Biotechnol. Bioprocess Eng. 1999, 4, 41-45 Effects of Mixing Conditions on the Production of Microbial Cellulose by Acetobacter xylinum Hei Chart Lee* and Xia Zhao Division of Chemical Engineering, College of Engineering, Sun Moon University, Asan, Chungnam 336-840, Korea Microbial cellulose has many potential applications due to its excellent physical properties. The production of cellulose from Acetobacter xylinurn in submerged culture is, however, beset with numerous problems. The mest difficult one has been the appearance of negative mutants under shaking culture conditions, which is deficient of cellulose producing ability. Thus genetic instability of Acetobacter xylinurn under shaking culture condition made developing a stable mutant major research interest in recent years. To find a proper type of bioreactor for the production of microbial cellulose, several production systems were developed. Using a reactor system with planar type impeller with bottom sparging system, it was possible to produce 5 g/L microbial cellulose without generating cellulose minus mutants, which is comparable to that of static culture system. Key words: Acetobacter xylinurn, microbial cellulose, bioreactor design, negative mutants, shaking culture INTRODUCTION Acetobacter xyiInum, a gram-negative aerobic bacte- ria, generally under static culture conditions secretes cellulose fibrils as part of its normal metabolic activity. Under electron microscope microbial cellulose (MC, or bacterial cellulose) characteristically appears as a form of separate ribbon-like fibrils in contrast to the cellulose of high plants consisting of bundles of micro- fibrils [1]. It possesses not only excellent physical properties, such as high degree of polymerization and preferential orientation, but also strong mechanical and absorbent properties. Moreover, the fibrils of microbial cellulose are composed of pure cellulose, which is devoid of lignin, hemicellulose, and other substances, thus it can be purified more easily than natural cellulose. Using those characteristics of micro- bial cellulose, therefore many potential commercial applications are being developed. At present, microbial cellulose has found practical applications such as sensitive diaphragms for stereo headphones, additives for food and composite paper and textile products, thickener for paint, composite membrane, binder and dietary fiber supplement in food processing and also as a temporary skin substitute in skin burn treatment [2-8]. Although Acetobacter xylinurn has proved to be the greatest potential for the commercialization in indu- strial applications, reported values of cellulose produc- tivity are too low for large scale production [9-15]. For this reason the subject of how to improve cellulose productivity of Acetobacter xyIinum has already ab- sorbed interests of many researchers. Investigations have been made on isolating high cellulose-producing strain [16], mechanism of cellulose biosynthesis and genetic structure involving cellulose secretion [3,17-24]. * Corresponding author Tel: +82-418-530-2376 Fax: +82-418-541-7426 e-mail: heichan@emega'sunm~176 However, relatively few reports have discussed in details the relationship between cellulose production and culture conditions [11,14], a few were about the influence of nutritional sources on cellulose produc- tivity after the first report of Schramm and Hestrin [14]. In the literatures concerning the production of microbial cellulose by A. xytinurn [3,9-15], peptone, yeast extract and (NH4)2SQ were used as nitrogen source, while glucose, mannitol, sucrose, fructose, citrate, or ethanol as carbon source, KH2PO4 and Na2HPO4 as phosphate source, and MgSO4 or FeC13 as mineral elements. These medium compositions are the simple variations of Schramm and Hestrins medium or made by adding single component such as citrate, ethanol, and so on. Therefore, for these newly developed applications to be economically feasible, a commercial scale fermentation process for large-scale production of microbial cellulose needs to be devel- oped. By static culture of Acetobacterxylinurn, however, about 2,000 square feet of air-liquid interface area is required for the production of one pound cellulose per day, therefore, static culture system is considered to be inefficient from the industrial point of view [3]. In general, relatively smaller amount of cellulose is produced in shaking and agitated culture than in static culture and this is closely related to the genera- tion of negative mutants which does not produce microbial cellulose. These mutants are known to occur with agitation in the culture. Insertional sequences were found in the genes of negative mutants [25,26]. In addition to the reports on developing culture medium and isolating bacterial strains which is stable under shaking and agitated culture conditions [27-31], some reports have been made on the environmental conditions causing mutant generation [32,33]. However, none was on the bioreactor configuration which can produce microbial cellulose without generating mutants deficient of microbial cellulose producing ability. Developing a submerged fermentation system for the production of microbial cellulose is the subject of this research.
