Isolation Of A Cellulose-producing Strain, Fermentation And Application Of Bacterial Cellulose

Bacterial cellulose (BC), a kind of nano-biomaterial, is produced by some microorganisms. It is of particular network structure composed of ultra-fine ribbon-shaped fibers. In comparison with plant cellulose, BC displays high purity, high crystallinity, high water-holding capacity, high mechanical strength and controllable during the course of biosynthesis and so on. Therefore, BC can be applied in areas where plant cellulose can hardly be used. Nowadays BC has been one of the most active topics in the materials field. But domestic research on BC mostly rests on the level of lab, which lags behind far away from abroad. In this dissertation, the screening of suitable cellulose-producing strains, systematical research on fermentation technologies of successive enlargement and preliminary application in paper-making and immobilization of microorganism were reported.Twelve cellulose-producing strains were efficiently isolated from rotten fruits. Strain NUST1292 belonged to Acetobacter according to morphologic, biochemical and physiological characteristic and the 16S rDNA sequence analysis. Then it was mutagenized with UV. The mutant Acetobacter NUST4 had higher productivity of cellulose. The morphology, structural units, chemical and crystal structure of fermentation products were investigated using TEM, AFM, GC-MS, FT-IR, XRD and CP/MAS ¹³C-NMR.The optimum fermentation technologies of successive enlargement from static culture and shake flask culture, 13L stirred-tank reactor to 500L reactor were systematically researched. The main results were as follows: (1) Using optimum technology, BC production by Acetobacter NUST4 reached 9.87g·L^-1 in stationary culture for 7 days; (2) Acetobacter NUST4 was repeatedly domesticated in the stirred-tank reactor. Acetobacter NUST4.1, a high BC producer under the shake condition was efficiently isolated. Using optimum technology, BC production by Acetobacter NUST4.1 reached 7.16g·L^-1 in shake flask culture for 5 days. In the same conditions, BC yield reached 10.9g·L^-1 in stationary culture for 5 days. Influence of culture modes on BC structure and properties was characterized by SEM, FT-IR, XRD, purity and water-absorption capacity measurement, TG-DTA and mechanical tensile test; (3) In order to solve two problems on serious clump forming and spontaneous appearance of cellulose nonproducers in agitated culture, carboxymethyl cellulose sodium (CMC-Na), alginate sodium (NaAlg) and agar were added into the medium in shake flask and 13L reactor. It's surprising that the addition of these polymers contributed to forming uniform suspension and enhancing BC production. Influence of addition of three polymers on BC structure and properties in the shake flask was characterized by SEM, FT-IR, XRD, TG-DTG and mechanical tensile test. With the addition of 0.06% CMC-Na in the stirred-tank reactor, BC production reached the maximum. Then using optimum fermentation technology, the production of 3.35g·L^-1 was achieved, which was 3.1-fold higher than the control. Later, using two-step fermentation technology, BC production was 8.44g·L^-1; (4) Acetobacter NUST4.1 had stable cellulose-producing performance in enlargement test with 500L reactor.Application of BC produced in static (Y-SBC) and stirred-tank reactor (Y-ABC) by homogenate treatment adding to the different kinds of pulp in paper-making was investigated. The fiber structures of different papers were observed by SEM and paper properties were analyzed. The results showed that adding Y-ABC into waste paper pulp could enhance mechanical and water-resistant properties, and adding Y-SBC and Y-ABC into unbleached reed-wood pulp could also enhance mechanical and water-resistant properties, and paper properties adding ABC was better than that adding SBC.Application of BC produced by two-step fermentation method in immobilization carrier of white rot fungi Z-4 to decolorize and degrade five dyes with different structures was investigated. Free and immobilized cells could efficiently degrade triarylmethane dyes. Especially they could thoroughly degrade malachite green. Comparing with free cells, immobilization could enhance the ability to decolorize and degrade dyes, endure dye with higher concentration of malachite green, keep the stable rate of decolorize in the condition of different pH value, efficiently decolorize under the condition when immobilized-cells was added into the medium in company with dye but free cells needed culture for 3 days beforehand. The method can reduce the medium consumption and the possibility of contamination, showing high efficient and great economic benefit. It shows that BC is a fine immobilization carrier.