Phytotoxic and cyto-genotoxic activity of essential oil from leaf residues of Eucalyptus urophylla and the hybrid E. urophylla x E. camaldulensis on Lactuca sativa and Sorghum bicolor

The use of agrochemicals has influenced the increase in agricultural productivity. However, the concern about damage to the human health and to the environment, resulting from the indiscriminate use of pesticides, has increased worldwide. Alternative methods for controlling pests and diseases have been proposed to maintain productivity and quality of life. A possibility is to use compounds produced by the secondary metabolism of plants, such as the essential oils. Some of these substances perform inhibitory or stimulatory activity on the development of other organisms. Plants from the genus Eucalyptus have been investigated and were chosen to be studied in the present work, due to the traditional knowledge regarding the potential of the essential oils from some species, as well as, for the need to use residues from their production. Therefore, were determined the yield, identified the compounds from the essential oils of E. urophylla and the hybrid E. urophylla x E. camaldulensis, and evaluated their biological activity through bioassays, investigating the phyto-cyto-genotoxicity and mutagenicity using Lactuca sativa and Sorghum bicolor as model plants. The essential oil from the hybrid provided a higher yield. Eucalyptol was the major compound identified for both oils, representing more than 85% of the compounds present. Both E. urophylla and the hybrid showed phyto-cyto-genotoxic, mutagenic effects, and clastogenic and aneugenic mechanisms of action, Research, Society and Development, v. 10, n. 11, e242101119646, 2021 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v10i11.19646 2 promoting epigenetic changes in the meristematic cells of L. sativa. The results point out to the bioherbicidal potential of these essential oils.


Introduction
Technological advances have led to high agricultural productivity, and to lower production costs (Alves and Tedesco, 2015). However, the technologies achieved have generated discussions about their consequences for human health and to the environment Alves et al., 2018;Pinheiro et al., 2015).
Debates have been held regarding the contamination of rivers, groundwater, soils, surrounding cultures, lakes and air (Flores et al., 2004;Silva et al., 2012). We must also consider problems with the aquatic and terrestrial fauna (Flores et al., 2004;Silva et al., 2012). Several cases of intoxication and damage to human health resulting from contact with agrochemicals (either by ingestion or skin contact), have generated the dilemma regarding the benefits and harms of these substances (Carson, 2002;Hendges et al., 2019;Santos et al., 2019).
To maintain an elevated and outstanding productivity and to mitigate the harmful effects of agrochemicals, natural products with biological activity have been evaluated in studies that aim at producing bio agrochemicals. The essential oils from plants with medicinal effects and/or food use are detached among several prominent compounds in this type of research Pinheiro et al., 2015;Vasconcelos et al., 2019).
Species from the genus Eucalyptus are known for their essential oils, which serve as the basis of medicinal solutions with anti-inflammatory and antibacterial action and in the treatment of respiratory diseases (Mallard et al., 2018). Studies have confirmed antimicrobial (Dhaliwal et al., 2004;Falahati et al., 2005;Moreira et al., 2005;Oluma and Garba, 2004), insecticide (Erler et al., 2006), antibacterial (Ramezani et al., 2002), phytotoxic and cytotoxic  activities. Within this genus, the species E. urophylla and the hybrid E. urophylla x E. camaldulensis are cultivated on a large scale aiming (with their stem) at meeting the needs of the pulp, coal and wood industries (Oliveira, 1998). However, their roots and leaves are residues that are discarded in the production process and there is no study reporting the influence of chemical composition and the action of essential oils of these species on model plants.
Another important fact concerns the planting of eucalyptus trees in India, which are cultivated as part of the local government's social forestry program. These trees are planted on abandoned lands and on the margins of state and national roads. However, dry leaves fall and, depending on the number of individuals found in a certain area, especially in the dry period of the year, they can cause forest fires (Aich et al., 2019). The use of leaf residues from eucalyptus trees for the extraction of essential oils can both increase their economic value and reduce the risk of forest fires.
Researchers have been carrying out bioassays to assess the biological activity, as well as, to elucidate bioactive compounds Alves et al., 2018;Pinheiro et al., 2015). The phyto-cyto-genotoxicity and mutagenicity analysis, besides the determination of mechanisms of action are types of tests performed to determine the herbicidal potential of the studied compounds. These tests present clear and efficient results in a short period of time, use model organisms (both mono and eudicotyledonous), present low cost and use, as test organisms, plants that show results correlated with those found in mammals dos Santos et al., 2018;Silveira et al., 2017).
Given the above, the objective was to evaluate the chemical composition and the biological activity of the essential oils of E. urophylla and the hybrid E. urophylla x E. camaldulensis. The tests were carried out with the plant models Lactuca sativa L. and Sorghum bicolor (L.) Moench, through phytotoxicity and cytotoxicity assessments. Also assessed the similarity of the variables between the two species, since studied a hybrid and its parent.

