary substances by plants are highly influenced by the environment. Such a metabolic response suggests that plants, as a defense strategy, promote the release or increase of their secondary metabolites in order to adapt to the environment and geographic area in which they are inserted.

2.3.3 Vegetable residue

Hsueh et al. [57] confirmed the allelopathic effect of B. pilosa L. var. radiata Sch. Beep. on Cyperus rotundus L. To verify such activity, the authors evaluated, in a greenhouse, the allelopathic effect of residues of B. pilosa var. radiata on C. rotundus(1); interspecific competition between B. pilosa var. radiata and C. rotundus (2); the sprouting of tubercles of C. rotundus, in the field, in an area with or without removal of plant residues of B. pilosa var. radiata (3); and in addition to the effects of the use of B. pilosa var. radiata as a cover crop on the reproduction of C. rotundus (4).

The experiment 1 was carried out in pots under semi-natural conditions (greenhouse). The soil used was collected in an area of vegetable cultivation severely infested by C. rotundus, which was mixed with different proportions of residues of B. pilosa var. radiata, 0, 1.4, 2.8, and 4.2 g/pot, equivalent to 0, 0.1, 0.2, and 0.3 kg/m², respectively. Pre-sprouted tubers of C. rotundus were planted at densities of 3, 6, and 9 pre-sprouted tubers, corresponding to low, medium, and high density treatments, respectively. After 4 weeks, the plants were harvested, divided into shoots and roots and dried for 48 hours at 80°C. Inhibition of C. rotundus seedling growth was observed in the proportion of 0.1 kg/m in the density of 3 plants/pot. The dry mass of shoot, root, and total of C. rotundus were 72, 51, and 61% lower than the control (without B. pilosa residue-treatment 0 kg/m²), respectively.

The results of this experiment indicated that the phytotoxicity of B. pilosa residues to shoot and root (including the number of tubers) and seedling growth of C. rotundus was density dependent. The residues exhibited greater phytotoxicity for the seedlings at the lowest density (3 plants/pot) and the growth reduction decreased with increasing density, providing evidence for the existence of density-dependent phytotoxicity in the residue of B. pilosa. Weidenhammer [111] were the first to provide experimental evidence that the density-dependent phytotoxicity of allelochemicals can be used to distinguish allelopathy from intraspecific competition (or other microbial activities). Allelochemicals can cause a greater reduction in growth in the target plant at low density than at high density due to the dilution of the phytotoxicity of allelochemicals. Studies have indicated that leachate or allelopathic plant residues can inhibit the growth of C. rotundus, as verified by Babu [112] when testing the leachate from fresh leaves of E. globulus which significantly reduced the dry mass of C. rotundus root. [112]. Residues of Helianthus annuus L., Sorghum bicolor (L.) Moench and Brassica campestris L. reduced plant density and dry mass of C. rotundus [113, 114]. Due to the complexity of the ecological environment, it becomes difficult to distinguish allelopathy from leachate or waste from competition (including intraspecies or plant-microbial interactions) [115, 116].

The influence of B. pilosa residue on the shoot/root ratio of C. rotundus also showed a density-dependent phytotoxic effect, since the proportions decreased with increasing plant density for all investigated B. pilosa proportions. Ratio values greater than 1 indicate that root growth was more suppressed by B. pilosa residue than shoot growth, and the default biomass allocation was altered. According to the optimal partition theory (OPT), plants tend to partition more biomass in the root than in the shoot when there is a deficiency of the main nutrients and, thus, result in a lower proportion of the shoot/root ratio [117]. Williams [118] showed that C. rotundus allocated more biomass to the root instead of prioritizing high-density growth. The shoot/root ratio was inverse to the tendency for the application of B. pilosa residues in each density treatment, presuming that the nutrient absorption capacity of the C. rotundus root was impaired due to the deleterious effects of B. pilosa allelochemicals, such as phenolic acids, which resulted in an increase in shoot/root ratio [119, 120]. Also, according to Duke [121] plant growth can be stimulated at high density due to the phenomenon of hormesis (stimulation in subtoxic concentration), observed in the treatment with 9 plants/pot.

The relationship between soil characteristics and allelochemicals can affect the retention, transport, and transformation processes of allelochemicals in soil [122]. Batish [15] from experiments in pots found that although the content of soil organic matter, available nitrogen, pH, and EC were altered by the addition of allelochemical residues, the phytotoxicity of the residues of Chenopodium murale L. had negative effects on chickpeas and peas. In the field, Iqbal and Cheema [123] reported an improvement in cotton yield and a reduction in the population of C. rotundus when established in the area, intercropping allelopathic plants with cotton. However, few studies have investigated the simultaneous effect of interspecific competition and allelopathy, considering that in natural or semi-natural conditions the distinction of these processes is difficult.

