Composition of the essential oil of Stachys sericantha, S. gaziantepensis, and S. mardinensis (Lamiaceae) from Turkey

ABSTRACT

The genus Stachys L. one of the largest genera of Lamiaceae and Stachys species are used by many people for the therapeutic value of their essential oils. In this study, water-distilled essential oil of three species of Stachys was analysed. The analysis was performed by using gas chromatography-flame ionisation detector (GC-FID) and gas chromatography-mass spectrometry (GC-MS) systems simultaneously. The major components found were hexadecanoic acid (23.7%), dodecanoic acid (11.3%), and caryophyllene oxide (10.7%) for S. sericantha P.H. Davis (endemic); α-pinene (53%), β-pinene (8.2%) for S. gaziantepensis M. Dinc and S. Doğu (endemic); Menthyl acetate (15.3%), isomenthone (15.0%), pulegone (10%), spathulenol (7.0%), and caryophyllene oxide (6.7%) for S. mardinensis (Post) R. Mill.

KEYWORDS:

• Essential oil
• GC-FID
• GC-MS
• Stachys sericantha
• Stachys gaziantepensis
• Stachys mardinensis

Introduction

The genus Stachys L. one of the largest genera of the subfamily Lamioideae (Lamiaceae) includes about 300 species of annuals and perennials occurring in all parts of the world, except for Australia and New Zealand.^[1] The Stachys genus is represented by 87 species (112 taxa) belonging to 15 sections and 2 subgenera in Turkey; 52 species are endemic to Turkey and endemism ratio is 46%.^[2–5] Most of the endemic taxa are east Mediterranean elements. The species name originates from the Greek and means “an ear of grain” referring to the inflorescence spike found in many members. They are consumed as herbal remedies in alternative medicine and wild tea (mountain tea) in Mediterranean regions. Many Stachys species are used in decoctions or infusions for the treatment of skin, stomach, ulcer, asthma, rheumatic disorders, and vaginal tumours. Some members of genus have been reported to be used as anti-inflammatory and antibacterial agents. Moreover, their antianxiety, antioxidant, and antinephritic properties have also been reported.^[6] According to ethnobotanical studies, Stachys species are generally consumed as herbal tea and a rich carbohydrate source in Europe and East Asia. A lot of Stachys species in Turkey are widely used as herbal teas due to their antibacterial properties. Essential oil composition of the species is one of the main reasons of their consumption as tea in Anatolian ethnobotany^[7,8]; however, the species also consist of glycosides, saponins, polyphenols, tannins, phenolic acids, flavonoids, and diterpenoids together with essential oils, mono and sesquiterpenoids. Therefore, the synergistic effects of the component chemicals could be the main reason of consumption of their flowers and aerial parts as tea in Anatolian culture.^[6]

The essential oil compositions of the Stachys genus have been well documented in the literature and they mainly consist of sesquiterpenes and oxygenated sesquiterpenes. The main components of the essential oil of the species were observed to be germacrene D, caryophyllene, cadinene, spathulenol, and caryophyllene. Moreover, the monoterpenes such as α-pinene, β-pinene, phellandrene, and carvacrol were also extracted from Stachys species.^[6,9]

S. sericantha P.H. Davis is a member of section Eriostomum, endemic and Mediterranean element. The plant is perennial 35–70 cm long and corolla purple-pink coloured, 13–16 mm. It grows on stone slopes, in between 20 and 1300 m altitude in Antalya province and the flowering time is from June to July.^[2] The plant is locally known as “dikenli cay” in the regions where it grows.^[10] In Turkey, S. sericantha is used in the form of herb, infusion, and decoction as a remedy for the treatment of cold, stomach ailments, fever, and cough.^[6] S. gaziantepensis M. Dinc and S. Dogu is an endemic perennial plant, included in section Infrarosularis. The plant is 10–30 cm long and corolla yellow coloured, 10–12 mm. It grows on marble rocks between 1060 and 1300 m altitude in Gaziantep province. The flowering time is between second half of May and first half of June. S. gaziantepensis is also used as herbal tea for the treatment of cold in the regions where it grows.^[11] S. mardinensis (Post) R. Mill is a member of section Fragilicaulis and Iranian-Turan element. The plant is perennial 10–60 cm long and corolla yellowish, 12–20 mm. It grows on sloping limestone rocks and cliff crevices, between 450 and 1200 m altitude in Mardin and Malatya provinces. The flowering time is from May to July.^[2] The plant is locally known as “Kaya Pungu” or “Punge Tehta” and is used for headaches in the regions where it grows.^[10,12] In the present paper, the chemical composition of the essential oils obtained from the aerial parts of endemic S. sericantha, endemic S. gaziantepensis, and S. mardinensis from Turkey has been reported. Most probably, this could be the first report on the essential oil chemical compositions of S. gaziantepensis and S. mardinensis.

