Antinociceptive effect of Aristolochia trilobata stem essential oil and 6-methyl-5-hepten-2yl acetate, its main compound, in rodents

1 Introduction

Pain is defined by the International Association for the Study of Pain as “an unpleasant sensory and emotional experience associated with actual or potential tissue damage” [1]. It has the function of protecting the organism from noxious stimuli [2]. However, sometimes its etiology is heterogeneous and the underlying pathophysiology mechanisms are complex, making treatment difficult and interfering with the quality of life for people with these pain syndromes [3]. Besides, the wide range of interindividual variability in the perception of pain makes the management of pain a major challenge for medicine [4].

Natural products emerge as interesting therapeutic resources for the development of new drugs for the management of certain painful states [5]. The search for new chemical entities as therapeutic alternatives for pain or inflammatory conditions has constantly progressed [6], [7] and presents the medicinal plants as the major source [8], [9], [10], [11].

Aristolochia trilobata L. (Aristolochiaceae), popularly known as “mil-homens”, is found from Central America down to South America, having various applications in folk medicine [12]. A. trilobata is used by humans to treat stomach aches, colic, poisoning, diabetes and also for skin affections [13], [14]. There are few pharmacological data on this plant species. Previous studies have shown that a chloroform extract of A. trilobata leaves had antiphlogistic potency close to that of the non-steroidal anti-inflammatory drug indomethacin [14].

Considering the antinociceptive potential of essential oils (EOs) and their constituents and the absence of publications demonstrating the antinociceptive effect of A. trilobata, this study evaluates the antinociceptive property of stem EO of A. trilobata and its major compound, 6-methyl-5-hepten-2yl acetate (sulcatyl acetate, SA), in animal models of nociception.

2 Materials and methods

3 Results and discussion

In this study, we evaluated the antinociceptive activity of the EO obtained from the stem of A. trilobata (EO) and 6-methyl-5-hepten-2yl acetate (SA), its main compound, both in male mice.

According to Santos et al. [15], our sample of the EO of A. trilobata presented an average yield of 0.22±0.05% (v/w), and is composed mainly of 6-methyl-5-hepten-2-yl acetate (SA) (21.49±0.43%), germacrene D (15.07±0.23%), bicyclogermacrene (8.84±0.45%), linalool (6.85±0.42%), (E)-caryophyllene (5.58±0.12%), (E)-β-ocimene (5.56±0.067%) and p-cymene (4.68±0.10%).

In the pretreatment with EO or SA, all doses significantly (p<0.001) reduced the number of writhing movements (nociceptive behavior) induced by acetic acid solution (Figure 1). As expected, MOR (5 mg/kg, i.p.), an opioid analgesic drug, produces an inhibition of nociceptive behavior (p<0.001). In this model, pain is elicited by the injection of an irritant such as acetic acid into the peritoneal cavity, which produces episodes of characteristic stretching (writhing) movements, and inhibition of the number of episodes by analgesics is easily quantifiable. Despite being a non-specific test, acetic acid-induced writhings are a highly sensitive and useful test for analgesic drug development [17].

Figure 1:

Effects of EO or SA on the acetic acid-induced writhing test in mice. Vehicle (control), EO (25, 50 or 100 mg/kg), SA (25 or 50 mg/kg) or MOR (5 mg/kg) were administered i.p. 1 h before acetic acid injection. Each column represents mean±SEM (n=8, per group). ^***p<0.001 versus control (ANOVA followed by Dunnett’s test).

Posteriorly, the antinociceptive effects of EO and SA were evaluated in a model of persistent nociception, the formalin test, which has two distinctive phases that can possibly indicate different types of pain [18]. The early and late phases of the formalin test have obvious differential properties, and therefore this test is useful not only for assessing the analgesic substances but also for an initial investigation the mechanism of analgesia [20]. The early phase, named non-inflammatory pain, is a result of direct stimulation of nociceptors and reflects centrally mediated pain; the late phase, named inflammatory pain, is caused by local inflammation with a release of inflammatory and hyperalgesic mediators [18]. In the present study, we found that EO and SA produced antinociceptive activity both in the early (p<0.05) and late (p<0.05–0.001) phases of the formalin test (Figure 2). However, EO was effective in the early phase only in higher dose (p<0.05).

Figure 2:

Effects of EO or SA on the formalin-induced nociception in mice. Vehicle (control), EO (25, 50 and 100 mg/kg), SA (25 or 50 mg/kg) or aspirin (200 mg/kg) were administered i.p. 1 h before formalin injection. (A) The first phase and (B) the second phase of formalin-induced nociception in mice. Each column represents mean±SEM (n=8, per group). ^*p<0.05, ^**p<0.01 or ^***p<0.001 versus control (ANOVA followed by Dunnett’s test).

Several studies have shown the antinociceptive and anti-inflammatory effects of terpenes or EOs, as reviewed by Almeida et al. [21], de Cássia da Silveira e Sá et al. [22], De Sousa [23] and Guimarães et al. [6], corroborating with our findings. Studies performed with linalool and p-cymene, major compounds found in EO of A. trilobata, demonstrated the antinociceptive activity of these monoterpenes in different experimental models of nociception [3], [11], [24], [25], [26]. Furthermore, the major compound, which has its pharmacological activity described for the first time in this study, can act synergistically with these compounds, thus contributing to the pharmacologic response observed. This therapeutic property has gained prominence among research groups of academic community and industry, which increased interest in developing new options for pain management. Furthermore, the high market acceptance of drugs containing terpenes and EOs for pain control, such as Acheflan^® and Salompas^®, motivates further research in this area and has led to an increase in patent filings in several countries [7].

Previous studies suggested that the CNS depression and the non-specific muscle relaxation effect can reduce the response of motor coordination and might invalidate the behavior test results [19]. We observed that EO- or SA-treated mice did not show any significant motor performance alterations with all doses tested (data not shown). Thus, we refute the hypothesis of a possible relaxing or motor deficit effects of EO and SA at the therapeutic doses.

Together, our results suggest that EO and SA, its main compound, modulate neurogenic and inflammatory pain in the tests used (Figure 3). Thus, it was observed that the SA is responsible, at least in part, for the analgesic profile of this EO. The antinociceptive actions demonstrated in A. trilobata in this study support the ethnomedicinal use of this plant. More studies are needed to determine the ED50 and the dose conversion ratio of compounds with standard drugs.

Figure 3:

Summary of the data obtained.