Studies on Cassytha filiformis from Benin: isolation, biological activities and quantification of aporphines

Cassytha filiformis (Lauraceae), a widely distributed parasitic plant, contains several aporphine alkaloids and is often used in African folk medicine to treat cancer, African trypanosomiasis and other diseases. Samples of C. filiformis from Benin were studied. We isolated and identified 6 aporphines and analysed the in vitro cytotoxic properties of four of them on different cancer and non-cancer cell lines. The major alkaloids (actinodaphnine, cassythine, and dicentrine) were also shown to possess antitrypanosomal properties in vitro on Trypanosoma brucei brucei. In order to elucidate their mechanism of action, the binding mode of these molecules to DNA was studied. The results of the optical measurements showed that these three alkaloids bind to DNA and behave as typical intercalating agents. Biochemical experiments also indicated that they interfere with the catalytic activity of topoisomerases. These interactions with DNA may explain, at least in part, the effects observed on cancer cells and on trypanosomes. In order to quantify these alkaloids in samples from different origins, a sensitive and accurate procedure based on an alkaloid extraction coupled to a HPLC-UV-MS determination has been developed for the separation and quantification of the major aporphines in C. filiformis. The extraction parameters and the liquid chromatography conditions were optimized in order to improve the selectivity of the method, which was completely validated using cassythine, one of the major aporphines in our samples, as reference standard. This procedure was successfully applied to the determination of these pharmacologically interesting aporphines in seven different batches of C. filiformis. The results showed variations in the total alkaloid content in samples, which ranged from 1.1 to 4.3%.