Materials and methods

Plant material

Plant materials were collected during the flowering period from Antalya (S. sericantha), Gaziantep (S. gaziantepensis), and Mardin (S. mardinensis) provinces of Turkey (Table 1). The voucher specimens are kept at the Herbarium of the Department of Biology, Necmettin Erbakan University, Konya, Turkey (NEÜ Herb.).

Table 1. Stachys spp. used in this study.

Stachys spp.	Plant part	Collection site	Collection date	Yield (%)	Herb. No
* S. sericantha	Aerial part	Antalya: Konyaaltı	2010	tr	3338
*S. gaziantepensis	Aerial part	Gaziantep: Şehitkamil	2012	0.11	3467
S. mardinensis	Aerial part	Mardin: Samandağ	2010	0.02	2472

*Endemic.

Isolation of essential oil

The essential oil from air-dried plant materials was isolated by hydrodistillation for 3 h, using a Clevenger-type apparatus to produce a small amount of essential oil of S. sericantha which was trapped in n-hexane. The yields of oils were 0.11% for S. gaziantepensis and 0.02% for S. mardinensis (Table 1). The obtained oils were dried over anhydrous sodium sulphate and stored at +4°C in the dark until analysed and tested.

Gas chromatography-mass spectrometry analysis

The gas chromatography-mass spectrometry (GC-MS) analysis was carried out with an Agilent 5975 GC-MSD system. Innowax FSC column (60 m × 0.25 mm, 0.25 μm film thickness) was used with helium as carrier gas (0.8 ml/min). GC oven temperature was kept at 60°C for 10 min and programmed to 220°C at a rate of 4°C/min, and kept constant at 220°C for 10 min and then programmed to 240°C at a rate of 1°C/min. Split ratio was adjusted at 40:1. The injector temperature was set at 250°C. Mass spectra were recorded at 70 eV. Mass range was from m/z 35 to 450.

GC analysis

The GC analysis was carried out using an Agilent 6890N GC system. Flame Ionisation Detector (FID) detector temperature was 300°C. To obtain the same elution order with GC-MS, simultaneous auto-injection was done on a duplicate of the same column applying the same operational conditions. Relative percentage amounts of the separated compounds were calculated from FID chromatograms. The analysis results are given in Table 2. The essential oil components were identified by comparison of their relative retention times with those of authentic samples or by comparison of their relative retention index (RRI) to series of n-alkanes. Computer matching against commercial (Wiley GC/MS Library, MassFinder 3 Library)^[13,14] and in-house “Baser Library of Essential Oil Constituents” built up by genuine compounds and components of known oils, as well as MS literature data,^[15,16] was used for the identification.

Results and discussion

As in our samples, the yield of essential oil of Stachys species is lower than other Lamiaceae family members (e.g. Origanum, Satureja, Sideritis, Salvia, Thymus). However, the consumption pattern of the species is reported to be quite similar to the other members of the family. Most of the species of Stachys have a pleasant smell. They are consumed as herbal tea in major parts of the world and Turkey, and due to their moderate antibacterial, antifungal, anti-inflammatory, and antioxidant capacity, they are widely used as herbal remedy in alternative medicine.^[6]

In this study, the water-distilled essential oils from aerial parts of the three Stachys species were characterized by gas chromatography-flame ionisation detector (GC-FID) and GC-MS. The compounds identified from the essential oils along with their relative percentages are listed in Table 2. According to our findings, the essential oil of S. gaziantepensis of which 89.8% of the composition was determined, was characterized by its high proportion of monoterpene hydrocarbons (71.1%). Of the 18 components detected, the most abundant compounds were determined to be α-pinene (53%) followed by β-pinene (8.2%). In S. mardinensis oil, 16 components were identified representing 82.8% of the total oil. The oil consisted mainly of oxygenated monoterpenes (62.0%) and oxygenated sesquiterpenes (15.7%). Menthyl acetate (15.3%), isomenthone (15.0%), pulegone (10%), menthol (8.4%), spathulenol (7.0%), and caryophyllene oxide (6.7%) were the major components. In S. sericantha oil, 59 components were accounted for 82.5% of the total composition of the oil. The oil consisted mainly of Fatty acid + esters (41.0%) and oxygenated sesquiterpenes (22.8%). Hexadecanoic acid (23.7%), dodecanoic acid (11.3%) and caryophyllene oxide (10.7%) were found as major components.

