Callus Culture and Camptothecin Production of Camptotheca acuminata - Long term culture

Callus cultures of Camptotheca acuminata Decne. were initiated from hypocotyl and leaf explants excised from 2-mo-old plants in vitro. The optimal culture medium for callus growth for both explants was woody plant medium (WPM) containing 1~3 mg L-1 NAA or 2,4-D. The calli appeared compact in NAA medium, but friable in 2,4-D medium. Comparing HCPT (10-hydroxy camptothecin) and CPT (camptothecin) contents of calli derived from 19 sources including the hypocotyls, leaves, and stems of different donor plants, we found that CPT existed in all 3 organ-derived calli but HCPT was present only in stem-derived calli. Both CPT and HCPT contents in different callus lines varied greatly. CPT and HCPT contents in a superior line were 0.0307% and 0.0021%, respectively, while those of the poor lines were close to 0. Investigating the stability of CPT content in long-term cultures, 6 calli subcultured monthly in NAA and 2,4-D media for over 3 yr were analyzed once every 6 mo. Only 1 callus line cultured in NAA medium maintained a stable CPT content of between 0.01% and 0.014%. Four lines gradually lost the ability to produce CPT, especially those cultured in 2,4-D medium. This indicates that the type of auxin used and the callus line are very critical for CPT production via cell culture.