The alkaloid fraction from Buxus sempervirens leaves shows strong in vitro activity against Plasmodium falciparum

JB Althaus; G Jerz; P Winterhalter; M Kaiser; R Brun; TJ Schmidt

doi:10.1055/s-0033-1351906

Planta Medica, Inhaltsverzeichnis

The alkaloid fraction from Buxus sempervirens leaves shows strong in vitro activity against Plasmodium falciparum

JB Althaus ¹, G Jerz ², P Winterhalter ², M Kaiser ³, R Brun ³, TJ Schmidt ¹

¹University of Münster, Institute of Pharmaceutical Biology and Phytochemistry (IPBP), Corrensstraße 48, D-48149 Münster, Germany
²Technical University of Braunschweig, Institute of Food Chemistry, Schleinitzstraße 20, D-38106 Braunschweig, Germany
³Swiss Tropical and Public Health Institute (SwissTPH), Socinstraße 57, CH-4002 Basel, Switzerland

Abstract

In ethnopharmacology, Buxus sempervirens (European Box, Buxaceae) is known as a plant with antimalarial activity [1,2]. On different continents, species of the genus Buxus were used in different preparations to cure malaria [3].

In the course of our ongoing screening of plant extracts for antiprotozoal activity, a CH₂Cl₂ extract from the leaves of B. sempervirens showed selective in vitro activity against the NF54 strain of Plasmodium falciparum (IC₅₀= 2.79 and 20.2 µg/mL for antiplasmodial and cytotoxic activity, respectively).

Separation of the extract by acid/base extraction into a basic and a neutral non-polar fraction (extraction with acetic acid/water followed by neutralization and extraction with CH₂Cl₂) led to a much more active and equally selective fraction with alkaloids (IC₅₀= 0.36 and 7.3 µg/mL, respectively) while the fraction of non-polar neutral constituents was markedly less active than the crude extract (Table 1). Thus, the activity of the crude extract can clearly be attributed to alkaloid constituents. Identification of the main triterpene-alkaloids and characterization of the complex pattern of this alkaloid fraction was performed by UHPLC/+ESI-QTOF MS analyses.

*Tab. 1:* *In vitro* activity of CH₂Cl₂ extract of *B. sempervirens* leaves and its main fractions against *P. falciparum* and cytotoxicity data against L6 rat skeletal myoblasts. Data represent IC₅₀ values in µg/mL. Chloroquine and Podophyllotoxin are positive controls.
	P. falciparum NF54 strain	L6 cells
CH₂Cl₂ extract	2.79	20.2
Alkaloid fraction	0.361	7.31
Lipophilic fraction	7.76	33.9
Chloroquine (pos. control)	0.003	-
Podophyllotoxin (pos. control)	-	0.008

ESI-MS/MS target-guided larger scale preparative separation of the alkaloid fraction (4 g) was performed by 'high-speed- ' and 'spiral coil-countercurrent chromatography'. Subfractions containing all representative alkaloids are currently under evaluation for their in vitro antiplasmodial activity. The results of these tests will enable us to identify and isolate the most active alkaloids for further testing and evaluation.

References:

[1] Leporatti ML et al., Journal of Ethnopharmacology, 14, 53 – 63, (1985).

[2] Orhana IE et al., Industrial Crops and Products, 40, 116 – 121, (2012).

[3] Athar A in: The Alkaloids, Cordell AC (Ed.), Vol. 66, pp.191, Elsevier, 2008

This work is part of the activities of ResNetNPND: http://www.uni-muenster.de/ResNetNPND/