本研究的目標包含(1)研究柵藻於兩種不同刺激狀態下提升微藻體內之油脂含量的效率，第一種為缺乏氮源以及第二種為加入暗發酵產氫菌廢液刺激。(2)利用柵藻處理暗發酵產氫菌的廢液的可能性。暗發酵產氫菌生可產出合成氣體，是未來的能源發展前景，但暗發酵產氫菌生產合成氣體的同時，留下許多可再利用含有機酸廢液，這些有機酸有潛力可作為柵藻異營培養時的碳源來源。。
本研究以控制組、缺氮組、0.3x廢液組以及0.5x廢液組進行實驗比較，並利用柵藻於缺氮源刺激與暗發酵產氫菌廢液刺激之生長、色素含量、油含量以及培養液中有機酸含量來決定處理程序的效率。控制組之生物量產量為0.155g / L/day，其油脂含量達30.2wt％。缺氮組生產之生物量產量為0.155g / L /day，而其油脂含量為32.6wt％。0.3x廢液組生物量產量為0.177g / L /day，其油脂含量高達43.73wt ％。0.5x廢液組生物量產量為0.085g / L /天，其油脂含量為15.13wt ％。
由實驗結果獲知利用0.3x廢液刺激柵藻，其生物量生產力生與油脂含量最高。除此之外，0.3x廢液對暗發酵產氫菌廢液有機物之移除率較0.5x廢液組高，其中乳酸、甲酸、乙酸、丙酸和丁酸的移除率為84.01 %、100 %、100 %、84.01 % 和67.69 %。

This study aims to study the effects of two stress, nitrogen deprivation and organic acid addition, on the metabolism products of Scendesmus abundans. The organic acids were produced by the dark fermentation of hydrogen production bacteria. Dark fermentation is a future prospect for the synthesis of renewable fuel gas (syngas) but its effluent contains many organic acids which are generally considered as waste products.
Four different sets of experiments have been conducted: control (sufficient nitrogen), nitrogen depletion, 0.3 x effluent addition, and 0.5x effluent addition. Biomass productivity, pigment yields, lipid contents, and organic acid removal rates were compared to select the optimal stress condition. Biomass productivity produced by the control set was 0.155 g/L/day and lipid abundance was 30.2 wt%. Nitrogen deprivation produced a biomass productivity of 0.155 g/L/day and a lipid abundance of 32.6 wt%. The addition of 0.3x effluent produced a biomass productivity of 0.177 g/L/day and a lipid abundance of 43.73 wt% while 0.5x effluent produced a biomass productivity of 0.085 g/L/day and a lipid abundance of 15.13 wt%. These results show that 0.3x effluent of the hydrogen production fermentation had the highest biomass productivity and lipid abundance compare to the others. Besides, 0.3x effluent also has a better potential to decrease amounts of organic acids comparing to 0.5x effluent. 0.3x effluent removal efficiency of lactic acid = 84.01 %, formic acid = 100 %, acetic acid =100 %, propionic acid = 84.01 % and butyric acid = 67.69 %。

Abdel-Raouf, N., Al-Homaidan, A., & Ibraheem, I. (2012). Microalgae and wastewater treatment. Saudi journal of biological sciences, 19(3), 257-275.
Adams, C., Godfrey, V., Wahlen, B., Seefeldt, L., & Bugbee, B. (2013). Understanding precision nitrogen stress to optimize the growth and lipid content tradeoff in oleaginous green microalgae. Bioresource technology, 131, 188-194.
Arita, C. E. Q., Peebles, C., & Bradley, T. H. (2015). Scalability of combining microalgae-based biofuels with wastewater facilities: a review. Algal Research, 9, 160-169.
Axelsson, M., & Gentili, F. (2014). A single-step method for rapid extraction of total lipids from green microalgae. PloS one, 9(2), e89643.
Azwar, M., Hussain, M., & Abdul-Wahab, A. (2014). Development of biohydrogen production by photobiological, fermentation and electrochemical processes: A review. Renewable and Sustainable Energy Reviews, 31, 158-173.
