Investigation of the mutagenic and antimutagenic effects of Origanum compactum essential oil and some of its constituents

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Abstract

In the present study, the chemical composition of Origanum compactum essential oil was determined by gas chromatography and mass spectrometry, and its mutagenic and antimutagenic activities were investigated by the somatic mutation and recombination test (SMART) in Drosophila melanogaster. No significant increase in the number of somatic mutations was observed with the essential oil tested using both the standard (ST) and high bio-activation (HB) cross. In order to investigate the antimutagenic effect of the essential oil, we have tested the effect on the indirect-acting mutagen urethane (URE), as well as the direct-acting mutagen methyl methanesulfonate (MMS). O. compactum essential oil showed a strong inhibitory effect against URE-induced mutagenicity, especially with the HB cross. However, only a weak inhibitory effect on the mutagenicity induced by MMS was observed. These results suggest that the detected antimutagenicity could be mediated by an inhibitory effect on metabolic activation. The essential oil was fractionated to identify the components responsible of the suppressing effect detected. Seven fractions were obtained: two of them showed the most potent inhibitory effect against URE-induced mutagenicity and were further fractionated. The sub-fractions obtained from the second chromatographic fractionation were tested for their antimutagenic activity, together with carvacrol and thymol. The highest antimutagenic effect obtained with the sub-fractions was similar to the effect of the crude essential oil, as well as to the effect of carvacrol alone. These results suggest the absence of a synergic antimutagenic effect between the components of O. compactum essential oil and indicate that carvacrol was the most active oil component.

Introduction

Recently, cancer chemoprevention has become a promising alternative to control cancer, now that a large number of modulating factors including antimutagens and anticarcinogens have been widely studied to assess their ability to suppress or prevent the process of carcinogenesis. It has been accepted that plants and their products represent one of the main sources for compounds with chemopreventive potential and, indeed, several secondary plant metabolites have demonstrated chemopreventive activity [1].

Essential oils are isolated from many plants by mechanical pressing or hydro- and steam-distillation. They are complex mixtures of odorous and volatile compounds from secondary plant metabolism: monoterpenes, sesquiterpenes, alcohols, ethers, aldehydes, esters and ketones are the main constituents of these mixtures. The essential oils are often responsible for a plant's distinctive scent or taste and are widely used as flavour enhancers in many food products and as odorants in fragrances [2]. Moreover, the essential oils and their ingredients are lipophilic compounds that are able to cross cell membranes and therefore absorbed through the skin [3], which is the reason why they have a long history of use for many medical applications [4]. Nowadays, the essential oils have been shown to possess a range of biological activities including antibacterial, antioxidant, antifungal and insecticidal activity [5], [6], [7]. However, just a few of the essential oils have been studied for their antimutagenic activity [8].

All over the world, the Origanum genus was the most commonly found among oregano species. Forty-nine taxa divided into 10 sections belong to this genus, most of them having a very local distribution around the Mediterranean [9]. Origanum compactum with very compact pinkish flowers is the endemic oregano in Morocco. In general, the oregano spices are widely used to enhance the flavour of many different foods, but O. compactum for its part is used as herbal remedy in folk medicine, especially as infusion for many purposes [10]. No published data are available on the mutagenicity and antimutagenicity of O. compactum oil and only a few studies with some of its components have been performed. Carvacrol was suggested to be antimutagenic against 4-nitro-o-phenylenediamine and 2-aminofluorene using the Ames test [11]. Carvacrol is also able to prevent mutagenicity of some known mutagens in human lymphocytes by inhibiting chromatid exchange [12]. The thymol together with the carvacrol γ-terpinene were reported for their protective effect against 2-amino-3-methylimidazo[4,5-f]-quinoline and mitomycin C using the comet assay [13].

In this study, we first evaluated the mutagenic activity of O. compactum oil and then investigated the antimutagenic potential of the crude essential oil against a promutagen and a direct alkylating agent using the somatic mutation and recombination test in Drosophila melanogaster. To identify the components responsible of the detected antimutagenic activity, the Origanum essential oil was separated into a series of fractions and each fraction was evaluate for its antimutagenic activity. A second fractionation was performed for fractions that had demonstrated the highest level of antimutagenic activity. Determination of the chemical composition of the essential oil as well as the series of the fractions obtained was elaborated by gas chromatography and gas chromatography–mass spectrometry. Finally, we evaluated the antimutagenic activity of the major constituents of the most potent fraction, mainly carvacrol and thymol, to compare the antimutagenic activity of the crude Origanum essential oil to that of its constituents.

Section snippets

Compounds and plant material

The Origanum was collected from the north of Morocco, only flowering tops of the plant were used. Oil of O. compactum was produced by PRANAROM International Company, Belgium. Urethane (URE) [CAS no. 51-79-6] and methyl methanesulfonate (MMS) [66-27-3] as well as carvacrol [499-75-2], thymol [89-83-8], pentane, HPLC grade [109-66-0] and diethyl ether [60-29-7] were purchased from Sigma Aldrich. Acetone [67-64-1] was from Riedel-deHaën.

Extraction of the essential oil and identification of its constituents

The essential oil was extracted by hydro-distillation. This

Results

The different constituents of the O. compactum essential oil were identified and quantified by GC and GC–MS. The chemical composition of the essential oil is presented in Table 1. The essential oil was particularly rich in carvacrol 22%, γ-terpinene 22%, thymol 19% and p-cymene 13%.

The Origanum essential oil was first analyzed for the evaluation of its genotoxic/mutagenic activity. The numbers of small spots, large spots and twin spots together with the total number of spots are given in Table 2

Discussion

The somatic mutation and recombination test in D. melanogaster was performed to study the mutagenicity and antimutagenicity of O. compactum essential oil, using the ST and the HB cross. The larvae from the HB cross are characterized by a high level of cytochrome P450 [17]. P450 plays the main role in the bio-activation of numerous classes of compounds, including drugs, carcinogens and pesticides, as well as natural products and plant constituents [18], [19], [20].

Data from the wing SMART assay

Acknowledgements

This work was supported by PRANAROM International Company and the Ministry of Wallone region, Belgium (Direction générale des technologies, de la recherche et de l’énergie, Promotion de l’innovation technique, Jambe-Namur). Acknowledgements to the Manuscript Review Committee at Biomatecus and Dr B. Edderkaoui for revision of manuscript and language.

References (32)

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