For the interspecific competition tests, the modified protocol of Snaydon [124]. A plastic pot (13.5 cm in diameter and 13 cm in depth) was similarly subdivided by a plastic plate that served as an underground partition. The plastic plate was sealed with neutral silicone gel on the sides and bottom of the vessel. Another 30 × 30 cm plastic plate was fixed vertically to the upper edge of the pot, acting as a partition above the ground. This arrangement, with the arrangement of B. pilosa var. radiata and C. rotundus were designed to compare different competitions: NO competition, SHOOT competition, ROOT competition, and FULL competition. For each treatment, 1.2 kg of soil was mixed thoroughly with (AC treatment) or without (N treatment) fine powdered activated carbon (pure grade) in a 50:1 ratio, added to the two subdivisions of each pot. All pots were sprayed daily with water (50 ml per pot) and no fertilizer was added during the experiment.

From the relative arrangement of the subterranean partition associated with the disposition of B. pilosa and C. rotundus, four competition modes were obtained: (1) both the shoot and root of the two species were separated (NO competition), (2) only the root was separated (SHOOT competition), (3) only the shoot was separated (ROOT competition), and (4) neither the shoot. The findings of this experiment showed that under SHOOT competition, the growth of B. pilosa and C. rotundus was not suppressed with the addition of activated charcoal (AC treatment) compared to the AC treatment under NO competition. However, shoot and root growth of C. rotundus was reduced in the N treatment, presuming that this reduction under SHOOT competition may be a result of phytotoxicity in the leachate from the shoot of B. pilosa. In the ROOT and FULL competitions, the growth of C. rotundus increased, while B. pilosa decreased in the AC treatment compared to N. C. rotundus tends to express predominant root competition with coexisting plants, which was confirmed by Tuor and Froud-Williams [125] who demonstrated that this species can significantly reduce the dry mass of the aerial part and the height of corn and soybean when in conditions of complete competition in relation to situations without competition. Horowitz [126] indicated that citrus seedling growth was significantly inhibited by C. rotundus despite being fertilized with nitrogen. According to the author, the phytotoxic substances produced by C. rotundus may partially contribute to the competition with citrus. Although previous studies demonstrated that C. rotundus could compete with coexisting plants through allelopathy, the allelopathic effects of C. rotundus on B. pilosa were not observed, since the growth of C. rotundus was suppressed by B. pilosa when activated carbon was not applied.

C. rotundus shoot/root ratio responses to the allelopathic effects of B. pilosa roots did not increase under the influence of B. pilosa residues, as observed in experiment 1. This can be explained in two ways. The first one establishes that the allelochemicals in the residues and the exudates of the root can be different. Deba [101] found that the main phenolic compounds in the leaves, stems, and roots of B. pilosa may be similar in composition but different in content. And the second considers that allelopathy combined with root competition promoted a different inhibition mechanism in this experiment.

El-Rokiek [127] pointed out that phenolics (ferulic acid, caffeic acid, for example) present in mango leaves can inhibit seedling growth and tuber sprouting of C. rotundus. Fifteen phenolic compounds isolated from B. pilosa have also been reported to possess the allelochemicals of phenolic compounds (e.g., caffeic, ferulic, p-coumaric, p-hydroxybenzoic, salicylic acid, among others) [60, 128]. Such phenolics have been reported to cause deleterious damage to plant roots. In this regard, Einhellig [129] in his studies indicated that salicylic acid seems to cause damage to the membrane structure and permeability of root cells, while caffeic acid decreases the levels of nitrogen, phosphorus, potassium, iron and molybdenum in cowpea. Ferulic acid was responsible for inhibiting nitrogen uptake in the roots of corn seedlings [119], while transcinnamic, ferulic, and p-coumaric acids reduced net nitrogen uptake and plasma membrane H⁺-ATPase activity [130]. Such activity is possibly associated, as presented elsewhere, with PHT, a supposed allelochemical of polyacetylene, which supposedly releases phytotoxic radicals acting as an inhibitory mechanism. The presence of PHT has already been verified in leaves of B. pilosa, with reports of suppression of the growth of seedlings of A. syriaca, C. album, P. pratense, and T. pratense with LC50 of 0.66, 0.83, 2.88, and 1.43 ppm, respectively [99]. The inhibitory effect was also correlated with nutrient deficiency conditions [122], in which plants with allelopathic effect exhibited greater inhibitory effects on neighboring plants, and contributed to stimulate the allelochemical exudation of B. pilosa by competing for nutrients with C. rotundus (under ROOT and FULL competitions).

