Comparative Study of Three Achillea Essential Oils from Eastern Part of Turkey and their Biological Activities

Essential oils obtained by hydrodistillation were analyzed both by gas chromatography (GC) and gas chromatography-mass spectrometry (GC-MS). The main constituents found in Achillea oil were as follows: A. filipendulina Lam.: 43.8% santolina alcohol, 14.5% 1,8-cineole and 12.5% cis-chrysanthenyl acetate; A. magnifica Hiemerl ex Hub.-Mor.: 27.5% linalool, 5.8% spathulenol, 5.5% terpinen-4-ol, 4.7% α-terpineol and 4.7% β-eudesmol; A. tenuifolia Lam.: 12.4% artemisia ketone, 9.9% p-cymene, 7.1% camphor, 5.9% terpinen-4-ol, 4.7% caryophyllene oxide and 4.5% α-pinene. Furthermore, the Achillea essential oils were evaluated for antimalarial and antimicrobial activities. A. magnifica and A. filipendulina oils showed strong antimalarial activity against both chloroquine sensitive D6 (IC50= 1.2 and 0.68 g/mL) and chloroquine resistant W2 (IC50= 1.1 and 0.9 g/mL) strains of Plasmodium falciparum without any cytotoxicity to mammalian cells up to IC50=47.6 g/mL against Vero cells. whereas A. tenuifolia oil showed no antimalarial activity up to a concentration of 20 mg/mL. All three Achillea oils showed no antibacterial activity against human pathogenic bacteria up to a concentration of 200 g/mL. A. tenuifolia and A. magnifica oils demonstrated mild antifungal activity against Cryptococcus neoformans (IC50= 45, 20 and 15 g/mL, respectively).

magnifica Hiemerl ex Hub.-Mor. from eastern part of Turkey. Out of the three species, A. magnifica, an endemic species, is recognized by local names as "karcivanı" and its dried flowering parts are mixed with honey and consumed for stomach disorders [4].
The plant material was identified by Prof. Dr. Z. Aytaç. Voucher specimens were deposited at the Herbarium of Gazi University, Faculty of Science and Letters. Detailed information on the plant materials used are given in Table 1.

Previous Studies
Essential oils of Achillea species have been the subject of several investigations. A. millefolium L. has been the most widely studied species because of its economic value and therapeutic properties. Previous phytochemical investigations of Achillea species have revealed the presence of sesquiterpene lactones, proazulenes, sesquiterpenoids, flavonoids, triterpenes, coumarins, peroxides, phenolic and polyacetylene compounds [5].
Although the essential oil composition of A. filipendulina, A. tenuifolia and A. magnifica have been recently reported in the literature and the antibacterial activity of A. filipendulina has been studied [5][6][7][8][9][10], the antimalarial activity of A. tenuifolia and antimicrobial activity of A. tenuifolia and A. magnifica were investigated for the first time.

Present Studies
Isolation of the Essential Oil: Essential oils were hydrodistilled from dried aerial parts for 3 h using a Clevenger apparatus. The yields were calculated on a dry weight basis as given in Table 1.

GC-MS and GC-FID Analysis:
The GC-MS and GC-FID analysis were carried out with an Agilent 5975 GC-MSD and Agilent 6890N GC systems, respectively. Analysis conditions and identification of the oil components are similar to our earlier studies [1].
Biological Activity: The in vitro antimalarial [11] and antimicrobial [12] activity was performed as previously described. Antimalarial standards chloroquine (Aldrich-Sigma, ST, Louis, MO) and artemisinin (Aldrich-Sigma, ST, Louis, MO) and antimicrobial standards ciprofloxacin (ICN Biomedicals, Ohio) for bacteria and amphotericin B (ICN Biomedicals, Ohio) for fungi were purchased from commercial sources. Air-dried aerial parts of three Achillea species, A. filipendulina Lam., A. magnifica Hiemerl ex Hub.-Mor., and A. tenuifolia Lam., were analyzed by GC and GC-MS and the individual identified components with their relative percentages are given in Table 2.
In the oil of the A. filipendulina, 53 components were characterized representing 93% of the total oil. This oil was characterized by a relatively high content of santolina alcohol (43.8%). 1,8-Cineole (14.5%) and cis-chrysanthenyl acetate (12.5%) were found as other main constituents.
In the current study, these three Achillea essential oils were evaluated for their antimalarial and antimicrobial activities. A. magnifica and A. filipendulina oils showed strong antimalarial activity against both chloroquine-sensitive D6 (IC50= 1.2 and 0.68 g/mL) and chloroquine-resistant W2 (IC50= 1.1 and 0.9 g/mL) strains of Plasmodium falciparum when the comported to positive standards chloroquine (IC50= 0.018 g/mL for D6 and IC50= 0.16 g/mL for W2 clones) and artemisinin (IC50= 0.0037 g/mL for D6 and IC50= 0.0035 g/mL for W2 clones) without exhibiting any cytotoxicity at IC50 values of 47.6, 15.867 and 5.288 g/mL against Vero cells. however, A. tenuifolia did not show antimalarial activity Achillea oils showed no antibacterial activity against human pathogenic bacteria up to a concentration of 200 g/mL. A. tenuifolia and A. magnifica oils demonstrated mild antifungal activity against Cryptococcus neoformans (IC50= 20 and 15 g/mL, respectively) and A. tenuifolia oil demonstrated weak antimycobacterial activity against Mycobacterium intracellulare with an IC50 value of 200 µg/mL. Bioassay-guided investigations are warranted out to identify active antimalarial compounds from A. magnifica and A. filipendulina essential oils.
To the best of our knowledge, this is the first report of chemical composition these three Achillea species from eastern (Malatya and Ağrı) and south-eastern (Şırnak) region of Turkey and their biological activities of A. magnifica, A. tenuifolia and A. filipendulina essential oils were evaluated.