Plant Material
Leaves from adult clones of E. urophylla (parental) and the hybrid E. urophylla X E. camaldulensis were harvested in the forest garden of the Center of Agricultural Sciences and Engineering of the Federal University of Espírito Santo, located at Highway ES 482, Km 43, coordinates 20°47'43.7"S 41°24'20.9"W. For the biological effect analysis, seeds of L. sativa (lettuce) "Crespa Grand Rapids -TBR" (ISLA) and S. bicolor (sorghum) "precious AL" (Br seeds) were purchased from local stores.

Essential Oil
For the oil extraction, the leaves were weighed, crushed with distilled water and placed into the Clevenger apparatus.
Subsequently, the oil obtained was weighed and its compounds were identified by gas chromatography with a mass spectrometer (Mendes et al., 2018). Helium was used as a carrier gas in a capillary column of fused silica Rtx-5MS (30 m long, 0.25 mm internal diameter). The temperatures were 220°C for the injector and 300°C for the detector, being the 60°C the initial column temperature, programmed for an increase of 3°C per minute until reaching 240°C, the maximum temperature.
The obtained mass spectra was compared with reference data from the equipment database. This procedure was performed separately for the two studied species. Research, Society andDevelopment, v. 10, n. 11, e242101119646, 2021 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v10i11.19646 For component quantification, the essential oils were analyzed on a gas chromatograph equipped with a flame ionization detector (GC-FID), using nitrogen as carrier gas a the stationary phase was the capillary column Rtx-5MS (30 m long and 0.25 mm internal diameter).
The yield of each essential oil was expressed in percentage. It was calculated through the comparison of the quantity (g) of leaves used to obtain the oil with the quantity (g) of oil obtained.

Phytotoxicity Assay
To perform the phytotoxicity test, five concentrations (3000, 1500, 750, 375 and 187.5 μg L -1 ) of each essential oil obtained were analyzed. A completely randomized design was used with four replications per treatment; each replication consisted of 25 seeds. The analyses were carried out with lettuce and sorghum. Distilled water and dichloromethane (solvent used in the preparation of the concentrates) were used as negative controls (C-), and glyphosate, commercial herbicide, was used as a positive control (C+), at the concentration indicated by the manufacturer, 0.1%.
Evaluated the germination speed index (GSI) of the seeds following the number of seeds germinated per plate every 8 hours during the first 48 hours of exposure to the treatments. The germination percentage (GP) and the percentage of oxidized roots (O) were evaluated based on the germinated seeds/oxidized roots per plate after 48h of exposure to the treatments. Root (RG) and shoot (SG) growths were measured after 48h and 120h of exposure to the treatments, respectively .

Cyto-Genotoxic and Mutagenic Assay
The tests were performed on lettuce roots after 48 hours of exposure to the treatments, with the same experimental design described on item 2.3. Only lettuce was used, since it presents large chromosomes in small quantities, easy manipulation of roots and preparation of slides and because it has a correlation with the response in the cell cycle of other eukaryotes (Silveira et al., 2017).
For the cellular evaluation, roots were collected and fixed in ethyl alcohol:acetic acid (3:1). After 24 hours of fixation, the slides were prepared using 2% of acetic orcein as dye. Approximately 1000 cells were evaluated in each slide. We classified them according to the phase of the mitotic cycle and regarding the cellular damage presented.
To assess cytotoxicity, the mitotic index (MI) was calculated by dividing the number of cells in division per the total number of cells evaluated. The genotoxic effect was assessed based on the amount of observed chromosomal alterations (CA).
CA were calculated from the ratio between the number of cells with CA and the total number of cells evaluated, as well as, by the frequencies of each observed CA, which are calculated by the ratio of the quantity of each CA to the total cells in division observed. The mutagenic effect was measured from the nuclear alterations (NA), calculated from the ratio between the number of cells with NA and the total number of cells evaluated, as well as, by the frequencies of each observed NA, calculated using the ratio between the quantity of each NA and the total number of cells evaluated .

