Hairy root cultures of Cassia obtusifolia clones transformed with Agrobacterium rhizogenes strain 9402 were established to investigate the anthraquinone production. Seven anthraquinones, together with betulinic acid, stigmasterol and sitosterol were isolated from the hairy roots. Their chemical structures were elucidated on the basis of chromatographic and spectral data. The effects of culture conditions and rare earth element Eu(3+) on the production of six free anthraquinones have also been investigated. It was found that changes of the elements in the culture medium and addition of rare earth element Eu(3+) can greatly influence the contents of free anthraquinones in the hairy roots.