Composition of the Essential Oils from the Flowers and Leaves of Salvia sclarea L. (Lamiaceae) Cultivated in Slovak Republic

Composition of the Essential Oils from the Flowers and Leaves of Salvia sclarea L. (Lamiaceae) Cultivated in Slovak Republic

Farkas, Pavel

Abstract

The chemical composition of the essential oils obtained from the flowers and leaves of Salvia sclarea L. ( Lamiaceae) cultivated in Slovak Republic have been studied by GC and GC/MS methods, which resulted in the identification of 34 and 23 components, respectively. The major constituents in the flower oil were linalool (18.9%), linalyl acetate (13.7%), sclareol (15.7%), α-terpineol (6.5%), germacrene D (5.0%) and geranyl acetate (4.3%), while the major components in the leaf oil were germacrene D (28.8%), bicyclogermacrene (12.5%), spathulenol (10.1%), caryophyllene oxide (6.2%) and α-copaene (6.0%).

Key Word Index

Salvia sclarea, Lamiaceae, essential oil composition, linalyl acetate, linalool, germacrene D, bicyclogermacrene, spathulenol.

Introduction

Salvia, the largest genus of the Lamiaceae family, includes about 900 species wide spread over the world. Salvia sclarea is of economical importance as flavoring agent in the food industry, perfumery and cosmetic industries (1). Recent studies have shown that the clary sage oil has some interesting biological properties. The oil inhibits hyperalgesia and has appreciable anti-inflammatory effect in experimental animals. The oil has been used where inflammatory disease is present in the oral cavity (2). Some terpenoids isolated from whole plants, roots, leaves, flowers and the oil of S. sclarea has been shown to exert significant microbiostatic activity against Staphylococcus aureus, Escherichia coli, Staphylococcus epidermis, Candida albicans and Proteus mirabilis (3-6). The chemical composition of the clary sage oil has been reported by different researchers (6-19) using GC and GC/MS methods, and retention indices to characterize the oil. Analyzed oils usually contained high quantities of monoterpenic esters and alcohols.

The most important components in the oils were alcohols: linalool, α-terpineol, and esters: linalyl acetate, α-terpinyl acetate, geranyl acetate. However, the amount of these components varied very widely (linalool 2.6-32.8%, α-terpineol 1.5-47.4%, linalyl acetate 8.2-81.1%, and α-terpineol acetate trace-22.1%). The clary sage oil from plants cultivated in Sardinia (Italy) (6) was characterized by higher content of alcohols, mainly α-terpineol (47.4%), while the contents of linalyl acetate and α-terpinyl acetate were lower (12.7% and 22.1%, respectively). These authors have also studied the antimicrobial activity of the oils concerning their use in the pharmaceutical preparations for local application. The composition of clary sage oils from flowers and leaves has been reported in papers (7,8). In these studies, the chemical composition of the lab-distilled oil within a four year span was compared, and oxidation products were found in the older oil. Carruba et al. (9) found that oil from the inflorescences of S. sclarea was rich in linalyl acetate (35-53%), linalool (26-29%) and germacrene D (4-11%), whereas the oil from the leaves contained germacrene D (69%) as the main component. Lalande (10) compared the chemical composition of French and American clary sage oils. The French oil contained linalool and linalyl acetate in amount 10-20% and 60-70%, respectively. The American oil contained 25% of linalool and 55% of linalyl acetate. Chemical composition of the clary sage oils isolated from plants cultivated in United States, France and Russia are reported in a review (11). The content of linalool in United States, French and Russian clary sage oil was 20.3-28.5%, 9.0-16.0% and 10.4-19.5%, respectively, while the content of linalyl acetate was 44.9-53.4%, 49.0-73.6% and 45.3-61.8%, respectively. Elnir et al. (12) determined 23.5-31.0% of linalool and 34.4-44.6% of linalyl acetate in a commercial clary sage oil. Dzumayev et al. reported the composition of the cultivated clary sage oils produced over four separate seasons (13). The amounts of linalool and linalyl acetate were 22.0-36.0% and 25.0-51.0%, respectively. These authors also investigated the composition of oils produced from different plant parts (inflorescence, calyces) both wild and cultivated plants with white and rose-violet bracts. In reference 14 authors determined 33.2% of linalool and 16.9% of linalyl acetate in clary sage oil produced from wild plants grown in Spain. Souleles et al. (15) identified 72 components in the clary sage oil from plants collected in wild at Thessaloniki (Greece), where amount of linalool and linalyl acetate was 17.2% and 14.3%, respectively. Esteban et al. (16) compared the composition of clary sage oil obtained by automatic thermal desorption (ATD) with that of produced by simultaneous distillation-extraction (SDE). The composition of these oils was quite different. The contents of linalool, linalyl acetate and sclareol obtained by ATD method were 3.9%, 15.2% and 74.2%, however the contents of the same compounds in the oil obtained by SDE method was 16.9%, 25.0% and 20.5%, respectively. Ronay et al. (17) compared composition of the clary sage oil and volatile concentrate obtained by supercritical fluid extraction (SFE) of clary sage. Authors determined 14.9% of linalool and 10.3% of linalyl acetate in the oil and 0.9% of linalool, 8.2% of linalyl acetate and 50.0% of sclareol in the extract. Foray et al. (18) found that the level of linalyl acetate in French clary sage oil was higher (81.1%) than that of normally occurs in commerce. Demirci et al. (19) reported linalool and linalyl acetate as the major components in the oil obtained by hydrodistillation of both wild and cultivated forms of S. sclarea. Pesic and Kankovic (20) analyzed clary sage flower oil through three different phases of herbal development: at full blossom, at seed formation, and at full seede maturity, in all parts of the inflorescence. This oil contained linalool and linalyl acetate 15.4% and 67.4%; 12.9% and 74.1%; 9.4% and 79.1%, respectively.