Batista, A. P., Moura, P., Marques, P. A., Ortigueira, J., Alves, L., & Gouveia, L. (2014). Scenedesmus obliquus as feedstock for biohydrogen production by Enterobacter aerogenes and Clostridium butyricum. Fuel, 117, 537-543.
Bishop, W., & Zubeck, H. (2012). Evaluation of microalgae for use as nutraceuticals and nutritional supplements. J Nutr Food Sci, 2(5), 1-6.
Červený, J., Šetlík, I., Trtílek, M., & Nedbal, L. (2009). Photobioreactor for cultivation and real‐time, in‐situ measurement of O2 and CO2 exchange rates, growth dynamics, and of chlorophyll fluorescence emission of photoautotrophic microorganisms. Engineering in Life Sciences, 9(3), 247-253.
Chen, F. (1996). High cell density culture of microalgae in heterotrophic growth. Trends in biotechnology, 14(11), 421-426.
Chen, M., & Blankenship, R. E. (2011). Expanding the solar spectrum used by photosynthesis. Trends in plant science, 16(8), 427-431.
Chiu, S.-Y., Kao, C.-Y., Tsai, M.-T., Ong, S.-C., Chen, C.-H., & Lin, C.-S. (2009). Lipid accumulation and CO2 utilization of Nannochloropsis oculata in response to CO2 aeration. Bioresource technology, 100(2), 833-838.
Cohen, Z., Norman, H. A., & Heimer, Y. M. (1995). Microalgae as a Source of ω3 Fatty Acids1. In Plants in human nutrition (Vol. 77, pp. 1-31): Karger Publishers.
Durvasula, R., Hurwitz, I., Fieck, A., & Rao, D. S. (2015). Culture, growth, pigments and lipid content of Scenedesmus species, an extremophile microalga from Soda Dam, New Mexico in wastewater. Algal Research, 10, 128-133.
Gao, C., Zhai, Y., Ding, Y., & Wu, Q. (2010). Application of sweet sorghum for biodiesel production by heterotrophic microalga Chlorella protothecoides. Applied Energy, 87(3), 756-761.
George, B., Pancha, I., Desai, C., Chokshi, K., Paliwal, C., Ghosh, T., & Mishra, S. (2014). Effects of different media composition, light intensity and photoperiod on morphology and physiology of freshwater microalgae Ankistrodesmus falcatus–A potential strain for bio-fuel production. Bioresource technology, 171, 367-374.
Girbal, L., Croux, C., Vasconcelos, I., & Soucaille, P. (1995). Regulation of metabolic shifts in Clostridium acetobutylicum ATCC 824. FEMS microbiology reviews, 17(3), 287-297.
Gong, H., Tang, Y., Wang, J., Wen, X., Zhang, L., & Lu, C. (2008). Characterization of photosystem II in salt-stressed cyanobacterial Spirulina platensis cells. Biochimica et Biophysica Acta (BBA)-Bioenergetics, 1777(6), 488-495.
Griffiths, M. J., Garcin, C., van Hille, R. P., & Harrison, S. T. (2011). Interference by pigment in the estimation of microalgal biomass concentration by optical density. Journal of microbiological methods, 85(2), 119-123.
Guckert, J. B., & Cooksey, K. E. (1990). Triglyceride accumulation and fatty acid profile changes in Chlorella (Chlorophyta) during high Ph‐induced cell cycle Inhibition 1. Journal of Phycology, 26(1), 72-79.
Guedes, A. C., Amaro, H. M., & Malcata, F. X. (2011). Microalgae as sources of carotenoids. Marine drugs, 9(4), 625-644.
Ho, S.-H., Chen, C.-Y., & Chang, J.-S. (2012). Effect of light intensity and nitrogen starvation on CO2 fixation and lipid/carbohydrate production of an indigenous microalga Scenedesmus obliquus CNW-N. Bioresource technology, 113, 244-252.
