Research On The Effects Of The Growth And Alkaloid Content Of Fritillariac Ussuricnsis's Callus Treated With Physical And Chemical Factors

Pingbeimu, Bulbus Fritiilariae ussuriensis, is one of the four Fritillaria medicinal materials (Bulbus Fritillariae thunbergii,Bulbus Fritillariae cirrhosae,Bulbus Fritiilariae ussuriensis,Bulbus Fritillariae pallidiflorae). Its original plant is the Fritillaria ussuricnsis Maxim belong to Fritillaria genus, Liliaceae family, and the medicinal part is its bulb whose functions as a traditional Chinese medicine (TCM) are cleaning up fever, nourishing lung, expectorant, releasing stasis. Pingbeimu is one of common and great consumed TCM in China, it is usually used to cure upper respiratory disease, such as tracheitis, tuberculosis, pneumonia and chincough, and there are plenty of compounding prescriptions contained Pingbeimu. In recent years, the wild resource provide too little amount of the Pingbeimu to meet the demand. Similar to most of our country's cultivated TCM, it is mainly cultivated with individual, that result in the following problems: spreading around, low techniques, short of standard and intensive farming, the Pingbeimu quality, thus, has been not stable and difficult to supply, etc.The research aim to those problems, to induce the Pingbeimu's effective component, the alkaloid, with the advanced technique of tissue culture, and develop the producing media of high effective component. The methods may get rid of the restriction of the field conditions, climate conditions, techniques in traditional cultivation.The experiment is based on biotechnological theory, the optimum medium is screened first from the four media of MS, B₅, N₆ and H according to the induced ratio of the bulb's callus in the media. After the optimum medium is screened, the Orthogonal experimental design was applied to study the effects of the proportion and concentration of the plant hormones to the callus inducing and the subculture, and it is fatherly researched that what kinds of effects of different carbohydrates, nitrogen substances, phosphate, light or dark culture and adding precursor substances they are to the alkaloid content in the callus. During the culture, the main targets were biomass and the total alkaloid content in the callus, the optimum cultural conditions of the Pingbeimu callus growth and the total alkaloid production has been identified. It is the first reported that adding precursor substances induced the alkaloid production in the callus.After investigating the targets, such as the fresh or dry weight, the total alkaloid content in the callus, the whole results indicate: 1. the optimum Fritillaria callus induced medium is MS+ NAA0.5mg/L+2, 4-D1.0mg/L+BA1.5mg/L, the induced rate is 68.7%; 2. the optimum Fritillaria callus production medium is MS+2.4-D0.5mg/L+ BA0.05mg/L, and the optimum sugar concentration is 4%, NH4+:N03-=1:1, the total nitrogen concentration is 60 mmol/L,KH2P04 concentration is 1.5 mmol/L; 3. It is first discovered that the three kinds of amino acids as precursor substances could induce the alkaloid content rise in the callus, the increasing abilities as the following order:â‘ L-tyrosine 10mg/L could induce the alkaloid content in the callus to 0.4235%, 41.2% more than control;â‘¡L-phenylalanine 10mg/L could induce the alkaloid content in the callus to 0.4122%, 37.4% more than control;â‘¢L- tryptophan 20mg/L could induce the alkaloid content in the callus to 0.4025%, 34.2% more than control.