The Study Of Growth And Metabolism Property Of Microbe In Chitosan/Alginate Microcapsules

At present, the study of microencapsulated culture of mammalian cell and microbe has been the hot spot in the fields of biology and medicine science. But in the process of microcapsule preparation and microencapsulated cell culture, the high cost of membrane materials, the complicated preparation procedure, the lower membrane strength and mass diffusion restriction restrain its extensive application.This paper was aimed at microencapsulated cell culture. In order to sovle theabove problems, the preparation, membrane properties of microcapsules andmicroencapsulated culture property were studied under the base of the mild andefficient impulsive electrostatic droplet generation technique. Owing to their goodbiologic characteristics and economical property, chitosan and sodium alginate wereused as microcapsule materials. Experimental results were showed as follows:1. The influence of preparation conditions on chitosan/alginate microcapsules'morphology and membrane properties had been systematically investigated. Themicrocapsules prepared with 2.0% sodium alginate solution had fine sphere, evenparticle size and smooth surface. With the increase of the concentration ofchitosan solution, membrane formation time and the decrease of the particle sizeof calcium alginate gel beads, the microcapsule membrane stability was increased,while its permeability was decreased. But considering maintaining of cell activity,15 minutes was suitable for membrane formation time. As for the chitosanmolecular weight, the microcapsule membrane properties were the combinedeffect of the chitosan molecular weight and membrane formation time. Whenmembrane formation time was less than 15 minutes, with the increase of thechitosan molecular weight, the microcapsule membrane stability was increased,while its permeability was decreased. 2. Glucose and citric acid are the necessary carbon source for microbial cell culture, and L-glutamic acid is the nitrogen source. In this work, using these subjects as the low molecule models, and a series of poly(ethylene glycol)s (PEG) of different molecular weight as the high molecule models to study the molecular mass cut-off(MMCO), we investigated the diffusivity of chitosan/alginate microcapsule. Glucose, citric acid and L-glutamic acid could diffuse through the membrane freely, and the gel structure in the microcapsules was depolymerizated by competitively chelating Ca²⁺ with the ingress of citric acid and L-glutamic acid. The molecular mass cut-off(MMCO) of microcapsules prepared with 2.0% sodium alginate and fifty thousand molecular weight,0.2% chitosan was 1500.3. Using E.coIi, yeast and B.licheniformis as the cell models, the method of microencapsulated microbial cell had been established. Meanwhile the growth and metabolism property of microencapsulated cell culture had been studied. Cell growth and metabolism had a shorter lag in microcapsules compared with free culture because of the mass diffusion effect of the microcapsule membrane, but the microcapsules provided a steady micro-environment for cells and the microencapsulated culture existed advantage over free culture when cell culture entered the steady stage. We obtained higher cell biomass and the microcapsule morphology maintained fine through batch culture, so there was enough membrane stability for chitosan/alginate microcapsule. The phenomenon of microbial cell aggregation inside chitosan/alginate microcapsule and the "S" variety rule of microcapsule particle size with the incubation time had been discovered.4. The influence of preparation conditions on culture property of the cell-loaded chitosan/alginate microcapsules had been systematically investigated. There was an optimum initial inoculation density for microencapsulated microbial cell culture. Although the influence of the chitosan molecular weight and concentration on microencapsulated cell culture existed in the beginning stage of initial culture, it showed no difference in steady stage and batch culture. The influence of microcapsule particle size on microencapsulated culture property was remarkable. Reducing microcapsule volume was beneficial to oxygen and mass diffusion through microcapsule and weakened the inhibition effects of metabolites, which was favorable for microencapsulated cell culture.In conclusion, the chitosan/alginate microcapsule is an ideal microreactor for cellculture. It provides a new method for the cell-loaded microcapsule technique in thefields of bio-chemistry engineering and medicine science.