The role of centrifugal partition chromatography in the removal of β-asarone from Acorus calamus essential oil

Abstract

Β-asarone is a phenylpropane derivative present in the rhizomes of Acorus calamus, that was proved to exhibit toxic effects in humans. Because of its presence the whole plant that is commonly used in traditional medicine for its sedative, anti-inflammatory, neuroprotective and other properties has limited application nowadays. In the study, qualitative and quantitative analysis of a collection of nine essential oil (EO) samples of European and Asian origin was performed. The final content of β-asarone in the tested samples ranged between 0.265 and 1.885 mg/mL. Having in mind a possible application of the EO as a biopesticide, this research aimed at the development of CPC-based purification protocol that could help remove β-asarone from EO. It was proved that the biphasic solvent system composed of n-hexane/EtOAc/MeOH/water, 9:1:9:1 (v/v/v/v) was capable of the removal of the toxic constituent in the CPC chromatograph operated in the ascending elution mode with 2200 rpm and a flow rate of 5 mL/min. The chromatographic analysis that lasted only 144 min effectively separated β-asarone (purity of 95.5%) and α-asarone (purity of 93.7%) directly from the crude Acorus calamus rhizome EO.

Figure 1

The isomers of asarone found in Acorus calamus EO.

Figure 2

The total ion chromatogram (HPLC–MS) in positive ionization mode of the commercial essential oil sample obtained from the rhizomes of Acorus calamus.

Figure 3

The β-asarone concentration in the analysed EOs.

Figure 4

The linear map of original variables in the space of the two relevant dimensions of the discussed problem (A) and the linear map of samples in the same coordinate system (B).

Figure 5

Cluster dendrogram, drawn on the basis of the Euclidean distances between studied samples, concerning relative concentrations of α-, β- and γ-asarone.

Figure 6

The CPC chromatogram (CPC-DAD) of the essential oil obtained from the rhizomes of Acorus calamus by hydrodistillation in the Clevenger apparatus. Monitored wavelength, λ = 254 nm.

Figure 7

(A) HPLC-DAD chromatogram of the standard solution of Acorus calamus EO and UV spectra of α- and β-asarone; (B) HPLC-DAD chromatogram and UV spectrum of the α-asarone fraction obtained by CPC separation; (C) HPLC-DAD chromatogram and UV spectrum of the β-asarone fraction obtained by CPC separation.

Figure 8

Map showing locations and origins of Acorus calamus accessions harvested from different regions of Poland. (S1–S3)—sample code, N—North, BY—Belarus, CZ—Czech Republic, DE—Germany, RU—Russian Federation, SK—Slovakia, UA—Ukraine. The base map of Poland and the neighbouring countries was obtained from http://diva-gis.org/data.