Analysis of Artemisia annua extracts and related products by high performance liquid chromatography-tandem mass spectrometry coupled to sample treatment miniaturisation.

ABSTRACT

Artemisinin, the main antimalarial compound of Artemisia annua L., is currently attracting increasing interest for its antiproliferative properties, but its content is highly variable, depending on several genetic, environmental and processing conditions. Aim of the present study is to analyse the artemisinin content in different plant extracts, to test their in vitro activity on cell proliferation and then to correlate these data to the active principle concentration. For this purpose, an innovative miniaturised sample pretreatment strategy based on microextraction by packed sorbent (MEPS) was developed and coupled to an original advanced method based on liquid chromatography with diode array detection and tandem mass spectrometry (LC-DAD-MS/MS). The method was fully validated, granting consistent data. Good linearity was found over a suitable concentration range, i.e. 5-1000ng/mL. Extraction yields (>85%), precision (RSD < 3.5%) and accuracy (recovery 88-93%) were all within acceptable levels of confidence. After validation, the method was successfully applied to the determination of artemisinin in A. annua extracts. Analyte content was widely variable (up to twenty-fold) according to the starting material and the extraction procedure, ranging between 5.9µg/g and 109µg/mL. The cytotoxic activity of all analysed extracts was also tested on human leukemic cells by viable cell count and cell cycle analysis. Artemisinin concentrations and biological activity were carefully evaluated and the observed antiproliferative effects varied according to artemisinin content in each extract type. This highlights the need to quantitatively analyse the main active constituent of plant extracts and the obtained data have shown to be promising for the choice of the related herbal product dosage.