Critical Technology For Efficient Production Of High-quality Bacterial Nanocellulose In Large Scale

Bacterial nanocellulose?BNC?is a high value-added biomaterial mainly synthesized by acetic acid bacteria.It possesses unique structure and properties and is endowed with great potential in a variety of areas including textile manufacturing.However,many potential applications of BNC are restricted by its relatively high price.The high price is mainly due to the high cost of culture medium including carbon source,small scale production,low production rate and high labour intensity of the traditional static production technology.Additionally the difficulty in property regulation restricts its application as a material.In this study,the following three aspects were performed to efficiently produce high quality BNC in large scale:a new approach was developed to produce BNC by biorefining the lignocellulosic waste of sweet sorghum bagasse?SSB?,the dependence of BNC productivity and properties on critical bioprocess conditions was investigated,and pilot scale-up of production of flocculent and membranous BNC was also studied to promote the BNC production.1.Process of BNC production from the agricultural waste sweet sorghum bagasse?SSB?after ionic liquid pretreatment was developed,and the related mechanisms were studied.?1?Six kinds of ionic liquids?ILs?were firstly compared in detail and showed a diversity of inhibition on cellulase and bacterial culture along with increasing IL concentration.The results showed that IL of[EMIM]Fmt had higher dissolving capacity of cellulose and large enhancement in cellulase saccharification of SSB.The fermentable sugar yield from SSB was enhanced by more than 7.5 times.The highest productivity of BNC was obtained after cultivation with the hydrolysate of the[EMIM]Fmt-pretreated SSB.?2?Dilute acid pretreatment and[EMIM]Fmt-pretreatment of SSB were compared in terms of their effects on SSB component and structure and saccharification enhancement.It was shown that after[EMIM]Fmt-pretreatment,the component of SSB rarely changed.However,the morphology of SSB changed to be floc-like,and its crystallinity decreased.Moreover,all of the surface area,porosity,and pore size largely of SSB increased.Compared to dilute acid pretreatment,[EMIM]Fmt-pretreatmentperformedsignificantlybetterin saccharification enhancement of SSB.?3?The effect of two kinds of SSB hydrolysate on the BNC productivity and properties were compared.Both of the BNC productivity and yield from the SSB hydrolysate using[EMIM]Fmt-pretreatment were significantly higher than that from dilute acid pretreatment.By comparing the structure and properties of BNC,it was shown that BNC from the SSB hydrolysate using[EMIM]Fmt-pretreatment featured lower average viscometric degree of polymerization?DP_v?,lower crystallinity,lower thermal stability and lower water content.However,the Young's modulus of BNC from the SSB hydrolysate using[EMIM]Fmt-pretreatment was 1.3 times of that using dilute acid pretreatment.Through the above studies,an effective process for preparing BNC with low-cost raw material SSB was developed,and a new raw material preparation route was established for large-scale production in the future.2.The effects of key culture factors on BNC productivity and properties were investigated.?1?Komagataeibacter xylinus was cultivated in eight culture media based on different carbon sources,viz.seven biomass-derived sugars and one sugar mixture,and the productivity and quality of the BNC produced in the different media were compared.Highest volumetric productivity,yield on consumed sugar,DP_v and thermal stability were achieved using media based on glucose or maltose.Growth in media based on xylose,mannose,or galactose resulted in larger fibril diameter and higher crystallinity.Growth in medium based on a synthetic sugar mixture promoted BNC productivity and yield,but decreased fibril diameter,DP_v,crystallinity,and thermal stability.?2?The dependence on strain and culture conditions of the productivity and quality of BNC was investigated for four strains in static and agitated cultures with an initial pH in the range 4.0-6.0.Strain DHU-ATCC-1 displayed highest productivity[1.14 g/?L×d?]in shake flasks,and strain DHU-ZGD-1186 in static cultures[0.55 g/?L×d?].Static cultures of DHU-ZGD-1186 exhibited superior BNC yield on consumed glucose?0.79 g/g?.The BNC from DHU-ZGD-1186exhibited higher