Statistical Analysis
The results were subjected to analysis of variance and the means were compared by the Dunnett's test (p <0.05), the most suitable test for comparing treatments with controls. All data were processed with the GENES statistical program (Cruz, 2013).

Quantification and Characterization of Essential Oils
The essential oil of the hybrid showed a yield of 0.77% m/m, while E. urophylla presented 0.20% m/m per fresh leaf fraction. The hybrids have a high genetic potential for production, because this technique allows exploring heterosis (superiority of the crossing in relation to either two parents), also called hybrid vigor. Cimanga et al. (2002) studied the essential oils of different species of eucalyptus and obtained a yield of 0.53% m/m from the essential oil of E. urophylla, using the same extraction method and comparing it with the weight of fresh leaves. The variation observed may be due to environmental conditions, plant and leaf age as well as the genetic factors (Xavier et al., 1993). Cimanga et al. (2002) also studied the essential oil of E. camaldulensis, the other parent of the studied hybrid. The data obtained showed lower yield of this parent (0.30% m/m of yield) when compared to E. urophylla, which presented 0.53% m/m. The individuals from the current research were harvested in the same place (same environmental conditions) and followed the same pattern of size and age of individuals and leaves (minimizing the physiological differences). Can infer that the yield of the hybrid (0.77% m/m), because it is higher than that of the parents', results from the additive effect of the combination of parental genes, determining a higher yield in the hybrid oil.
Regarding the chemical characterization, were identified four compounds in each oil (four peaks appeared for each oil). The major compound in the essential oil of E. urophylla was the oxygenated monoterpene eucalyptol (87.9%), followed by the oxygenated sesquiterpene viridiflorol (5.1%), the hydrocarbon monoterpene α-pinene (4.3%) and the hydrocarbon sesquiterpene aromadendrene (2.6%) ( Table 1). Eucalyptol was also the major compound identified in the hybrid (89.8%), followed by the oxygenated monoterpene terpineol (6.73%), hydrocarbon monoterpene α-pinene (1.9%) and the oxygenated monoterpene linalool (1.4%) ( Table 1). Eucalyptol has been described in the literature as the major compound of both the essential oils of E. urophylla and E.
camaldulensis (Cimanga et al., 2002;Pereira, 2010), which was also observed in the oil of the hybrid studied in the present research. In addition, the presence of α-pinene, eucalyptol and terpineol in both parents has already been described in the literature, while the presence of linalool was observed only in the parental E. urophylla (Cimanga et al., 2002;Pereira, 2010).
The hydrocarbon monoterpene α-pinene is present in both parents. According to Pereira (2010), this compound represented 1.7% of the oil from E. camaldulensis and 5.0% of the oil from E. urophylla. Thus, the percentage of α-pinene identified in E. urophylla in the present study, is similar to that found by that author. The percentage of α-pinene identified in the hybrid in the present study, is more similar to that found in the parental E. camaldulensis by Pereira (2010). Linalool has been described as part of the composition of the oil from E. urophylla, representing 2.5% of the total oil composition (Cimanga et al., 2002). Thus, there is a decrease in the proportion observed in the hybrid, when compared to the parental.
Other authors have already reported terpineol as part of the essential oils of the parental species, however, as explained previously, in lower proportions than that found in the hybrid in the present study, indicating the presence of a possible additive effect in its production.

