Study Of Rapid Propagation In Vitro Of The Endangered Medicinal Herbs Notopterygium Incisum

Notopterygium incisum Ting ex H.T.Chang(Apiaceae)is a perennial herbaceous herb endemic to China.Investigations of phytochemication have revealed that both rhizomes and roots of N.incium contained are used in the treatment of rheumatism strong cold,headache,rheumatism bi pain,shoulder pain.However,increased demand for its medicinal purpose inn recent years has led to excessively excavate and deforestation,natural habitat of N.incisum have been severely damaged.Though natural regeneration of N.incisum takes place through seed,the seed germinaton rate is extremely low due to its long time for dormancy breaking.Propagation of N.incisum could be developed by vegetative propagation through stem cutting,but the method is time-consuming and requires large part of the plant as explants,massive loss of the species could be happened in its natural habitat.An urgent need to develop a reproductive propagation protocols was posed.Leaves of N.Incisum plants collected from Xiaojin County of Sichuan Provincewas used as explant,aimed to develop an efficient propagation protocol to get a large number of plants within a short time through study of callus induction,callus proliferation,shoot induction,rooting and acclimatization.The results of the study are as the followings:1.Leaf is the best explant for in vitro propagation of N.Incisum.The best suitable sterilization method is using 75% alcohol for10 s combined 0.1% mercuric chloride for 10 min,the sterilization rate can reach 95%.2.Best callusing was achieved from the leaf of N.incisum in natural habitat of culture in MS medium supplemented with 1.5mg/l 2,4-D and0.2mg/l 6-BA,callus induction rate was 84.44%.When used the leaves in vitro as the explant,the optimum medium for callus induction was MS+3.0 mg/l NAA +1.0mg/l 6-BA,Callus induction rate was78.922%.And the Callus is embryonic callus,and can be differentiation form buds.3.Maxium number of shoots(4.7),shoot induction rate(76.85%)and shoot weight(1.308 g)were found on MS medium with 1.5mg/l 6-BA and 0.2mg/l NAA derived from leaf-derved callus.While,the optimum medium for direct shoot organogenesis was MS supplemented with 2.0mg/l 6-BA and 0.2mg/l NAA,the shoot induction rate was 36%,the average number of shoot 1.5.4.The IBA concentration is 0.6 mg/l,the rooting rate was92.28%.Root length maximum up to 2.14 cm at the 0.4mg/l IBA.While the increasing rate of base callus increases with the increase of concentration of IBA.The increasing rate of base callus up to 93.61% at the 1.0mg/l IBA.Considering the roots induction rate and increasing rate of base callus,the optimum IBA concentration was 0.2mg/l.Low concentration of activated carbon for adventitious roots induction rate had no significant effect,but higher concentration showed a inhibition action.While,low concentration of activated carbon for root length and root number has a significant role in promoting.As has been said above,we can concluded that the best medium was 1/2MS + IBA0.2mg/l+ activated carbon 0.5 g/l,and the root induction rate was 80.58%,average root number was 17.59,root length is 3.07 cm.5.A month after acclimatization,new root and leaves was developed after 30 d late.The survival rate of tissue culture seedling with basal callus increased was 47.8%,while the survival rate of tissue culture seedling with basal callus has not increased was 83.8%.6.Tests for the notopterol and isoimperatorin content in medicinal part of 3 years old cultured seeding plant and seed seeding plant,wild plant is conducted,it revealed seed seeding plant > cultured seeding plant > wild plant,and the difference was significant.There is no significant difference between volatile oil content in three strains planting.The notopterol and isoimperatorin content in the undergroundpart of three strains planting samples were significantly higher than that of aboveground part,and notopterol content were higher than isoimperatorin in the underground part of three strains planting.The above results show that the cultured seeding under the condition of artificial cultivation,its chemical quality is higher than wild plants and satisfy the standard of pharmacopoeia.Therefore,cultured seeding can be used as seedling sources for artificial planting.