基於日益嚴重的全球暖化與氣候變遷問題，開發二氧化碳捕捉與封存的相關技術乃刻不容緩的議題。其中，利用微藻進行生物固碳與其他下游產品的應用在最近受到相當廣泛的重視。本研究從南台灣淡水域中篩得的22株本土斜邊柵藻 (Scenedesmus obliquus)中選出七株進行詳細的親緣鑑定與二氧化碳固定能力測試。在這七株本土柵藻中，S. obliquus CNW-N展現了較佳的生長速率與二氧化碳固定能力，因此被選為進行後續深入研究的目標藻株。本研究首先利用反應曲面法 (Response surface methodology; RSM) 進行批次培養條件的優化，選用跟生長與二氧化碳固定效率相關的四個重要因子(例如：二氧化碳濃度、二氧化碳流速、鎂離子濃度、與光照強度)進行混成實驗，並利用比生長速率與二氧化碳固定效率的表現做為評估的依據，尋找四個影響因子中交疊的部分。在最適化範圍中 (2.5%二氧化碳濃度搭配流速0.4 vvm、鎂離子濃度1.7-2.7 mM、光強度180-250 μmol m-2 s-1)，比生長速率與二氧化碳固定效率可分別大於1.22 d-1與800 mg L-1d-1。再者，本研究利用半批次與連續式操作等工程策略進一步提升其生長與二氧化碳固定效率。在最佳連續流操作條件下，其生質體產率與二氧化碳固定效率可分別高達1139.5 mg L-1d-1 與1988.6 mg L-1d-1，此成果表現優於大多數目前發表在國際期刊之數據。
此外，本研究亦致力於利用缺氮與其他工程策略提升微藻S. obliquus CNW-N的二氧化碳固定能力與含糖量。在前面最佳批次條件下，首先乃利用氮源缺乏策略來使其藻體內的糖量累積，在兩天的缺氮下其細胞內總糖可以從20.9%迅速提升至49.4%。 為了進一步增加其未來進行酒精生產的可行性，本研究進一步利用不同的工程操作策略 (例如：二階段培養、半批次操作、與連續式操作) 來提升其二氧化碳固定效率與胞內含糖量。在最佳的半批次操作下，其生質體生產速率、總糖累積速率、與二氧化碳固定效率可分別達883.8 mg L-1 d-1、467.6 mg L-1 d-1 (49-52%之胞內含糖量)、與1546.7 mg L-1 d-1。此數據表現亦明顯優於目前大多數文獻發表之成果。再者，在不同工程操作結合缺氮策略下，其胞內總糖大多皆為葡萄糖(佔約80-82%)，十分適合進一步進行生質酒精發酵或是其他相關產品之生產。
除此之外，本研究亦著眼於評估利用富含糖量的S. obliquus CNW-N藻體做為料源來進行後段之酒精生產，測試不同的酸水解條件並結合後段的兩階段水解醣化醱酵程序(separate hydrolysis and fermentation; SHF)來進行酒精生產。研究中發現2%之稀硫酸能夠非常有效率的水解S. obliquus CNW-N的乾、濕藻體，在起始濃度為40 g L-1(濕藻體)與60 g L-1(乾藻體)的條件中達到約96-98%的葡萄糖產率。為了增加未來商業化的可行性，本研究決定選擇未經過去水步驟的濕藻體做為後段之酒精發酵料源。再經過2%硫酸水解前處理與兩階段水解醣化醱酵程序，酒精濃度可達8.55 g L-1 (起始葡萄糖濃度約為16.8 g L-1)，達到幾乎99.8%的理論產率。因此，此富含糖類的微藻S. obliquus CNW-N極適合做為後段酒精生產之料源。
最後，本研究將室內小型的管狀反應器放大至50升並進行微藻S. obliquus CNW-N之戶外培養測試。在夏季陽光充足的環境下，最大的二氧化碳固定效率、總糖含量、與總糖生產效率可分別達430.2 mg L-1 d-1、45.5%、和111.8 mg L-1 d-1。此外，為了進一步提升其表現，本研究亦在夜間(晚上六點至早上六點)進行人工補光的實驗測試，發現在有夜間補光(500 μmol m-2 s-1)的環境下，其最大的生質體生產速率、二氧化碳固定速率、與總糖累積速率可分別提升至390.1 mg L-1d-1、682.7 mg L-1d-1、和185.7 mg L-1d-1，亦明顯優於大多數相關之文獻值。另外，利用戶外培養的S. obliquus CNW-N在缺氮條件下培養可迅速累積葡萄糖(可佔總糖中的82%)，表示其適合用來進行後續的酒精發酵。因此，將S. obliquus CNW-N於夜間補光的戶外光反應器系統放大培養，可同時進行微藻減碳與生物燃料(酒精)之生產。

Due to the problems of climate change and global warming, technology on CO2 capture and storage is urgently needed. In particular, biofixation of CO2 by microalgae and the downstream utilization of resulting microalgae biomass have recently become a hot research topic. In this study, seven out of twenty-two indigenous Scenedesmus obliquus isolates obtained from southern Taiwan were selected based on their CO2 fixation ability. Among them, S. obliquus CNW-N displayed the highest cell growth rate and highest CO2 removal ability, thereby being selected for detailed CO2 fixation studies. Response surface methodology was used to optimize the four critical parameters (e.g., CO2 concentration, CO2 flow rate, magnesium concentration, and light intensity), which significantly influence the CO2 fixation performance. Two performance indexes (e.g., specific growth rate and CO2 fixation rate) were used to assess the microalgal-CO2 fixation ability. An overlay counter plot was used to determine the optimal region of four vital parameters by simultaneously considering all the performance indexes. The optimal region (i.e., 2.5% CO2 , 0.4 vvm, magnesium concentration, 1.7-2.7 mM; light intensity, 180-250 μmol m-2 s-1) was obtained based on the criteria of specific growth rate >1.22 d-1 and CO2 fixation rate > 800 mg L-1d-1. Moreover, using engineering strategies of semi-continuous and continuous cultivation further improves the biomass productivity and CO2 fixation rate to 1139.5 mg L-1 d-1 and 1988.6 mg L-1 d-1, respectively. This performance is better than the results from most of the related studies.
