Assessment of Anticancer Properties of Essential Oils from sand dunes of Peniche (Portugal)
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1
ESTM & GIRM, Polytechnic Institute of Leiria, Portugal
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2
University of Hull, School of Biological, Biomedical and Environmental Sciences, United Kingdom
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3
Faculty of Pharmacy/CEF and CNC, University of Coimbra, Portugal
1. Introduction
An essential oil (EO), also defined as essence, volatile oil, etheric oil or aetheroleum, is a complex mixture of volatile constituents biosynthesised by aromatic plants. These compounds are mainly monoterpenes, sesquiterpenes and phenylpropanoids.
EOs can be synthesized by all plant organs (root, stem, twigs, leaves, flowers, buds, fruits and seeds) and stored in structures classified into 6 groups: (1) secretory cells; (2) osmophores; (3) secretory cavities; (4) secretory ducts; (5) glandular trichomes; (6) epidermal cells.
Essential oils are characteristic as being non-polar or weakly polar liquids, soluble in alcohol and in dimethyl sulfoxide (DMSO), and less dense than water.
They may have many biological activities, including anticancer, anti-inflammatory, antimicrobial, antiviral, antioxidant and/or insect repellent.
The aim of this study was to determine the efficacy of the essential oils from six species of Peniche (Portugal) sand dunes as potential cytotoxic/cytostatic therapeutic agents in breast and colorectal cancer cell line models.
2. Materials and methods
2.1 Plant material
Aerial parts of Artemisia campestris, Crithmum maritimum, Eryngium maritimum, Juniperus turbinata, Otanthus maritimus and Seseli tortuosum were collected during flowering stage in July 2013, in the sand dunes of Consolação beach, Peniche (Portugal). Voucher specimens were deposited in the Herbarium of Medicinal Plants, Faculty of Pharmacy of the University of Coimbra.
2.2 Essential oils isolation
Essential oils were isolated by hydrodistillation for 3h using a Clevenger-type apparatus, according to the procedure described in the European Pharmacopoeia (Council of Europe, 1997).
2.3 Chemical characterization of essential oils
The essential oils were chemically characterized by gas chromatography (GC) and gas chromatography–mass spectrometry (GC–MS).
2.4 Cell Culture
Human cancer cell lines RKO (colorectal) and MCF7 (breast) were maintained in medium (Dulbecco’s Modified Eagle’s), supplemented with 10% fetal bovine serum (FBS) and 1% sodium pyruvate. Cells were grown at 37˚C, under a 5% CO2 atmosphere and at 90% humidity. The subculture was made three times a week. Cells regularly tested negative for Mycoplasma infection.
2.5 In vitro inhibition of RKO and MCF7 cells growth
RKO and MCF7 cell lines were seed into 96 well microtiter plates to determine the activity of every essential oil, at a density of 1x104 and 5x103 cells per well, respectively.
The serial doubling dilution of the EO was prepared in DMSO. For stock solution were used 25µL EO and 75µL of DMSO (1:4). The final concentration used was 1 µL/mL and the dilution was 1:10.
2.6 MTS Assay for cellular viability
MTS (3-(4, 5-dimethyl-2-thiazolyl)-2, 5-dyphenyl-2H-tetrazolium bromide) colorimetric assay measures cell respiration and the amount of formazan produced is proportional to the number of living cells present in culture. It is a sensitive, quantitative and reliable colorimetric assay. This method was employed as test for quantification of cell proliferation after 72 hours incubation at 37˚C and 5% CO2. For this purpose, the medium of each well was removed and 100µL of fresh medium was added in addition to 20µL of MTS solution. After 4h incubation, the absorbance was determined using a microplate reader (Bioteck ELx800) at a test wavelength of 490 nm.
3. Results
To determine the antitumor activity of EOs against RKO and MCF7 cells, MTS assay was carried out, and cell viability was calculated comparing to the vehicle (DMSO), corresponding to 100% proliferation (fig. 1-6).
