Biosynthesis Of Cytidine Triphosphate(CTP) By Immobilized Yeast Cells

Cytidine monophosphate(CMP) is converted to Cytidine triphosphate(CTP) by dissociative beer yeasts and compound enzymes from yeasts that has been studied in this paper. And based on these studies, biosythesis of CTP by immoblizied yeast cells would be carried out, meanwhile, effects on several kinds of carriers, and the concentration of CMP , initial glucose, phosphate buffer and pH, temperature, reaction time are also studied. Then hope to feel about a optimal production process of CTP.We can find that the results about the best technological parameters on the synthesis of CTP are comparatively accordant to dissociative beer yeasts and compound enzymes from yeasts by orthogonal optimal method, included 60.0 mmol/L CMP, 150.0 mmol/L initial glucose, 250.0 mmol/L phosphate buffer(sodium), 8.0 mmol/L magnesium chloride, 3.0 mmol/L ATP, the temperature is 35℃. There are some differents in pH and reaction time. The optimal pH 6.0 and reaction time 2.0 h is used in dissociative beer yeasts, but they are pH 8.0 and reaction time 2.5 h in compoundenzymes from yeasts, meanwhile, the conversion rate of the former is about 80%, less than the latter, about 90%.Through a choice about immobilizing carriers and the compare about several kinds of entrpping matrix, it is shown that k-carrageenan is better than other carriers. By orthogonal experiments, the results show that the carrier included in 2.7% k-carrageenan, 25% concentration of cells , and added to 0.3% potassium ion, put in 2.0% potassium chloride about 4.0 h , has good intensity of mechanism and the conversion rate.Cell membrane permeability about the immoblized waste beer yeasts and immobilized new beer yeasts is also studied. The results show that the activity of cells is greatly improved after freezed above three days , three times to be freezed and thawwed, and added to appropriate surfactant.The effects of conversion caused by some conditions during synthetical process are researched by HPLC and orthogonal optimization tests of the mostly factors are also carried out. Under the optimal conditions, about 45.7% CMP was transformed to CTP.Based on batch reactions, the system of continuous column reaction is founded. When the diluted rate is 1.89 X 10-41/s, there is a optimal coversion rate. And the best yield is 57.6%. The immobilized cells can be operated four or five batches.