In experiment 3, tuber sprouting of C. rotundus in the field was evaluated. In order to prevent the interference of invasive C. rotundus plants, this was carried out in a three-year strip and vegetation (24 m long and 1.5 m wide) containing residues of B. pilosa var. radiata. In this range, eight 30 cm × 30 cm blocks were randomly defined. In half of them, the shoots and litter were removed in order to keep the soil surface uncovered (VN), and in the other, they were kept (VS). On adjacent land, four 1 m × 1 m plots were randomly selected in which weeds, including C. rotundus tubers, were removed by hand weeding. In the center of each plot (30 cm × 30 cm) an opaque plastic sheet was placed. This treatment was conducted in order to compare tuber sprouting with the treatments previously described. Twenty-five tubers (0.5–1.0 cm in diameter) of C. rotundus were planted in each block.

The highest percentage of tuber sprouting, sprouts per block, and dry mass per sprout occurred in the treatment with opaque plastic sheet cover, followed by the area with tubercles and the presence of B. pilosa vegetation, and the area without sprouts and litter. The percentage of tuber sprouting (52%), average sprouts per block (18 sprouts per block) and dry mass per sprout (5.26 mg) reduced when C. rotundus tubers were planted in areas containing B. pilosa. The clean treatment, without the buds and litter of B. pilosa, indicated that even in the absence of these, the allelochemicals remained in the soil and continued to inhibit the sprouting of C. rotundus tubers.

B. pilosa exhibited a strong phytotoxic effect on C. rotundus tuber reproduction. The percentage of tuber sprouting, average number of sprouts per block, and dry mass per sprout were lower in treatments with and without removal of sprouts and litter than in treatments with opaque plastic. Scavo [122] highlights that in natural environments, the concentration, movement, and persistence of allelochemicals have the power to influence the phytotoxicity of potential donor plants on the target species. Water-soluble allelochemicals such as phenolics have been reported to have a short residence time in soil due to rapid leaching and degradation [128, 129].

The founds of Hsueh et al. [57] indicated that the use of land without cover, such as the VN treatment, blocked the entry of allelochemicals, such as phenolics and PHT, into the soil. It is estimated that the phytotoxic effect of B. pilosa remaining in the soil is degraded within a brief period, when the release of allelochemicals from the plant surface and litter to the evaluated block ceases. Off-block replenishment of B. pilosa allelochemicals was also limited due to low rates of chemical diffusion in the soil, sorption of soil particles and organic matter, and microbial degradation [131].

Pre-cutting of C. rotundus plants before the beginning of the experiment predicted a break in the apical dominance of the tubers [132, 133], however, there was a stimulus for the sprouting of dormant tubers. Additionally, the fact of belonging to the C4 photosynthetic pathway plant group [134] the C. rotundus shoots grew faster than the B. pilosa seedlings. Plant residues from sorghum stalks (15 ton/ha) incorporated into the soil had a lower inhibitory effect on the density of C. rotundus than surface-applied residue at the same concentration as Mahmood and Cheema [135]. Later, Khaliq [113] tested the combination of sorghum, sunflower and brassica residues (7.5 ton/ha each) reporting a reduction in C. rotundus plant density by 87% compared to the control. Boz [135] reported a decrease in the density of P. oleracea L., Amaranthus retroflexus L., and Echinochloa colonum L. when using wheat and rye straw, however, they did not show an inhibitory effect on C. rotundus.

The effects of vegetation and coverage with B. pilosa var. radiate residues were also evaluated for the reproduction of C. rotundus. For this, a field infested by C. rotundus was divided into 24 plots (3 m long and 1 m wide), each one separated by a 0.8 m wide ditch. Soil samples were collected for each plot in order to investigate the numbers and dry mass of C. rotundus tubers in the top 15 cm of soil. All plots were mowed and the seeds of B. pilosa (8 g/polt) were randomly sown in half of the plots. In another 12 plots, B. pilosa was not sowed. At 69 days after sowing, all above-ground parts of the plant were cut, weighed, and kept in the plot, and tuber counts and dry mass were weighed. Half of the B. pilosa and C. rotundus residue plots were covered with an opaque plastic sheet while the other half was not covered.

In the studies of Hsueh et al. [57], although the tubers proliferated before the cover crops were cut, the density of C. rotundus plants was significantly reduced in the treatment with B. pilosa residue, and decreased with increasing dry mass of the residue. The density of C. rotundus plants was higher in both species of cover crops in the opaque plastic treatment. No negative effect has been reported associated with the increase in temperature promoted by the presence of opaque plastic [136], on the reproduction of tubers, provided that this does not exceed a temperature of 45°C for a period of more than 7 hours per day [137]. Despite this, according to Campbell [98] and Stevens [138] the phytotoxicity of polyacetylenes such as PHT, due to the characteristics of photosensitization and high activity, can be reduced in the dark and at temperatures above 30°C. Thus, it is possible that the allelochemicals of B. pilosa are degraded more quickly in treatments with opaque plastic covering, both for B. pilosa and for C. rotundus due to the increase in temperature under the leaf.