For all the EOs tested, antitumor activity was shown, as indicated by a decrease in cell viability – except for J. turbinata and C. maritimum which had the least effect on cell viability for MCF7 and RKO cell lines, respectively (table 1).
4. Discussion and Conclusion
In this study, different EOs from sand dune plants have been investigated for their antitumor activity using colorectal and breast cancer cell lines.
All essential oils tested showed antitumor activity as measured by their effect on cell viability. The most noticeable decrease in viability was shown by S. tortuosum and E. maritimum EOs, as indicated by IC50 average values of 3.29E-03 and 8.59E-04 (for RKO) and 1.85E-03 and 2.72E-03 (for MCF7), respectively.
Some of the tested EOs, such as J. turbinata for MCF7 cell line and C. maritimum for RKO cell line, showed lower activity (IC50 average value of 3.85E+1 for MCF7 cells and 8.76E00 for RKO cells, respectively).
There are several studies on the antioxidant activity of plant extracts. For example, Aniya and collaborators (2000) investigated the antioxidant and hepatoprotective action in water extracts of A. campestris (from Okinawa Islands), and observed that these extracts provide protection against liver toxicity. The literature available for potential antitumor activity of EOs is more limited, however, there are some studies, about citrus fruits, cherry, mint and herbs that indicate potential of antitumor activity against different types of human cancers, including breast cancer, liver cancer and melanoma. In these studies, this effect is associated with the specific chemical compounds of the EOs, including monoterpenes (Crowell, 1999).
It would be interesting to study the role of individual chemical components from the EOs in any potential effects, namely anticancer activity, in their action. For this, it is necessary to analyse in detail the chemical composition of each EO. These would then be evaluated independently and compared to the complex EO mix.
Finally, the data here presented indicates that these EOs have an anti-proliferative effect on the cell lines used. Further work needs to be undertaken to investigate the effect of the EOs in known signalling pathways that regulate cell proliferation and viability, such as apoptosis, DNA damage-induced cell cycle arrest or by decreased proliferation signalling.
References
Aniya, Y., Shimabukuro, M., Shimoji, M., Kohatsu, M., Gyamfi, M.A., Miyagi, C., Kunii, D., Takayama, F., Egashira, 2000. Antioxidant and hepatoprotective actions of the medicinal herb Artemisia campestris from the Okinawa Islands. Biol. Pharm. Bull. 23 (3), 309–312.
Council of Europe. (1997). European Pharmacopoeia. (3rd ed). Strasbourg: Council of Europe.
Crowell, P.L., 1999. Prevention and therapy of cancer by dietary monoterpenes. J. Nutr. 129 (3), 775–778.
Keywords:
Sand dune plants,
Peniche,
essential oils,
Anticancer properties,
Artemisia campestris,
Crithmum maritimum,
Eryngium maritimum,
Juniperus turbinata,
Otanthus maritimus,
Seseli tortuosum
Conference:
IMMR | International Meeting on Marine Research 2014, Peniche, Portugal, 10 Jul - 11 Jul, 2014.
Presentation Type:
Poster Presentation
Topic:
BLUE BIOTECH
Citation:
Poças
J,
Lemos
MF,
Cabral
C,
Salgueiro
L and
Pires
IM
(2014). Assessment of Anticancer Properties of Essential Oils from sand dunes of Peniche (Portugal).
Front. Mar. Sci.
Conference Abstract:
IMMR | International Meeting on Marine Research 2014.
doi: 10.3389/conf.fmars.2014.02.00011
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Received:
11 May 2014;
Published Online:
18 Jul 2014.
*
Correspondence:
Miss. Juliana Poças, ESTM & GIRM, Polytechnic Institute of Leiria, Peniche, Portugal, julianasacop13@gmail.com
Prof. Isabel M Pires, University of Hull, School of Biological, Biomedical and Environmental Sciences, Hull, United Kingdom, i.pires@hull.ac.uk