Chemical Composition and Microbial Activity of the Essential Oil of Calamintha nepeta (L.) Savi ssp. nepeta var. subisodonda (Borb.) Hayek from Serbia

Journal of Essential Oil Research: JEOR, Nov/Dec 2005 by Kitic, Dusanka, Stojanovic, Gordana, Palic, Radosav, Randjelovic, Vladimir

Abstract

An oil of Calainintha nepeta (L.) Savi ssp. nepeta van suhisodonda (Borb.) Hayek obtained by hydrodistillation was analyzed by GC and GC/MS. Eleven constituents were identified (96.5%). The main constituents in the oil were pulegone (75.5%), piperitenone oxide (6.0%), menthone (5.3%) and menthol (4.3%). The microbial activity of the oil was screened against Aspergillus niger, Escherichia coli, Staphylococcus aureus, Salmonella enteritidis, Bacillus subtilis and Pseudomonas aeruginosa. It was found that all mentioned microorganisms were sensitive to the oil.

Key Word Index

Calamintha nepeta ssp. nepeta var. suhisodonda, Lamiaceae, essential oil composition, pulegone, menthone, piperitenone oxide, menthol, antimicrobial activity.

Plant Name

Calamintha nepeta (L.) Savi ssp. nepeta var. stibisodonda (Borb.) Hayek [syii.: Calamintha tardarensvs Pavl. non Silic, Calamintha subnuda N. Randj. non (Wald. & Kit.) Host.]

Previous Work

Calamintha species has been the subject of intensive studies over the last few decades (4-45), particularly the oil of C. nepeta due to a great diversity in a chemical composition, microbial and fungicidal activities (4-31). It can be seen that the oil of C. nepeta has been found to be rich in monoterpenes. In all studies on oils obtained from wild growing species of the Calamintha genus, the compounds of monoterpenoids were dominant, especially the oxygenated p-menthane-type monoterpenoids. The monoterpenoids of the pinane, thujane and camphanetypes were present only as minor components or they were not detected. Two atypical oil samples have been characterized; one with isopinocamphone as the major component (38), and the other with carvone and 1,8-cineole as main components (22). The contents of sesquiterpenoids were present up to 5% while the other compound were very low (less than 2%).

Source

The aerial parts of flowering plant were collected in August 2000 on Vratarnica mountain (Knjazevac, South Serbia). A voucher specimen has been deposited in the Herbarium Moesicum Doljevac (HMD; No 395) (1).

Plant Part

Dried and pulverized aerial parts of the plant (100 g) were hydrodistilled for 2.5 h using a Clevenger-type apparatus (2). The oil was extracted from the distillate with Et^sub 2^O and then dried with anhydrous Na^sub 2^SO^sub 4^. After filtration, the solvent was removed by distillation under the atmospheric pressure and pure yellow oil, 1.0 m L, was kept at 4�C until analysis.

Present Work

The oil analysis was performed using GC and GC/MS. Constituents of the oil were identified by comparison of their mass spectra to those from the MS library (Wiley275.1) using a computer search and literature. For the purpose of the quantitative analysis the area percentage obtained by FID was used as the base without the use of correction factors.

Experimental

GC: The GC analysis of the oils was carried out on a GC HP-5890 II apparatus, equipped with the split-splitless injector, SPB 5 capillary column (30 m x 0.25 mm, 0.25 �m film thickness) with helium as the carrier gas (1 mL/min) and fitted to FID. Operating conditions: injector temperature 250�C, temperature program isothermal 50�C for 3 min, then 50�-250�C at 5�C/min and finally isothennal at 250�C for 15 min.

GC/MS: GC/MS analyses were performed on a Hewlett Packard apparatus, Model 5890, Series II at 70 eV coupled with a mass selective detector 8MSD 5971A, under the same gas-chromatograph conditions.

Identification procedure: The constituents were identified by comparison of their mass spectra to those from MS library Wiley 275.1. Obtained results were correlated with retention index(3). The areapercentage was obtained electronically from the GC-FID response without the use of internal standard or correction factors.

Antimicrobial activity: The oil was tested in vitro using the diffusion discs method with the following microorganisms: Aspergillus niger ATCC 16404, Escherichia coli 95 TORLAK, Staphylococcus aureus ATCC 6538, Salmonella enteritidis ATCC 13076, Bacillus subtilis 201 and Pseudomonas aeniginosa ATCC 9027.

Substrate for the bacteria was Antibiotic Medium 1 (Difco Laboratories Detroit Michigan USA), for the fungus: Tripton Soja Agar (Torlak, Beograd). The ethanolic solution of the oil (60 �L) in the dilutions of 1:101:201:30 and 1:50 (oil:abc. ethanol) was put on discs Antibiotica Test Blattchen (Schleicher and Schuell, Dassel, Germany; diameter 12.7 mm). The discs then were put on the antibiotic medium sowed with microorganisms and kept at 37�C. After 18 h the activities were determined on a Fisher-Lilly Antibiotic Zone Reader (Ficher Scientific Co. USA) by measuring the inhibition diameter around the discs.

Results and Discussion

Compared with other Calamintha species (3-45), and following the grouping of Lamiacea species, proposed by Kokldni et al., in oil-rich (> 2%), oil-intermediate (0.5-2%) and oil-poor (

The results of the oil analysis of C. nepeta ssp. nepeta var. subisodonda can be seen in Table I. About 96.3% (11 components) of the oil has been identified. Pulegone (75.5%) was the main compound of the oil; the other compounds with noticeable percentage in the oil were piperitenone oxide (6.0%), menthone (5.3%) and menthol (4.3%). According to the literature, the chemical composition of C. nepeta oil is independent of the nature of the subspecies nepeta or glandulosa (28). Generally, there are two main C. nepeta oil types. The first oil type is characterized by the dominance of pulegone along with menthone and menthol and/or their isomers, piperitenone and piperitone and their oxides (4,8,12,13,15,17,20-24,29,30,31), and second type is distinguished by dominance of piperitenone oxide and/orpiperitone oxide (11,16,18,26,27). By comparison with those types the oil of C. nepeta ssp. nepeta var. subisodonda belongs to the pulegone-rich group, which is the most common one.