Two easily accessible, natural occurring triterpenoids, betulinic and ursolic acid, were used as starting materials for the synthesis of novel cytotoxic agents. A set of 28 ethylenediamine-spacered carboxamides was prepared holding an additional substituent connected to the ethylenediamine group. The compounds were screened in SRB assays to evaluate their cytotoxic activity employing several human tumor cell lines. Betulinic acid-derived carboxamides 17–30 showed significantly higher cytotoxicity than their ursolic acid analogs 3–16. In particular, compounds 25 and 26 were highly cytotoxic, as indicated by EC₅₀ values lower than 1 μM.

A set of overall 40 carboxamides was prepared from five different natural occurring triterpenoids including oleanolic, ursolic, maslinic, betulinic, and platanic acid. All of which were derived from ethylene diamine holding an additional substituent connected to the ethylene diamine group. These derivatives were evaluated regarding their inhibitory activity of the enzymes acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) employing Ellman’s assay. We further determined the type of inhibition and inhibition constants. Carboxamides derived from platanic acid have been shown to be potent and selective BChE inhibitors. Especially the mixed-type inhibitor (3β)-N-(2-pyrrolidin-1-ylethyl)-3-acetyloxy-20-oxo-30-norlupan-28-amide (35) showed a remarkably low K_i of 0.07 ± 0.01 µM (K_i′ = 2.38 ± 0.48 µM) for the inhibition of BChE.

In this study, we report the synthesis of several amine-spacered conjugates of ursolic acid (UA) and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA). Thus, a total of 11 UA-DOTA conjugates were prepared holding various oligo-methylene diamine spacers as well as different substituents at the acetate units of DOTA including tert-butyl, benzyl, and allyl esters. Furthermore, three synthetic approaches were compared for the ethylenediamine-spacered conjugate 29 regarding reaction steps, yields, and precursor availability. The prepared conjugates were investigated regarding cytotoxicity using SRB assays and a set of human tumor cell lines. The highest cytotoxicity was observed for piperazinyl spacered compound 22. Thereby, EC₅₀ values of 1.5 µM (for A375 melanoma) and 1.7 µM (for A2780 ovarian carcinoma) were determined. Conjugates 22 and 24 were selected for further cytotoxicity investigations including fluorescence microscopy, annexin V assays and cell cycle analysis.