Studies On Immobilized Cell Of Cephalotaxus Mannii System

The application of immobilized cell of Cephalotaxus mannii to producing harringtonine is one of the most important problems in the study of cells of Cephalotaxus mannii. This article investigated the establishment of immobilized cell of Cephalotaxus mannii and the growth characteristics of immobilized cell. The optimization of the immobilization conditions was studied. And the effects of the age of cell on the cell growth and harringonine production of Cephalotaxus mannii immobilized cells was studied. Main results were as follows.1.Immobilization conditions was to be basically established by references and preexperiment. The appropriate concentration of sodium alginate was 20%. The appropriate concentration of sodium calcium chloride was 0.02g/mL. The inoculation capacity suitable to the immobilized particle is O.lOg/mL. The density of embedding was 0.2g/mL. The speed of rotary shaker was 120r/min. The age of cell was 15d. To study the conditions for immobilized cell of Cephalotaxus mannii and its effect on the growth and harringtonine synthesis of C.mannii. The growth cycle of the immo bilized cell of C. mannii was 40th day, the lag period of growth was 0-15th day, the slow growing time in 15-30th days, and the stable phase was 30-40th day. Immobilized cell of Cephalotaxus mannii in the lag phase had less consumption. The activity of immobilized cell was low. The activity was increased gradually with the development of immobilized cell. In the 0-15th day immobilized cell of C. mannii culture, the harringtonine increased, and the cells reach its optimal production volumes in the 15th day. In the 0-15th day immobilized cell of C. mannii culture, the harringtonine decreased.2. To study the optimization of the immobilized conditions of cell of C. Mannii. The research result show that the hardness of ball increases with the concentration of sodium alginate increases. The cells reach its optimal production volumes in the 20g/L of sodium alginate. Concentration of sodium alginate had effect on activity of immobilized cell of C. Mannii, higher concentration showed a inhibition which was unfavourable to synthesis of harringtonine. Solution of CaCl2 for concentration of 0.023g/mL, the immobilization cells of C. Mannii obtain maximum harringtonine yield. The production of harringtonine firstly increased and then decreased with the increased amount of vaccination of immobilized cell of C. Mannii. In the culture of 15th day, the production of harringtonine maximum, when inoculation quantity was O.lg/ml. Immobilized cells of C. Mannii synthesis of harringtonine maximum in the embedding density of 0.2g/ml. The activity of immobilized cell of Cephalotaxus mannii increased and medium browning decreased and increased yields of harringtonine with increasing shaker speed. In optimization conditions, the immobilized-cell could be reused 3 times, changed 70ml medium everytime, the production efficiency for harringtonine was 0.032mg/L d-1, the production efficiency for homobarringtonine was 0.014mg/L d-1.3.To study the growth and harringtonine synthesis of the immobilized cell of Cephalotaxus mannii in different age. Cells of Cephalotaxus mannii in different age were cultured at the optimized conditions of the immobilized cell culture, the biomass、activity of cell、activity of phenylalanine ammoiuum-lyse(PAL)、sucrose content、medium browning and production of harringtonine were determined. The research result shows that the younger age of cell was, the immobilized cell of Cephalotaxus mannii grew more rapidly and active, but the production of harringtonine was lower. On the contrary, the production of harringtonine and total phenols in medium increased and the colour of medium became deeper on the condition of the old cell being immobilized.In this case, the growth speed of cells and the consumption of surplus sucrose were in the lower level. Therefore immobilized cell of Cephalotaxus mannii in the age of 15d maximize the production of harringtonine in a short period.