在再生能源的眾多選項中，生質乙醇被視為一種極佳的石化燃料替代品。相較於傳統批次乙醇醱酵，本研究進行連續式乙醇醱酵，並結合固定化細胞技術，藉以促進醱酵生產速率之提升，以達成具有經濟發展之重要效益指標。本研究利用聚乙烯醇(PVA)包覆Zymomonas mobilis製成固定化細胞，應用於連續式乙醇醱酵程序中，並透過固定化細胞添加比例與水力停留時間(HRT)等操作因子之最適化實驗設計，藉以達到高乙醇生產速率與低葡萄糖殘留之操作系統。
高濃度葡萄糖醱酵系統中，葡萄糖濃度對於高濃度乙醇生產程序是非常重要的操作指標。本研究利用Fe2O3與聚乙烯醇進行固定化細胞基材修飾進行連續式乙醇醱酵，並針對不同水力停留時間(1到4小時)，不同葡萄糖進流濃度(100、125、150 g/L)與不同固定化細胞濃度(20、40、50%)進行連續式醱酵參數試驗。結果可知，在聚乙烯固定化材料中添加1% Fe2O3、葡萄糖進流濃度為125 g/L、固定化細胞添加量為40%(w/v)、且水力停留時間為2小時的操作條件下，可有效將乙醇生產效率從原本的15.74 g/L/h 提升至 31.09 g/L/h，並且此固定化生產系統可以被穩定操作達20天以上，其生產效率與細胞活性都無明顯降低。
本研究接著以薄膜蒸餾移除系統(VMD)進行同步移除乙醇之系統整合，藉以解決生產系統中產物抑制的問題，而本研究成功的將薄膜蒸餾移除系統(VMD)與連續式乙醇生產程序做一個完美的操作整合，並在300 g/L葡萄糖進流濃度條件下，其最大乙醇生產濃度為127.39 g/L、最大乙醇生產速率為63.69 g/L/h，且葡萄糖轉化效率高達84.93%。
此外，在醱酵生產乙醇的程序中，也伴隨著二氧化碳與有機酸等副產物的生成，而二氧化碳在此乙醇生產系統排放氣體中的含量接近100%。由於微藻具有快速生長速率、優異的二氧化碳固定效率與高含量碳水化合物/油脂累積能力，本研究透過微藻系統將二氧化碳進行固定吸收，藉由高碳水化合物之醣藻作為料源用以生產生質乙醇，進而解決二氧化碳這種低價值副產物對環境所造成的衝擊。由於在二氧化碳固定程序中，植物與微藻都必須以陽光做為光合作用之能量，因此，在系統放大生產時受到相當大的限制。有鑑於此，許多微生物在生長時往往需要消耗二氧化碳作為代謝所需，而琥珀酸便是其中一項相關的代謝產物。在琥珀酸醱酵程序中，1 mol二氧化碳經由PEP carboxykinase代謝合成路徑，能夠產生1 mol琥珀酸。由實驗結果可知，琥珀酸醱酵程序所消耗的二氧化碳速率為31.92 g/L/d是微藻系統的15倍，因此以微生物移除二氧化碳並代謝生產琥珀酸是非常具有效益之程序。
由於全世界的纖維料源非常豐富(如:蔗渣、稻稈等)，且此類物質之使用可避免與民爭糧的問題。研究最後，利用非糧食作物的纖維料源進行生質乙醇的生產。由研究結果可知，在120 g/L鹼處理蔗渣水解下，可以得到62.79 g/L的葡萄糖，透過連續式固定化細胞之乙醇醱酵系統，其葡萄糖轉化率為94.57%，最大乙醇濃度為26.82 g/L,且最大乙醇產率13.41 g/L/h，此結果也證明利用纖維水解物亦能夠成功的應用於所建立的連續式固定化細胞乙醇生產系統。

Bioethanol as one of renewable energy is considered an excellent alternative clean-burning fuel to replace gasoline. Continuous bioethanol fermentation systems have offered important economic advantages in comparison with traditional systems. Fermentation rates can be significantly improved when the continuous fermentation is integrated with cell immobilization techniques to enrich the cells concentration in the fermenter. Growing cells of Zymomonas mobilis immobilized in polyvinyl alcohol (PVA) gel beads were employed in an immobilized-cell fermenter for continuous bioethanol fermentation from glucose. The glucose loading, dilution rate and cells loading were varied in order to determine the best condition employed in obtaining both high bioethanol production and low residual glucose at high dilution rates.
