Background
Leishmaniasis, the third most important vector-borne diseases, is caused by a protozoan parasite of the genus
Leishmania, which is transmitted to human by the bite of sand flies. Leishmaniasis represents a complex disease with diverse clinical manifestations and poses a public health problem since it is a neglected tropical disease with current high worldwide incidence [
1,
2]. Globally, more than 12 million individuals are infected, with another 350 million at risk of infection, and nearly 2 million new cases are reported annually worldwide [
3]. The disease is prevalent in 16 developed and 72 developing countries; nevertheless 90 % of cases are reported in three regions: Sudan/Ethiopia/Kenya, India/Bangladesh/Nepal and Brazil with as many as 0.02 to 0.04 million deaths every year [
3,
4].
Leishmaniasis can be divided into three forms, varying in severity from self-healing cutaneous lesions, dermatological ulcers in cutaneous leishmaniasis (CL), destructive form of mucocutaneous leishmaniasis, to deadly form of visceral leishmaniasis (VL) [
5]. CL is characterized by ulcers on the skin that are often formed at the site of the insect vector bite. Those ulcers can undergo metastasis of the nasopharyngeal mucosa developing to tissue destruction, depending on the species of
Leishmania involved [
6].
Leishmania (Viannia) braziliensis and
Leishmania (V.) panamensis are responsible for cases of mucocutaneous leishmaniasis in the Americas, although
L. (V.) guyanensis and
L. (L.) amazonensis have been identified, especially, in immuno-compromised hosts [
6].
The first-line drugs for systemic treatment of leishmaniasis are parenterally administered antimonials such as the sodium stibogluconate (Pentostam®) and the N-methyl glucamine antimoniate (Glucantime®) [
7,
8] generally required for the treatment of CL in the New World due to the risk of mucosal involvement [
9]. This current chemotherapy presents several issues such as high cost, difficult administration and elevated toxicity, associated with serious side effects [
10], for instance musculoskeletal pain, gastrointestinal disturbances, mild to moderate headache, electrocardiographic QTs interval prolongation and mild to moderate increase of liver and pancreatic enzymes [
11]. Second-line drug Pentamidine and amphotericin B are not widely used due to their high toxicity and cost. Miltefosine, the first oral anti-leishmanial drug, is the treatment of choice for diffuse cutaneous leishmaniasis and New World cutaneous leishmaniasis caused by
Leishmania braziliensis but increasing resistance to this drug has been notified [
12].
All antileishmanial drugs except miltefosine have to be administrated parenterally. Most of these drugs are toxic, requires prolonged hospitalization and close monitoring, which makes the treatment costly and beyond the reach of most patients. Consequently, the development of alternative therapies is a priority for the treatment of leishmaniasis. As a strategy, the investigation of extracts and compounds, with biological activity, isolated from plants and used in traditional medicine is a promising in the research field for compounds with potential action for the prophylaxis and chemotherapy of CL [
13].
Essential oils (EOs) are complex mixtures of secondary metabolites isolated from plants. In these mixtures, there are 10–60 constituents at different concentrations, but usually only 2–3 major constituents determine the biological properties of the EO [
14]. Those compounds and their constituents present a broad pharmacological spectrum, and they are used as analgesics, sedatives, anti-inflammatory, and anti-spasmodic drugs, as well as antimicrobials, antiprotozoals and antihelmintics [
13,
15,
16]. It has been shown that several EOs or their constituents have inhibitory activity on protozoa, especially
Leishmania [
17‐
19]. For instance, Santos and colleagues demonstrated that copaiba oil from
Copaifera martii is a safer, shorter, less-expansive, and more easily administered antileishmanial drug [
18]. Therefore, the purpose of this present work was to analyze the effect of sixteen EOs biological potential on
L. amazonensis promastigotes forms and L6 cells and chemical constitution, by GC-MS, of those EOs that showed better leishmanicidal results.
Discussion
According to the classification of cytotoxicity and antileishmanial activity for extracts and fractions derived from plants and natural products defined by Study Program and Disease Control [
27], the evaluated EOs are classified as moderately toxic (100 < CC
50 ≤ 1000 μg/mL), except the EO from
S. guainensis, which was classified as toxic (10 < CC
50 ≤ 100 μg/mL). Regarding the antileishmanial activity only EOs from
S. guianensis,
C. dinisii,
M. chamomilla,
C. verbenaceae,
B. sarmientoi,
F. galbaniflua and
M. officinalis are considered moderately active (50 < IC
50 ≤ 150 μg/mL). The others are considered not active.