Phytotoxicity Assay
Phytotoxicity analyses demonstrated the toxic and allelopathic effects of both the tested oils. GP was significantly inhibited, differing even from C+, in lettuce seeds treated with the hybrid oil at a concentration of 3000μg L -1 , with a 17% inhibition compared to C- (Figure 1a). This is the low-sensitive parameter, which is only expressed in the acute phytotoxicity of the test compounds (Aragão et al., 2017;Costa et al., 2019). Therefore, as it is not very sensitive, this variable will be changed when the test agent is very toxic, that is, when it produces acute toxicity. Thus, the reduction of this variable demonstrated the strong bioherbicidal potential of the hybrid's essential oil. Research, Society andDevelopment, v. 10, n. 11, e242101119646, 2021 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v10i11.19646 8 All the treatments showed GSI means lower than those of C-, showing statistical difference in the treatments of the hybrid at the concentration of 3000μg L -1 in lettuce and sorghum, and at the concentration of 1500 μg L -1 in lettuce, equal to C+ (Figure 1). According to Pinheiro et al. (2015), this is the most sensitive parameter among those evaluated in the phytotoxicity tests; its inhibition indicates the toxicity of the test agent regardless of the results observed in the other measured parameters. The delay in the germination period represents a competitive disadvantage for the individuals in the field, as they will be submitted to a longer period of exposure to pathogens, abiotic stress and predators. The prolongation in the germination time determines competitive losses with pre-established plants in growth, making them more susceptible (Uhlmann et al., 2018).
The two oils tested promoted oxidation (O) in the lettuce roots, with no variation in sorghum (Figure 1). All the oil treatments caused an increase in O in lettuce, which was significant in the treatments with the essential oils of E. urophylla at the concentrations of 3000, 1500 and 750μg L -1 , 56, 29 and 33 times, respectively, when compared with C-, this increase was higher than that promoted by C+ (Figure 1a). This oxidative phenomenon, expressed from the darkening of the roots, has been related to the production, oxidation and release of phenolic compounds, which inhibit the individual's growth and development (Melo et al., 2001).
The hybrid's oil (3000μg L -1 ) caused significant inhibition in the RG of both model species, comparing to C-and equaling to C+, with 48% in lettuce and 60% in sorghum, when compared with C- (Figure 1 and 2). The essential oil of Eucalyptus urophylla inhibited the root development of sorghum by 60% and 50%, at the concentrations of 3000 and 1500μg L -1 , respectively, when compared to C- (Figure 1b). This result may be associated with the oxidative toxic effect described above, which promoted a decrease in the RG of the seedlings. Aragão et al. (2017) described this variable as the most sensitive among those related to growth. The essential oil of the hybrid did not promote significant inhibition in the SG of the model plants ( Figure 1). However, the essential oil of E. urophylla inhibited lettuce's SG by 60%, at the concentration of 3000 μg L -1 , which was equal to C+ (Figure 1a). This result may be related to the chemical composition of the essential oils, as well as to the synergistic effect between the compounds. Aragão et al. (2015), verified the allelopathic activity of E. grandis and E. citriodora, and reported the synergistic effect of the essential oil of E. grandis, which presents α-pinene as its major compound, providing greater phytotoxic effect. When we compare these results with ours, conclude that α-pinene, which is more abundant in E. urophylla than in the hybrid, provided greater phytotoxicity, when in synergism with the other compounds of the oil, providing greater inhibition in the SG of seedlings treated with E. urophylla, than with its hybrid.

