Composition and Antimicrobial Activity of the Essential Oil of Hypericum hyssopifolium ssp. hyssopifolium from Southeast France

Journal of Essential Oil Research: JEOR, Jul/Aug 2006 by Schwob, Isabelle, Viano, Josette, Jann-Para, Ghislaine, Bessière, Jean-Marie, Dherbomez, Michel

Abstract

The oil of Hypericum hyssopifolium ssp. hyssopifolium aerial parts was analysed by GC and GC/MS. It was found to be rich in sesquiterpenoids and characterized by spathulenol (19.5 %) and two alkanols, tetradecanol (10.2%) and dodecanol (9.3%). Furthermore, the oil was screened for its antimicrobial activity against five microbial strains. However, only a moderate antimicrobial effect of this oil was found against four of the five tested strains.

Key Word Index

Hypericum hyssopifolium ssp. hyssopifolium, Hypericaceae, essential oil composition, spathulenol, tetradecanol, antimicrobial activity.

Introduction

Among the various Hypericum species growing wild in France, Hypericum hyssopifolium Chaix is a rare species found only on calcareous and rocky soils. This species, with a SouthEuropean and West-Asian distribution, is only encountered in the south and it is only represented by the ssp. hyssopifolium Chaix (1). To the best of our knowledge, only two previous studies have been carried out on the chemistry of the species H. hyssopifolium but they were on the subspecies elongatum var. elongptum (2,3) that is not encountered in France and, no study has been done on the subspecies hyssopifolium.

Our laboratory is collecting data on the genus Hypericum to create a chemotaxonomic database of this genus. Moreoþr, as the oils of numerous species of this genus such as H. perforatum (4), H. coris (5), H. hircinum (6) and other species (7) have antimicrobial activities, we were interested on testing the biological activity of the oil of H. hyssopifolium subsp. hyssopifolium. Plants of the genus Hypericum are widely used in folk medicine (8) but only one recent study dealt with the antifungal activity of the essential oil composition of another subspecies of this species (3).

The purpose of this paper is to examine the essential oil composition of the aerial parts of H. hyssaptfolium ssp. hyssopifolium from southeastern France and determine its biological activity against three bacterial strains and two fungal strains.

Experimental

Plant material: The aerial parts of H. hyssopifolium ssp. hyssopifolium were han'ested during the flowering stage from a single location at Moustiers-Sainte-Marie(Provence-Alpes-Cote d'Azur, France) in June 1999. Fifteen individuals, representative of the population, were collected for the hydrodistillation. A voucher specimen has been deposited in the Herbarium of the Laboratory Dynamique et Ressources du Végétal, Equipe Biodiversité et Environnement of the University of Provence.

Oil isolation: The aerial parts were air-dried at room temperature (85% of moisture loss) then, powdered in a Waring blender during 3 30 s prior to hydrodistillation. An oil sample was isolated from this material by hydrodistillation for 2 h, by using a Clevenger-type apparatus. oil yield was then estimated on the basis of the dry weight of plant material. Hydrodistilled dry mass was about 40 g.

Oil analysis: Quantitative analyses of the oil were performed on a GC (Varian, model 3900GC) with a flame ionization detector (FID), equipped with a CP SIL 8CB fused silica capillary column (30 m x 0.25 mm, 0.25 µm film thickness). Oven temperature was programmed from 50°-220°C at 3°C/min, after an isothermal step at 50δC for 2 min. The carrier gas was hydrogen, with a flow rate of 0.5 mL/min. Injector and detector were heated to 220δC and 230δC, respectively. The injection volume was 0.1 µL for each of the three replicates.

Qualitative analysis was carried out on a GC/MS instrument with the following conditions: Hewlett-Packard, Model 5972, capillary GC-quadrupole MS system (EI, 70 eV) fitted with a 25 m x 0.2 mm fused silica DB-5 column. Temperature programme was 3°C/min from 60°-220°C. Helium was used as carrier gas at a flow of 1 mL/min.

Individual components were identified by comparison of both mass spectrum and their GC retention data with those of authentic compounds previously analyzed and stored in the data system (computer matching with the NBS 98K and WILEY 275 libraries). Other identifications were made by comparison of mass spectra with those in the data system libraries and cited in the literature (9). The retention indices were calculated for all constituents using an n-alkane homologous series (10). Quantitative data were calculated electronically from peak area measurements (FID) without the use of correction factors.

Antimicrobial activity: The growth inhibitory activity of various concentrations of oil solubilized with 2% of DMSO was evaluated in vitro every 30 min for 48 h by using the liquid diffusion method (11). The microbial growths were monitored in a liquid-phase system by determining the adjusted optical density (Perkin Elmer spectrophotometer) of cultures. Growth conditions were moderate agitation in a liquid Muller-Hinton medium at 37°C, for bacterial strains and, in a liquid Sabouraud medium at 28°C, for fungal strains. The strains of bacteria tested were from the Pasteur Institute: Escherichia coli (CIP: 54 127), Enterococcus hirae (CIP: 58 55) and Staphylococcus aureus (CIP: 53154). The fungal strains were from La Rochelle hospital: Candida albicans (CIP: 1180-7) and Saccharomyces cerevisiae (ATCC 28383). Eight concentrations of oil were tested in the culture medium: 0.78, 1.56, 3.125, 6.25, 12.5, 25, 50 and 100 µg/mL. Minimum inhibitory concentration (MIC) in µg/mL values were measured after 48 h incubation. The 50% growth inhibitory concentration (GIC-50) which is the oil concentration which reduces by 50% the maximum turbidity of the untreated control, was also determined and, results expressed in µg/mL.