Considering the chemical composition of the EO from
M. chamomilla, (E)-β-farnesene and (E,E)-α-farnesene were found as the major compound group representing 73.07 % of the total composition. These results corroborate with those reported by Machado et al. [
28] that found farnesene derivatives as the most representative constituents (22 %) and their bioassays using EO from
Lantana camara revealed a significant leishmanicidal activity against
L. amazonensis (IC
50/72 h = 0.25 μg/mL), except for the cytotoxic activity, in which the authors obtained high values on Brine shrimp (CC
50 10 μg/mL). Subsequently, Gawde
et al. [
29] observed that the chemical composition of
M. chamomilla was similar to the one found in our study (β-farnesene, α-bisabolol oxide B, chamazulene) but no leishmanicidal activity on
L. donavani was observed.
Studies on the chemical composition and biological activity of
M. peruiferum EO are scarce. The literature reports (E) and (Z)-nerolidol, α-bisabolol and (E, E)-farnesol as its major components [
30] but those compounds were not identified in the present study. Santos
et al. [
18] reported high levels of α-copaene in EO from
Copaifera reticulata as well as for EO from
M. peruiferum. The last one showed growth inhibitory activity for
L. amazonensis with IC
50/72 h values of 5 μg/mL for promastigotes and low cytotoxicity on J774G8 macrophages.
Ghannadi and Amree [
31] have already described the EO composition obtained from the fresh oleogum resin and latex of Iranian
F. galbaniflua (synonym F. gummosa) and the main constituents of this monoterpene rich oil were β-pinene (58.8 %). Other studies also indicate β-pinene as the major compound from the fresh oleogum resin and latex of this same specie [
32,
33], which corroborates our results. The presence of methyl 8-(14)-pimaren-18-ate, a diterpene esters hydrocarbons, has been reported on rosin, a solid form of resin obtained from pines and some other plants; and also in the Cretaceous resins from India and Myanmar [
34,
35]. To our knowledge, there is no antileishmanial activity reports related to this EO to date.
Rodilla et al. [
36] determined the chemical composition of EO from
B. sarmientoi. In accordance with our work, they identified guaiol as its major component. Studies with EO from
Endlicheria bracteolata, which has 72.12 % of guaiol in its composition, showed IC
50 of 7.93 μg/mL for
L. amazonensis and presented a CC
50 of 15.14 μg/mL for J774.G8 macrophages [
37]. The antileishmanial activity may be attributed to the presence of a hydroxyl group of alcohol characteristics in the guaiol, especially in the exocyclic portion of the molecule [
36].
The presence of linalyl acetate and linalool as the major compounds in
S. sclarea EO (total of 88.88 %) corroborate to the results presented by Pitarokili et al. [
38] that evaluated the EO composition of
S. sclarea originated from two localities in Greece, and by Kuźma et al. [
39] that evaluated the EO composition from
S. sclarea plants generated
in vitro. On the other hand, antileishmanial activity of linalool-rich EO from leaves of
Croton cajucara against
L. amazonensis was previously evaluated by Rosa et al. [
40], they were able to demonstrate morphological changes in
L. amazonensis promastigotes when treated with 15 ng/mL of that EO. In this study the cell lysis was observed within 1 h, indicating that the antileishmanial activity observed is directly related to the presence of linalool, due to the existence of a hydroxyl group in the organic alcohol function.
As in our study, the presence of the isomers of citral, neral and geranial are constantly reported in the chemical composition of the EO from
M. officinalis [
41‐
43]. Regarding the antileishmanial activity, Mikus et al. [
44] reported an IC
50/72 h of 7 μg/mL for
L. major, a CC
50/72 h of 25.5 μg/mL in HL-60 cells and SI of 3.6, higher than those observed in our study. Another study has already showed that citral presents activity against
T. cruzi, possibly by inducing cell membrane lysis with leakage of cytoplasm [
45].
The EO from
C. dinisii and
S. guianensis showed weak inhibitory effect on the protozoan
T. cruzi with values of IC
50/24 h = 209.30 μg/mL and 282.93 mg/mL, respectively. These values are higher when compared to those obtained in the study for
L. amazonensis, 54.05 and 48.55 μg/mL, respectively [
25].
The mechanism of action by which EOs inhibits parasite growth is still not well known, but previous studies have suggested that structural and morphological changes are caused by drugs that inhibit ergosterol synthesis, or interact with the membrane ergosterol [
19,
46]. Other studies indicated that the activity of essential oils on parasites is mainly due to terpene composition. Terpenes are responsible for the hydrophobic characteristic of EOs, thus allowing their diffusion through the parasite cell membrane, affecting intracellular metabolic pathways and organelles [
47].