According to Xuan [139] there is a strong correlation between the ability to compete and invasive capacity of a culture or native species with its allelopathic potential. Emphasis is given to B. pilosa, with recognized phytotoxic effect on its sympatric species, such as B. bipinnata L. and Pteris multifida Poir., in ecosystems [54, 140]. B. pilosa and B. pilosa var. radiata have been investigated in Southeast Asia for their ability to manage weeds. Hong [141] evaluated 10 allelopathic species for weed control in Vietnam. In this study, after the application of 2 t/ha of B. pilosa in the area and a reduction of 84.9% and 81.8% was verified for the density and dry mass of plants, respectively. The application of B. pilosa biomass also showed greater weed suppression than manual weeding and herbicide treatments stimulating the height of rice plants in relation to the control (without plant residue) and contributing with tiller numbers and panicle formation of rice grains (11.8–16.9%). in relation to herbicide treatment (5.6–7.8%). Furthermore, the introduction of B. pilosa plant material exceeded rice yield by more than 20% when compared to herbicide treatment and was similar to hand weeding. The data obtained allow us to conclude that the application of B. pilosa biomass in the rice field significantly suppresses the infestation and increases the rice yield. The weed reduction may have occurred due to the release of phytotoxins (allelochemicals) released from the decomposing biomass of B. pilosa in the soil. The allelopathic activity of B. pilosa on rice growth and yield was stimulated by the presence of allelochemical compounds and nutritional effects of B. pilosa, which generally contain nutrients (P, K, Ca, and Mg), which improved growth and rice productivity [141].

Krumsri [142] also examined the phytotoxic effects of B. pilosa residue on E. crus-galli with an increase in this when applied in the fresh form, in relation to the dry residue. In the same study, the authors found that both soil cover and incorporation with B. pilosa residues reduced the density of E. crus-galli when using plants harvested at 60 days of growth. Poonpaiboonpipat and Poolkum [143] reinforced the promising results of the B. pilosa var. radiata use in paddy field (4 ton/ha) in inhibiting weed growth (86.73%) and increasing productivity by 81.03%. Khanh [87] evaluated the effect of root exudates of B. pilosa during its initial growth stage (20-day exudates in agar culture) which promoted 70% inhibition on germination, root length and shoots of Leucaena leucocephala, and 50% inhibition in E. crus-galli, M. sativa, and O. sativa. B. pilosa was not inhibited. The results of these studies support the potential use of residues of B. pilosa and B. pilosa var. radiata, as well as the extract and essential oil, as natural herbicides in weed management in agroecosystems.

3. Conclusions

The potential of the Asteraceae family as a repository of species with phytotoxic or allelopathic activity is evident. Although the number of studies that associate allelopathy with metabolites present in the crude extract, essential oil, or in plant biomass residues is increasing and presents promising results, studies are still needed to prove the interaction of these compounds in the soil. According to Zhang [144] among the four pathways by which allelochemicals are released into the environment, plant residues exert the most negative effect. Still, the importance of allelopathy in nature requires further investigation, as allelopathic effects were smaller in studies where soil microorganisms were likely to be abundant and when the study duration was longer. For this, two points should be considered in future studies to confirm allelopathy. The first one is the identification of one or more phytotoxins produced by the plant under investigation, or the identification of a compound(s) that can be converted into a phytotoxin in the soil, after its release. The second refers to the quantitative determination of the compound(s), that is, if they are found in sufficient quantity (in time and/or space) in the soil in which the plant grows or grew, and if in these natural conditions can affect other species. This stage is especially difficult and challenging because, in the natural environment, plants are exposed to different interactions, effects of antagonism and synergism with other compounds in the soil, growth stages and physiological factors, state of the recipient plant, soil microbiota (especially with rhizosphere microorganisms), soil moisture, temperature, among others. Zhang [144] after a systematic review found that the effects of allelopathic were weaker between closely related species (or individuals of the same species) than between distant species, suggesting that allelopathy would favor the coexistence of closely related species, the opposite of the predicted effect in competition for resources. These notes further reinforce the need to investigate allelopathy and competition for resources together in order to explain the results of species coexistence.

Acknowledgments

The authors thank Universidade Paranaense and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brazil (CAPES) for the financial support and the fellowship.

Conflict of interest

The authors declare no conflict of interest.