Higher glucose fermentation rate has been considered as the most important target as it leads to higher bioethanol titer and better bioethanol productivity. To enable bioethanol fermentation at high glucose concentration with continuous-flow operations, the Z. mobilis cells were immobilized using polyvinyl alcohol (PVA) matrix modified with a certain amount of iron (III) oxide (Fe2O3) and the cell immobilization of Z. mobilis was simply performed by using the enriched cells culture media harvested at the exponential growth phase. The production of bioethanol was affected by the medium flow rates or hydraulic retention time (HRT) from 1 to 4 hour. In addition, the effects of both initial glucose loadings (100 g/L, 125 g/L and 150 g/L) and cell loadings (20% (w/v), 40% (w/v) and 50% (w/v)) were also investigated. On the other hand, the bioethanol production performance of Z. mobilis cells immobilized with cross-linked polymer of PVA with 1% (w/v), 2% (w/v) and 3% (w/v) of Fe2O3 and unmodified PVA-immobilized Z. mobilis was compared. Reusability of the immobilized Z. mobilis or stability of beads was examined and found that the immobilized cells could be utilized for more than 20 days without losing their activity. Biothanol productivity increased from 15.74 g/L/h to 31.09 g/L/h when the modified PVA-immobilized Z. mobilis with 1% (w/w) of Fe2O3 was employed and the glucose loading was 125 g/L. Moreover, the HRT of 2 hour and cell loading of 40% (w/v) were considered as the optimum conditions to obtain the best bioethanol fermentation performance.
The modified PVA-immobilized Zymomonas mobilis cells were used to enhance the efficiency of bioethanol production under very high gravity (VHG) conditions. Continuous bioethanol fermentation was integrated with in-situ bioethanol removal via vacuum membrane distillation (VMD) to overcome the problems associated with product inhibition and the resulting low bioethanol productivity during bioethanol fermentation. The developed VMD-integrated VHG fermentation system can be successfully operated under a feeding glucose concentration of up to 300 g/L (or 30% (w/v)), obtaining a maximum bioethanol concentration of 127.39 g/L (or 16.14% (v/v)), an bioethanol productivity of 63.69 g/L/h, and a glucose conversion of 84.93%.
In addition, the production of bioethanol by fermentation must be accompanied by some by-products, such as carbon dioxide (CO2) and organic acids. These by-products can lower the quality and usability of bioethanol as a biofuel and also increase the amount of wastes to cause environmental pollution. CO2 produced from fermentation is of high purity and is nearly a saturated gas (almost 100%). The majority of CO2 applications were dedicated to serving carbonated beverage and food processing/preservation. Beyond these traditional applications, one of the most potential ones is the production of algae-based biofuels through CO2 fixation by microalgae. The advantages of using microalgae CO2 fixation include rapid growth rate and high CO2 fixation capability when compared to conventional plants and high oil/carbohydrate production. The carbohydrate-rich microalgal biomass then can be used for bioethanol production in large scale applications.
Photosynthesis is often required for CO2 fixation. The need of solar energy supply in photosynthesis reactions appears to limit the application of biological CO2 sequestration due to scale-up problems. Furthermore, many microorganisms considered for CO2 fixation have fastidious growth requirements. Succinic acid is a common natural organic acid often found in humans, animals, plants and microorganisms. As 1 mol CO2 is theoretically required for the synthesis of 1 mol succinic acid. CO2 should play an important role in succinic acid production to promote the regulation of the PEP carboxykinase pathway. Compared to carbon capture by microalgae, this developed system encourages much higher CO2 fixation rate. The highest carbon fixation rate achieved for microalgae cultivation was 2.06 g/L/d, while it was 31.92 g/L/d (15 times higher) for succinic acid production. Therefore, production of succinic acid is also a feasible way of biological CO2 removal.
In the end, bioethanol has to be produced from non-food sources. Cellulosic biomass was used for bioethanol fermentation for the purpose of reducing greenhouse gas emissions and giving impacts to rural economy condition. Cellulosic resources are in general very widespread and abundant. Being abundant and outside the human food chain brings cellulosic materials such bagasse, rice straw, etc. relatively inexpensive feedstocks for bioethanol production. A 120 g/L of bagasse loading or corresponding to 62.79 g/L of glucose after hydrolysis was used to evaluate the feasibility of producing bioethanol with the modified-PVA immobilized Z. mobilis cells. The glucose conversion obtained was about 94.57% in average with a bioethanol titer and productivity of approximately 26.82 g/L and 13.41 g/L/h, respectively, during fermentation. The results demonstrate that the modified PVA-immobilized Z. mobilis cells are preferable cell immobilization system for cellulosic bioethanol production.

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