Table 2. The composition of the essential oil of S. sericantha, S. gaziantepensis, and S. mardinensis.

RRI	Compound	Ss %	Sg %	Sm %
1032	α-Pinene	0.3	53.0	–
1035	α-Thujene	–	0.9	–
1118	β-Pinene	0.9	8.2	–
1132	Sabinene	–	1.0	–
1174	Myrcene	–	3.7	–
1203	Limonene	–	3.0	–
1255	γ-Terpinene	–	0.6	–
1280	p-Cymene	–	0.7	–
1475	Menthone	–	–	1.5
1497	α-Copaene	0.1	–	–
1503	Isomenthone	–	–	15.0
1528	α-Bourbonene	tr	–	–
1535	β-Bourbonene	0.6	–	–
1553	Linalool	0.2	–	–
1574	Menthyl acetate	–	–	15.3
1591	Bornyl acetate	0.1	–	–
1594	Isomethylacetate	–	–	4.3
1600	β-Elemene	0.6	–	–
1610	Calarene (=β-gurjunene)	0.1	–	–
1612	β-Caryophyllene	4.2	–	–
1632	Neoisomenthol	–	–	2.2
1638	Menthol	–	–	8.4
1648	Myrtenal	0.1	–	–
1662	Pulegone	–	–	10.0
1664	Nonanol	0.1	–	–
1687	α-Humulene	0.9	–	–
1695	(E)-β-Farnesene	–	1.0	–
1704	γ-Muurolene	0.6	–	–
1706	α-Terpineol	0.7	–	–
1726	Germacrene D	0.6	3.5	–
1733	cis-Piperitone oxide	–	–	2.9
1740	Valencene	0.1	–	–
1740	α-Muurolene	0.3	–	–
1748	Piperitone	–	–	1.6
1755	Bicyclogermacrene	–	1.8	2.7
1765	(E)-2-Undecanal	0.2	–	–
1766	Decanol	tr	–	–
1773	δ-Cadinene	–	1.0	–
1784	(E)-α-Bisabolene	0.2	–	–
1785	7-epi-α-Selinene	–	tr	–
1804	Myrtenol	0.1	–	–
1838	(E)-β-Damascenone	0.2	–	–
1868	(E)-Geranyl acetone	0.3	–	–
1900	epi-Cubebol	0.1	–	–
1941	α-Calacorene	0.2	–	–
1949	Piperitenone	–	–	0.8
1958	(E)-β-Ionone	0.1	–	–
1973	Dodecanol	0.1	–	–
1984	γ-Calacorene	0.2	–	–
1992	2-Phenylethyl isovalerate	1.0	–	–
2008	Caryophyllene oxide	10.7	–	6.7
2037	Salvial-4(14)-en-1-one	0.4	–	–
2056	Ethyl tetradecanoate	0.1	–	–
2071	Humulene epoxide-II	1.4	–	–
2081	Humulene epoxide-III	0.2	–	–
2130	Salviadienol	0.7	–	–
2131	Hexahydrofarnesyl acetone	1.6	–	0.4
2144	Spathulenol	–	1.9	7.0
2187	T-Cadinol	–	–	2.0
2209	T-Muurolol	0.2	–	–
2210	Copaborneol	–	2.0	–
2239	Carvacrol	–	4.4	–
2255	α-Cadinol	–	1.8	–
2262	Ethyl hexadecanate	0.3	–	–
2278	Torilenol	1.2	–	–
2298	Decanoic acid	0.6		–
2312	9-Geranyl-p-cymene	–	–	2.0
2324	Caryophylla-2(12),6(13)-dien-5α-ol (=Caryophylladienol II)	1.2	–	–
2369	Eudesma-4(15),7-dien-4β-ol	1.0	–	–
2384	Farnesyl acetone	0.5	–	–
2389	Caryophylla-2(12),6-dien-5α-ol (=Caryophyllenol I)	1.2	–	–
2392	Caryophylla-2(12),6-dien-5β-ol (=Caryophyllenol II)	3.5	–	–
2445	14-Oxo-calamenene	0.2	–	–
2500	Pentacosane	0.7	–	–
2503	Dodecanoic acid	11.3	–	–
2512	Benzophenone	0.5	–	–
2607	14-Hydroxy-δ-cadinene	0.9	–	–
2622	Phytol	0.7	–	–
2670	Tetradecanoic acid	4.3	–	–
2679	Manool	0.2	–	–
2700	Heptacosane	0.7	–	–
2800	Octacosane	0.2	–	–
2822	Pentadecanoic acid	0.7	–	–
2857	Palmito-γ-lactone	0.4	–	–
2931	Hexadecanoic acid	23.7	1.3	–
	Monoterpene Hydrocarbons	1.2	71.1	–
	Oxygenated Monoterpenes	1.2	4.4	62.0
	Sesquiterpene Hydrocarbons	8.9	7.3	2.7
	Oxygenated Sesquiterpenes	22.8	5.7	15.7
	Diterpenes	0.9	–	–
	Fatty acid+esters	41.0	1.3	–
	Others	6.5	–	2.4
	Total	82.5	89.8	82.8