Ho, S.-H., Chen, C.-Y., Lee, D.-J., & Chang, J.-S. (2011). Perspectives on microalgal CO2-emission mitigation systems—a review. Biotechnology advances, 29(2), 189-198.
Hu, B., Zhou, W., Min, M., Du, Z., Chen, P., Ma, X., . . . Ruan, R. (2013). Development of an effective acidogenically digested swine manure-based algal system for improved wastewater treatment and biofuel and feed production. Applied Energy, 107, 255-263.
Hu, Q., Sommerfeld, M., Jarvis, E., Ghirardi, M., Posewitz, M., Seibert, M., & Darzins, A. (2008). Microalgal triacylglycerols as feedstocks for biofuel production: perspectives and advances. The plant journal, 54(4), 621-639.
Huang, G., Chen, F., Wei, D., Zhang, X., & Chen, G. (2010). Biodiesel production by microalgal biotechnology. Applied Energy, 87(1), 38-46.
Khalil, Z. I., Asker, M. M., El-Sayed, S., & Kobbia, I. A. (2010). Effect of pH on growth and biochemical responses of Dunaliella bardawil and Chlorella ellipsoidea. World Journal of Microbiology and Biotechnology, 26(7), 1225-1231.
Kim, D.-H., Han, S.-K., Kim, S.-H., & Shin, H.-S. (2006). Effect of gas sparging on continuous fermentative hydrogen production. International Journal of Hydrogen Energy, 31(15), 2158-2169.
Kwak, H. S., Kim, J. Y. H., Woo, H. M., Jin, E., Min, B. K., & Sim, S. J. (2016). Synergistic effect of multiple stress conditions for improving microalgal lipid production. Algal Research, 19, 215-224.
Li, Y., Horsman, M., Wang, B., Wu, N., & Lan, C. Q. (2008). Effects of nitrogen sources on cell growth and lipid accumulation of green alga Neochloris oleoabundans. Applied microbiology and biotechnology, 81(4), 629-636.
Liu, C.-H., Chang, C.-Y., Liao, Q., Zhu, X., & Chang, J.-S. (2013). Photoheterotrophic growth of Chlorella vulgaris ESP6 on organic acids from dark hydrogen fermentation effluents. Bioresource technology, 145, 331-336.
Liu, C.-H., Chang, C.-Y., Liao, Q., Zhu, X., Liao, C.-F., & Chang, J.-S. (2013). Biohydrogen production by a novel integration of dark fermentation and mixotrophic microalgae cultivation. International Journal of Hydrogen Energy, 38(35), 15807-15814.
Lo, Y.-C., Chen, W.-M., Hung, C.-H., Chen, S.-D., & Chang, J.-S. (2008). Dark H 2 fermentation from sucrose and xylose using H 2-producing indigenous bacteria: feasibility and kinetic studies. Water research, 42(4), 827-842.
Mandal, S., & Mallick, N. (2009). Microalga Scenedesmus obliquus as a potential source for biodiesel production. Applied microbiology and biotechnology, 84(2), 281-291.
Mandotra, S., Kumar, P., Suseela, M., & Ramteke, P. (2014). Fresh water green microalga Scenedesmus abundans: a potential feedstock for high quality biodiesel production. Bioresource technology, 156, 42-47.
Marquez, F. J., Nishio, N., Nagai, S., & Sasaki, K. (1995). Enhancement of biomass and pigment production during growth of Spirulina platensis in mixotrophic culture. Journal of Chemical Technology and Biotechnology, 62(2), 159-164.
Masojídek, J., Torzillo, G., Kopecký, J., Koblížek, M., Nidiaci, L., Komenda, J., . . . Sacchi, A. (2000). Changes in chlorophyll fluorescence quenching and pigment composition in the green alga Chlorococcum sp. grown under nitrogen deficiency and salinity stress. Journal of applied Phycology, 12(3-5), 417-426.
