Study Of Techniques Of Propagation In Vitro And Analysis Of Chemical Constituent On Dysosma Versipellis (Berberidaceae)

Dysosma versipellis (Hance) M.Cheng is a local med herb that belongs to the Berberidaceae. Several natural chemical have been isolated from Dysosma versioellis (HanceJM.Cheng. It is a source of highly valued podophyllotoxin which is currently being investigated for carinogenicity, cytotoxciry and antirumor activity. D. versioellis has been subjected to heavy collection from the wilds due to ever increasing demand. This report deals with the successful propagation of this species using in vitro techniques. The experiment with different explants of D. versipellis established systematically its tissue culture and plant regeneration systems. In addition, podophyllotoxin was determined by HPLC. The main results were as follows:The callus could be induced from the different explants of D. versipellis .The appearance time of callus and the rate of callus induction was different with the different explants. The petioles and leaves were better than root for callus induction., the rate of the callus induction is 75.6%,the optimal medium is MS + 2,4-D 0.5mg/L + KT 0.2mg/L.The optimal medium of callus differentiation is MS+ BA 0.5 mg/L + NAA 0.1 mg/L +GA 0.2 mg/L, rate of the callus differentiation could reach 59.2%. It was much easily to differentiate roots than buds from the callus. When we use the buds as explants to induce the organgenesis in vitro, the cluster buds formed directly at the basal parts of explants on MS medium with ZT 1.0mg/L+ GA 0.5 mg/L. Plantlets could grow healthy on this medium. The optimal medium for rooting was 1/2MS+ NAA 0.5+ AC 0.5 g/L, the rate of rooting can reach 100% .It is necessary to add some activated charcoal (0.5 g/L) into medium on themultiplication of the buds and roots. The effects of activated charcoal could be attributed to providing a dark environment in the medium, adsorption of certain inhibitory substances in medium.In order to improve the rate of plantlets survival, we covered plastic film on the plantlets when they transferred from the bottles. Above 70% plantlets could survival by this way. Combination of peat and perlite with a ratio of 1:1 was used for transplant substances. The plantlets could be transferred from the greenhouse to the field for one month.Content of podophyllotoxin was analyze and compare among callus, root differentiated from callus and the rhizome of the native D. versipellis by HPLC. Thecontent of podophyllotoxin in callus (20% compared with the native rhizome) was lower than that in roots cultured on the medium (25% compared with the native rhizome).The content of podophyllotoxin increased on differentiation medium contained with phenylalanine or amylose when the time passed, but it increased slowly. Content of podophyllotoxin in cultures on the medium supplemented with phenylalanine or amylose was lower than that on CK medium. The maximal product yields of podophyllotoxin in cultures on the medium supplemented with phenylalanine or amylose is up to 69.2% and 46 % of the native rhizome, which could approach 91.3% of native rhizome on CK medium for 3 months.