Cyto-Genotoxic and Mutagenic Assay
The essential oil of E. urophylla, promoted a significant increase of 1.5 and 3%, at the concentrations of 3000 and 187.5μg L -1 , respectively, in the frequency of cells in interphase (Figure 3). The increase in cells in interphase indicates the occurrence of a mitosis block Barroso Aragão et al., 2017), which may be related to the inhibition of DNA synthesis due to the increased damage to this molecule, aiming at minimizing tissue damage (Kordali et al., 2008). Aragão et al. (2015) also observed this result in treatments with the essential oils of E. grandis and E. citriodora. Figure 3. Percentage of each cell cycle phase of lettuce root meristems treated with the essential oils of E. urophylla and the hybrid E. urophylla X E. camaldulensis at the concentrations of 3000, 1500, 750, 375 and 187.5μg L -1 . Means followed by letter a were equal to C-(distilled water), means followed by letter b were equal to C-(dichloromethane-DCM) and means followed by letter c were equal to C+ (glyphosate), according to Dunnett's test (p <0.05).
The essential oil of E. urophylla caused a decrease in the percentage of prophase and metaphase in 31% and 21%, respectively, at the lowest concentration (187.5μg L -1 ) (Figure 3). This reinforces the previous result and may be related to the increased oxidation of the roots treated with this oil (Figure 1), since this phenomenon is related to the release of phenolic compounds, which are toxic to the plant (Aragão et al., 2017). In addition, the essential oil of E. urophylla resulted in an increase of 24% in the frequency of metaphases at the concentration of 750μg L -1 (Figure 3). This is related to the increase in chromosomal adherence, which alters the pattern of the phases of mitotic division and determines the cell permanence in metaphase (Aragão et al., 2017;Alves et al., 2018;dos Santos et al., 2018).
The most cytotoxic treatments were those of the essential oil of E. urophylla at the concentrations of 3000 and 187.5μg L -1 , which significantly inhibited MI by 13 and 23%, respectively (Figure 4). This parameter is a well-established endpoint as an indicator of toxic agents. Under stress conditions, cells will suffer a reduction in the number of cells in division, aiming at minimizing the contact with the toxic substance Aragão et al., 2017;Costa et al., 2017;Alves et al., 2018;Leme and Marin-Morales, 2009;Pinheiro et al., 2015). All the treatments were genotoxic, promoting a significant increase in CA, when compared to C-and C+. The treatments with E. urophylla were increased 9.4; 10.3; 11; 7.96; 4.15 times, at the concentrations of 3000, 1500, 750, 375 and 187.5μg L -1 , respectively (Figure 4). For the treatments with the hybrid's oil at the concentrations of 3000, 1500, 750, 375 and 187.5μg L -1 this increase was of 7.8; 8; 7.96; 6.27 and 7.15, respectively (Figure 4). CA are caused by the interaction between the genetic material of the cell and the chemical compound present in the environment, what is a genotoxicity endpoint (Bernardes et al., 2015).
The essential oil of E. urophylla presents greater mutagenic potential than the hybrid's oil. Both promoted an increase in the frequency of NA, however, the essential oil of E. urophylla, promoted a significant increase of 15.7; 8.5; 8.5 and 6.71 times, at the concentrations of 3000, 1500, 750, and 187.5μg L -1 , respectively. While the essential oil of the hybrid provided a significant increase of 5.5 and 11 times, at the concentrations of 3000, 1500μg L -1 , respectively, when compared to C-( Figure   4). NA are morphological alterations, which occur as a result of biochemical changes in the cell nucleus, as a defense Research, Society andDevelopment, v. 10, n. 11, e242101119646, 2021 (CC BY 4.0) | ISSN 2525-3409 | DOI: http://dx.doi.org/10.33448/rsd-v10i11.19646 11 mechanism for possible nuclear "errors" arising from interaction with the environment (by Alves et al., 2018;dos Santos et al., 2018;Fernandes et al., 2009).
The observed NA were micronucleus (MNC) and condensed nucleus (CN) (Figure 5a and 5b). CN was more frequent contributing on a larger scale in the total of NA (Figure 4). The MNC comes from chromosomes that are "lost" during cell division (not properly incorporated into the nucleus of the daughter cell) or it is formed by fragments of acentric chromosomes (also not incorporated into the nucleus of the daughter cells). Thus, these MNCs have the function to involve and eliminate these extra/loose portions of DNA from the cell cytoplasm (dos Fernandes et al., 2009). The cells treated with the essential oil of E. urophylla at the concentrations of 3000, 1500 and 750μg L -1 , presented a frequency of CN significantly higher than the frequency of C-, 14.71, 7.57 and 7.14 times, respectively. Whereas, the hybrid's oil promoted a significant increase of 12.8 and 10.4 times, at the concentrations of 3000 and 1500μg L -1 , respectively ( Figure   4). CN is the cytological expression of the occurrence of cell death (Andrade-Vieira et al., 2011;Costa et al., 2017). This mechanism has the main function of maintaining tissue homeostasis, in addition to eliminating cells that could trigger malignant processes in the body, as they contain changes in DNA (Silva et al., 2017). This variable is increased according to the increase in genetic damage (Kordali et al., 2008).