In this study, the composition of the flower and leaf oils of Salvia sclarea L. cultivated in Slovak Republic was determined.

Experimental

Plant material: Salvia sclarea was grown on the clay soils of Nitra locality in Slovak Republic. The locality of cultivation is situated in the mild climatic zone. Salvia sclarea was harvested at the full bloom stage. After collection the samples of flowers and leaves they were separately air-dried. A voucher specimen was lodged in Herbarium of the Department of Botany, Faculty of Pharmacy, Comenius University in Bratislava.

Oil isolation: Samples of dry flowers and leaves were separately subjected to hydro distillation for 4 h in accordance to European Pharmacopoeia(21). The isolated oils were dissolved in hexane and dried over anhydrous sodium sulfate.

Chromatographic analysis: Identification of the volatile components was carried out using a Hewlett Packard HP 5971A mass selective detector directly coupled to a gas chromatograph HP 5890 Series II. A capillary column Ultra 2,25 m x 0.20 mm, 0.33 µm film thickness (Hewlett Packard, USA) was used. The temperature program was as follows: 40°-250°C at 3°C/min. Injection port temperature was 250°C. Helium was used as a carrier gas with split ratio 1:50. Mass spectra were recorded at ionization energy (EI) 70 eV.

For determination of retention indices and quantification of the oil components a Hewlett Packard (HP 5890 Series II) gas chromatograph, equipped with a flame ionization detector was used. The percentage composition was calculated from FID area values without the use of correction factor. The oils were analyzed on the Ultra 2, 50 m x 0.32 mm, 0.5 µm film thickness (Hewlett Packard, USA) capillary column. The column was operated at a temperature program from 35°-250°C with a gradient of 2°C/min. The linear velocity of the carrier gas hydrogen was 36 cm/s (measured at column temperature 143°C). The linear retention indices (LRI) were calculated after Van den Dool and Kratz equation (22) using n-alkanes C^sub 9^-C^sub 23^.

The oil components were identified by comparison of their mass spectra and retention indices with that measured on authentic compounds and their mass spectra with ones from the library of spectra NIST, Wiley 138 and INRA Mass (LRSA, Dijon, France).

Results and Discussion

The contents of the oil from flowers and leaves cultivated in Slovak Republic were 0.11-0.27% and 0.29-0.54% (v/w), respectively on a dry weight of the plant material. Thirty-four and 23 components in the flower and leaf oils were identified by mass spectrometry and confirmed by their relative retention indices. Their relative percentages comprised 89.1% and 90.6%, respectively, of the total oils. Relevant qualitative and quantitative differences between both analyzed oils were found. Clary sage flower oil was characterized by high content of linalool (18.9%), sclareol (15.7%), linalyl acetate (13.7%), α-terpineol (6.5%), germacrene D (5.0%) and geranyl acetate (4.3%), whereas in the leaf oil germacrene D (28.8%), bicyclogermacrene (12.5%), β-caryophyllene (12.3%), spathulenol (10.1%), caryophyllene oxide (6.2%) and α-copaene (6.0%) were found as the main components. As is shown in Table I, linalool, linalyl acetate, sclareol, α-terpineol and geranyl acetate, which were the main components of the clary sage flower oil, were not found in the leaf oil. Contrary to that, clary sage leaf oil contained high amounts of germacrene D, bicyclogermacrene, β-caryophyllene and spathulenol. The smaller amounts of myrcene, limonene, (Z)-β-ocimene, (E)-β-ocimene, α-terpinolene, nerol, α-eudesmol, β-eudesmol, valencene, 13-epi-manoyl oxide were identified in the clary sage flower oil, but they were not found in the leaf oil. Contrary to that, the small amounts of δ-elemene, α-cubebene, viridiflorol and α-cadinol were identified in clary sage leaf oil, but they were not found in the flower oil. Similar qualitative and quantitative differences between the flower and leaf oil compositions were also reported in papers (7,8,9). Comparing the data presented and that from literature (6-20) it is evident, that the content of linalool and linalyl acetate in the clary sage oils from diverse sources is different. We suppose that this variability may be caused by different partial hydrolysis of linalyl acetate during the hydrodistillation as well as by the origin of the plant, locality of its cultivation, environmental conditions, developmental stage and used part of the plant.

Acknowledgement

This work was supported by grant No. 1/7282/20 and grant No. 1/8215/01 from Scientific Grant Agency (VEGA) of Slovak Ministry of Education.

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Pavel Farkas

Food Research Institute, Bratislava, Slovak Republic

Magda Hollá, Jozef Tekel* and Slavomír Mellen

Department of Pharmaceutical Analysis and Nuclear Pharmacy, Faculty of Pharmacy, Comenius University, Odbojárov 10, SK-83232 Bratislava 3, Slovak Republic

Stefánia Vaverková

Department of Pharmacognosy and Botany, Faculty of Pharmacy, Comenius University, Bratislava, Slovak Republic

* Address for correspondence

Received: May 2002

Revised: December 2003

Accepted: January 2004

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