Mata, T. M., Martins, A. A., & Caetano, N. S. (2010). Microalgae for biodiesel production and other applications: a review. Renewable and sustainable energy reviews, 14(1), 217-232.
Milne, T. A., Evans, R. J., & Nagle, N. (1990). Catalytic conversion of microalgae and vegetable oils to premium gasoline, with shape-selective zeolites. Biomass, 21(3), 219-232.
Mohan, S. V., Bhaskar, Y. V., Krishna, P. M., Rao, N. C., Babu, V. L., & Sarma, P. (2007). Biohydrogen production from chemical wastewater as substrate by selectively enriched anaerobic mixed consortia: influence of fermentation pH and substrate composition. International Journal of Hydrogen Energy, 32(13), 2286-2295.
Mohan, S. V., & Devi, M. P. (2012). Fatty acid rich effluent from acidogenic biohydrogen reactor as substrate for lipid accumulation in heterotrophic microalgae with simultaneous treatment. Bioresource technology, 123, 627-635.
Moon, M., Kim, C. W., Park, W.-K., Yoo, G., Choi, Y.-E., & Yang, J.-W. (2013). Mixotrophic growth with acetate or volatile fatty acids maximizes growth and lipid production in Chlamydomonas reinhardtii. Algal Research, 2(4), 352-357.
Mujtaba, G., Choi, W., Lee, C.-G., & Lee, K. (2012). Lipid production by Chlorella vulgaris after a shift from nutrient-rich to nitrogen starvation conditions. Bioresource technology, 123, 279-283.
Nakanishi, A., Aikawa, S., Ho, S.-H., Chen, C.-Y., Chang, J.-S., Hasunuma, T., & Kondo, A. (2014). Development of lipid productivities under different CO2 conditions of marine microalgae Chlamydomonas sp. JSC4. Bioresource technology, 152, 247-252.
Nalewajko, C., Colman, B., & Olaveson, M. (1997). Effects of pH on growth, photosynthesis, respiration, and copper tolerance of three Scenedesmus strains. Environmental and experimental botany, 37(2-3), 153-160.
Ortiz Montoya, E. Y., Casazza, A. A., Aliakbarian, B., Perego, P., Converti, A., & de Carvalho, J. C. M. (2014). Production of Chlorella vulgaris as a source of essential fatty acids in a tubular photobioreactor continuously fed with air enriched with CO2 at different concentrations. Biotechnology progress, 30(4), 916-922.
Ozmihci, S., & Kargi, F. (2011). Dark fermentative bio-hydrogen production from waste wheat starch using co-culture with periodic feeding: effects of substrate loading rate. International Journal of Hydrogen Energy, 36(12), 7089-7093.
Perez-Garcia, O., Escalante, F. M., de-Bashan, L. E., & Bashan, Y. (2011). Heterotrophic cultures of microalgae: metabolism and potential products. Water research, 45(1), 11-36.
Piorreck, M., Baasch, K.-H., & Pohl, P. (1984). Biomass production, total protein, chlorophylls, lipids and fatty acids of freshwater green and blue-green algae under different nitrogen regimes. Phytochemistry, 23(2), 207-216.
Quéméneur, M., Hamelin, J., Benomar, S., Guidici-Orticoni, M.-T., Latrille, E., Steyer, J.-P., & Trably, E. (2011). Changes in hydrogenase genetic diversity and proteomic patterns in mixed-culture dark fermentation of mono-, di-and tri-saccharides. International Journal of Hydrogen Energy, 36(18), 11654-11665.
Rai, M. P., & Gupta, S. (2017). Effect of media composition and light supply on biomass, lipid content and FAME profile for quality biofuel production from Scenedesmus abundans. Energy conversion and management, 141, 85-92.
Rajfur, M. (2013). Algae as a source of information on surface waters contamination with heavy metals. Ecological Chemistry and Engineering. A, 20(10).