Moreover, the nitrogen starvation strategy and engineering operation strategies were applied to achieve higher CO2 fixation rate and carbohydrate productivity of Scenedesmus obliquus CNW-N. The microalga was first cultivated with 2.5% CO2 feeding and a light intensity of 210-230 μmol m-2 s-1 under nitrogen rich medium for cell growth and CO2 fixation, followed by applying nitrogen starvation strategy to trigger carbohydrate accumulation. After two days of nitrogen starvation, the carbohydrate content of microalga S. obliquus CNW-N increased dramatically from 20.9 to 49.4%. Engineering strategies (e.g., two-stage, semi-batch, and continuous operations) were also utilized to further enhance the carbohydrate production performance of S. obliquus CNW-N so that the microalgal strain is more feasible for the use as feedstock for microalgae-based bioethanol production,. Under the optimal operation condition (i.e., semi-continuous operation), the highest biomass productivity (883.8 mg L-1 d-1), carbohydrate productivity (467.6 mg L-1 d-1 with 49-52% carbohydrate content per dry weight of biomass), and CO2 fixation rate (1546.7 mg L-1 d-1) were reached. This performance is also better than that obtained from most related studies. Under the nitrogen starvation condition and bioreactor operation strategies, the carbohydrate accumulated in the microalgal biomass was mainly composed of glucose, accounting for 80-82% of total carbohydrates. This carbohydrate composition is suitable for the production of bioethanol and any other fermentable products based on sugar platform.
The potential of using the carbohydrate-rich microalga S. obliquus CNW-N as feedstock for bioethanol production was evaluated via various acid hydrolysis conditions combined with separate hydrolysis and fermentation (SHF) process. Dilute acid hydrolysis with 2% sulfuric acid was shown to be effective and economical in saccharifying both wet and dried biomass of S. obliquus CNW-N, achieving a glucose yield of nearly 96-98% from the microalgal carbohydrates using starting biomass concentration of 40 g L-1 and 60 g L-1, respectively. To increase the feasibility of further commercialization, the wet biomass without dehydration step was used as the feedstock. Using the acidic hydrolysate of S. obliquus CNW-N biomass as substrate, the SHF process produced ethanol at a concentration of 8.55 g L-1 (initial glucose concentration=16.8 g L-1) and nearly 99.8% theoretical yield. These findings indicate the feasibility of using carbohydrate-rich microalgae as feedstock for fermentative bioethanol production.
To assess the commercial viability, the scale of the indoor vertical tubular-type photobioreactor (PBR) was increased to 50 liter PBR for outdoor cultivation of S. obliquus CNW-N. Under the phototrophic condition of sunny period during the summer season, the highest CO2 fixation rate, carbohydrate content, and carbohydrate productivity of 430.2 mg L-1d-1, 45.5%, and 111.8 mg L-1d-1, respectively, were obtained. The effect of external light compensation during night period (from 6 p.m. to 6 a.m.) on the performance of biomass and carbohydrate production was also investigated. The biomass productivity, CO2 fixation rate, and carbohydrate productivity of S. obliquus CNW-N were further improved to 390.1 mg L-1d-1, 682.7 mg L-1d-1, and 185.7 mg L-1d-1 when additional artificial light of 500 μmol m-2s-1 was used during the night period. This outdoor cultivaton performance is also better than most related reports. In particular, the microalgae-based sugars produced from large-scale cultivation system with external light compensation were mainly glucose (approximately 82% of the total sugars), suggesting that S. obliquus CNW-N is appropriate for the use as feedstock for ethanol fermentation with additional benefits of CO2 fixation.

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