DP_v?7800?,higher crystallinity index?84%?and cellulose I_?content,and higher tear index.Productivity,DP_v,crystallinity,and mechanical strength of BNC from shake flasks were typically similar to or even higher than the corresponding values of BNC from static cultures.?3?Seven agitator configurations in stirred-tank reactor?STR?were investigated to optimize the production of BNC in glucose-based media.At optimized stirring speed,the twin pitched blade?big?performed best.The K_La and shear force were two critical factors that determined the performance of an agitator configuration in BNC production.Further characterization indicated that the obtained BNC from the STR equipped with the optimal agitator at100 rpm maintained high degree of polymerization and crystallinity.?4?The effect of cultivation condition in STR on BNC productivity and DP_v was investigated.In a400-mL STR,highest BNC productivity and yield was achieved at 250 rpm,while highest DP_v was obtained at 150 rpm.Increasing aeration rate would decrease the DP_v significantly.The pH adjustment significantly enhanced the BNC productivity,but meanwhile it decreased the DP_v significantly.Through the above studies,comprehensive theory for regulating the yield and properties of BNC in the fermentation process has been developed.3.The up-scaling of submerged production of flocculent and membranous BNC was explored,and the BNC produced were characterized.?1?Scale-up of flocculent BNC production using a 75-L pilot STR was investigated to assess the possibility to improve BNC production and maintain its unique properties.Scale-up using a 75-L STR pilot bioreactor resulted in an enhancement of the BNC production rate from0.056 g/d in the shake flasks to 17.3 g/d in the 75-L STR,although the productivity decreased by 23%.During scale-up from shake flasks to 400-mL STR and further on to 75-L STR,the BNC fibers formed more bundles,and fiber diameter decreased,and The BNC from the 75-L STR exhibited higher degree of polymerization,higher degree of crystallinity,larger crystallites,and improved strength including higher tensile energy absorption index and superior stretch at break.?2?Scale-up of production of membranous BNC was investigated by using a newly designed 30-L rotary drum bioreactor.It is showed that compared with the traditional tray cultivation,the productivity of dry and wet BNC membrane from the rotary drum bioreactor cultivation were enhanced by 1.65 times and 3.73 times,respectively.The BNC hydrogel membrane from the rotary drum bioreactor cultivation showed anisotropic morphology and tensile properties,sparser network and larger fiber diameter.The degree of polymerization of BNC hydrogel membrane from the rotary drum bioreactor was lower,but was still 1.8 times higher than cotton.BNC hydrogel membrane from the rotary drum bioreactor also featured with significantly higher water content of 99.84 w/w%,much higher ultraviolet-visible light transmittance.BNC aerogel membrane was prepared via lyophilization and analyzed.Compared with that from the traditional tray static cultivation,the BNC aerogel from the rotary drum bioreactor had a much lower density of 1.49 mg/cm³.Its structure featured lower crystallinity index,and exclusively higher specific surface area?234 m²/g?.With the lower density and special structure,the thermal conductivity of the corresponding BNC aerogel film was determined to be only 72%of the thermal conductivity of the air.Through the above studies,the foundation for the large-scale production and application of BNC has been laid.In summary,the innovations in this paper include:?1?The critical technologies related to the efficient preparation of high-quality BNC in large-scale are thoroughly studied,and the idea on the improvement of BNC yield without the expense of quality of BNC is first proposed in the BNC production studies.?2?A new approach was developed to produce BNC with low cost by using SSB and[EMIM]Fmt-pretreatment.?3?The effects of key factors in the fermentation process on the yield and properties of BNC were clarified,which provided as theory for improving the BNC productivity and implementing the controllable production of BNC with regard to its structure and properties.?4?The pilot scale submerged production was for the first time carried out for both flocculent and membranous BNC,which provides important technical support for the further industrial production and application of BNC.