The CA observed were: c-metaphase (Figure 5c The aneugenic mechanism of action is promoted due to changes in the mitotic machinery of the cell, promoting changes in the mitotic spindle, involving the polymerization and depolymerization of the microtubules. Meanwhile, clastogenic agents interact with the genetic material of the cell, causing DNA damage (Bernardes et al., 2015;dos Santos et al., 2018;Fernandes et al., 2009). There was an increase in the frequency of chromosomes lost in all the treatments analyzed, with significance in treatments with the essential oil of E. urophylla at the 3 highest concentrations, 13.9, 10.94 and 13.6 times, respectively, which were equal to C+ (Table 2).
Observed multipolarity, an aneugenic alteration, in cells treated with the essential oil of E. urophylla at concentrations of 1500 and 375μg L -1 , with no significant increase (Table 2). This CA is related to the aneugenic action, since it results from the formation of multiple nucleation sites, which can be caused by the genetic imbalance of the cell, occurring in polyploid cells (Fernandes et al., 2009).
Chromosomal adherence was the most frequently observed alteration, with an increment in all the treatments. It showed a significant increase of 6.45; 6.94; 7.61 and 6.04 times in the treatments with E. urophylla at the concentrations of 3000, 1500, 750 and 375μg L -1 , as well as, 7.7; 6.22 and 6.08 times in cells treated with the hybrid's oil at the concentrations of 1500, 750 and 187.5μg L -1 , respectively (Table 2). Chromosomal adherence occurs due to cytological, genetic and epigenetic changes. The latter is indicated, since such alterations comes from changes in the phosphorylation pattern of histones, suggesting both aneugenic and clastogenic action (dos Freitas et al., 2016;Silveira et al., 2017).
C-metaphase was very frequent and presented an increase in all the treatments. Nevertheless, it did not show a statistical difference from C-, due to the low number of cells in C+ division, making the frequency of c-metaphases (which is obtained by the reason of the number of cells with c-metaphase observed by the total number of dividing cells) high in this treatment ( Table 2). The c-metaphases demonstrate aneugenic action of the test agent, since they express the complete inactivation of the microtubules and consequently the formation of the mitotic spindle dos Santos et al., 2018).
Another alteration that presented (non-significant) increase in all the treatments was the delay of chromosomes in telophase (Table 2). This alteration is related to the interference of the essential oils tested in the depolymerization of the microtubules, resulting in an uneven dragging of the chromosomes. Such alteration causes the cell to reconstitute a nuclear envelope with deformation, aiming at involving all the genetic material applicable to it and later that deformation is undone (Fernandes et al., 2009).
The frequency of chromosomal bridge was significantly increased in all of the treatments, 16 times, when compared to C-, in cells treated with the oil of E. urophylla at the concentration of 1500μg L -1 . A similar fact was observed in chromosomal fragmentation, which had a significant increase in the same treatment ( Table 2). The increase in the frequency of bridges and chromosomal fragments indicates the clastogenic action of the test agent (Fernandes et al., 2009). This result suggests the occurrence of the break-fusion-break cycle, where chromosomal fragmentation occurs leaving free cohesive ends in the chromosomes, which causes the fusion (bridge) of chromosomes. When these chromosomes are dragged by their centromeres to the cell poles, through the depolymerization of the alpha and beta tubulin filaments, the chromosome breaks again, forming new chromosomal fragments, enabling new fusion and future breaks (Silveira et al., 2017).
Elucidative experiments encompassing phyto-cyto-genotoxicity, mutagenicity and the mechanism of action of natural compounds is important to elucidate the best use of plant material/waste, as well as to minimize environmental damage resulting from forest fires. Means followed by letter a were equal to C-distilled water, means followed by letter b were equal to C-dichloromethane (DCM) and means followed by letter c were equal to C+ glyphosate, according to Dunnett's test (p <0.05). Source: Authors.

Conclusion
The yield and characterization of the essential oils of E. urophylla and its hybrid E. urophylla X E. camaldulensis indicated a possible additive effect of the inherited genes for such characteristics. The yield of the essential oil of the hybrid was superior to that of the parents.
Both E. urophylla and the hybrid showed phyto-cyto-genotoxic and mutagenic activities, as well as clastogenic and aneugenic mechanisms of action. In addition, they promoted epigenetic changes in the meristematic cells of lettuce, identified from the increase in chromosomal adherence. This was the first report of the action mechanism of essential oils of the E.
urophylla and E. urophylla X E. camaldulensis in model plant cells.
The results point out to the bioherbicidal potential of these oils, raising the possibility that the leaf residues of these species are destined for the extraction of essential oil, aiming at its application in the control of weeds.