Relative retention indices (RRI) calculated against n-alkanes % calculated from FID data.

tr Trace (< 0.1%); Ss = S. sericantha, Sg = S. gaziantepensis, and Sm = S. mardinensis.

According to our results in Table 2, monoterpene hydrocarbons (71.1%) are characterized by a high content in the oil of S. gaziantepensis while oxygenated monoterpenes (62.0%) are characterized by a high content in the oil of S. mardinensis. Fatty acid + esters (41.0%) and oxygenated sesquiterpenes (22.8%) are in large amount in the oils of S. sericantha. An earlier study^[9] published essential oil composition of 22 Stachys species (mountain tea) of which one of them is S. sericantha. It was identified to have 25 components which were accounted for 81.7% of the total composition of the oil. It consisted of sesquiterpenes (74.6%) and oxygenated sesquiterpenes (2.3%). Germacrene-D (32.4%), β-caryophyllene (23.2%), and α-cadinene (7.1%) were the main components. However, the essential oil composition of our samples was found to be quite different from those already reported. Both of S. sericantha are collected from Antalya region. But our samples are growing in Konyaaltı at 100 m and the other is growing in Kemer at 1200 m. Various factors, both endogenous and exogenous, can affect the composition of the essential oil of S. sericantha. We believe that the time of flowering and geographical and climatic factors may be very important.^[17]

According to the findings of Goren et al.,^[6,9] the oil composition of studied total 78 Stachys species in the world mainly consists of sesquiterpenes and oxygenated sesquiterpenes. The main components of the essential oil of the species were observed to be germacrene D, β-caryophyllene, cadinene, spathulenol, and caryophyllene oxide. Spathulenol and caryophyllene oxide were determined only in our S. mardinensis samples while caryophyllene oxide is found in S. sericantha. Moreover, the monoterpenes such as α-pinene, β-pinene, phellandrene, and carvacrol were also extracted from Stachys species.^[6] The monoterpenes(71.1%)α-pinene and β-pinene were also extracted from our S. gaziantepensis samples. As the main compounds, hexadecanoic acid in eight Stachys species and pulegone in one Stachys species (S. setifera) were reported by Goren et al.^[6] However, dodecanoic acid, menthol, isomenthone, and menthyl acetate are identified as the main compounds here in the oil of Stachys species for the first time.

Monoterpene compounds have therapeutic properties such as anti-inflammatory, antiseptic, antiviral, and antibacterial. In regard to cosmetics, monoterpene hydrocarbons are less valuable than oxygenated compounds in terms of their contribution to the fragrance of the essential oil. Conversely, the oxygenated compounds are highly odoriferous and, therefore, the most valuable.^[18]

Conclusion

In the Anatolia folk medicine, Stachys species are used by many people in various villages and towns for the therapeutic value of their essential oils. In this study, water-distilled essential oil of three species of Stachys (Lamiaceae) was analysed by GC-FID and GC-MS. The essential oil of S. gaziantepensis (endemic) was characterized by monoterpene hydrocarbons. Fatty acid + esters and oxygenated sesquiterpenes were found in S. sericantha (endemic) oil while oxygenated monoterpenes and oxygenated sesquiterpenes were identified in S. mardinensis oil.