Studies On Production Of The Inophyllums B Cell Cultures Of Calophyllum Inophyllum Lin

Inophyllums is a novel, naturally occurring, nonnucleoside inhibitor of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase first isolated from a tropical tree (Calophyllum inophyllum Linn). Investigation suggested that the resource of C. inophyllum be not abundance, distributed from Qongshang to Wangning city in the east of Hainang island. It is significance to develop inophyllums by cell culture of C. inophyllum instead of extracted directly from the trees.The chemical constituents of the essential oil of C. inophyllum Linn, have been analysed by GC-MS and 40 components with mainly the alcohol compounds and advanced alkane were identified. The measure technique of Inophyllums with HPLC was established by using Inophyllum B as standard.The calli could be induced from young leaves, stem and radicles of C. inophyllum seedling on MS medium but could not from endosperms. Of the explants above, the highest calli-inducing frequency with 95.6% was observed from stem. The calli growth was in relation to the concentration of inorganic salts in the medium. The best basic medium for the calli growth was B₅ following with MS and White. 1200 lx all-day light was the best light condition for the calli inducing with frequency of 100%. Relatively better growth responses were displayed by 2,4-D/6-BA ratios range from 6/1 to 4/1.Compared the biomass and inophyllums-containing,the high producing calli line CR2 with inophyllums concentration of 1.480mg/gDW was screened. The medium for CR2 growth was further optimized with quadratic orthogonal rotation combination device and the optimum medium Mo (basic B5 medium supplied with 2.0mg/L of 2,4-D, 0.32mg/L of 6-BA and 35g/L of sucrose) was obtained.In Mo medium, the process curve of CR2 cell suspension culture and inophyllums yield showed typical 'S'. The suspension culture process of CR2 cell could be divided into five phase: lag phase, exponential growth phase, linear growth phase, stationary phase and recession phase. The lag phase was longer and peak value of CR2 cell biomass and inophyllums production were occurred at the 21^st day and 24^th day respectively. The growth and inophyllums yield of CR2 cell were affected by beginning sucrose concentration, ratio of NH₄⁺ to NO₃^- and total nitrate. 3.5% of beginning sucrose concentration, 2/25 of ratio of NH₄⁺to NO₃^- and 27mmol/L of total nitrate were the most favorable combination to CR2 cell growth and inophyllums...