Ramanan, R., Kim, B.-H., Cho, D.-H., Ko, S.-R., Oh, H.-M., & Kim, H.-S. (2013). Lipid droplet synthesis is limited by acetate availability in starchless mutant of Chlamydomonas reinhardtii. FEBS letters, 587(4), 370-377.
Reitan, K. I., Rainuzzo, J. R., & Olsen, Y. (1994). Effect of nutrient limitation on fatty acid and lipid content of marine microalgae 1. Journal of Phycology, 30(6), 972-979.
Ren, H.-Y., Liu, B.-F., Kong, F., Zhao, L., Xing, D., & Ren, N.-Q. (2014). Enhanced energy conversion efficiency from high strength synthetic organic wastewater by sequential dark fermentative hydrogen production and algal lipid accumulation. Bioresource technology, 157, 355-359.
Renaud, S., & Parry, D. (1994). Microalgae for use in tropical aquaculture II: Effect of salinity on growth, gross chemical composition and fatty acid composition of three species of marine microalgae. Journal of applied Phycology, 6(3), 347-356.
Richmond, A. (2008). Handbook of microalgal culture: biotechnology and applied phycology: John Wiley & Sons.
Sagnak, R., Kargi, F., & Kapdan, I. K. (2011). Bio-hydrogen production from acid hydrolyzed waste ground wheat by dark fermentation. International Journal of Hydrogen Energy, 36(20), 12803-12809.
Santos, C., Ferreira, M., Da Silva, T. L., Gouveia, L., Novais, J., & Reis, A. (2011). A symbiotic gas exchange between bioreactors enhances microalgal biomass and lipid productivities: taking advantage of complementary nutritional modes. Journal of industrial microbiology & biotechnology, 38(8), 909-917.
Sato, A., Matsumura, R., Hoshino, N., Tsuzuki, M., & Sato, N. (2014). Responsibility of regulatory gene expression and repressed protein synthesis for triacylglycerol accumulation on sulfur-starvation in Chlamydomonas reinhardtii. Frontiers in plant science, 5, 444.
Schmidt, L. E., & Hansen, P. J. (2001). Allelopathy in the prymnesiophyte Chrysochromulina polylepis: effect of cell concentration, growth phase and pH. Marine Ecology Progress Series, 216, 67-81.
Seppälä, J. J., Puhakka, J. A., Yli-Harja, O., Karp, M. T., & Santala, V. (2011). Fermentative hydrogen production by Clostridium butyricum and Escherichia coli in pure and cocultures. International Journal of Hydrogen Energy, 36(17), 10701-10708.
Sharma, A. K., Sahoo, P. K., Singhal, S., & Patel, A. (2016). Impact of various media and organic carbon sources on biofuel production potential from Chlorella spp. 3 Biotech, 6(2), 116.
Suen, Y., Hubbard, J., Holzer, G., & Tornabene, T. (1987). TOTAL LIPID PRODUCTION OF THE GREEN ALGA NANNOCHLOROPSIS SP. QII UNDER DIFFERENT NITROGEN REGIMES 1. Journal of Phycology, 23, 289-296.
Turpin, D. H. (1991). Effects of inorganic N availability on algal photosynthesis and carbon metabolism. Journal of Phycology, 27(1), 14-20.
Valdez-Ojeda, R., González-Muñoz, M., Us-Vázquez, R., Narváez-Zapata, J., Chavarria-Hernandez, J. C., López-Adrián, S., . . . Medrano, R. M. E.-G. (2015). Characterization of five fresh water microalgae with potential for biodiesel production. Algal Research, 7, 33-44.
Wahidin, S., Idris, A., & Shaleh, S. R. M. (2013). The influence of light intensity and photoperiod on the growth and lipid content of microalgae Nannochloropsis sp. Bioresource technology, 129, 7-11.
Widjaja, A., Chien, C.-C., & Ju, Y.-H. (2009). Study of increasing lipid production from fresh water microalgae Chlorella vulgaris. Journal of the Taiwan Institute of Chemical Engineers, 40(1), 13-20.
Yap, B. H., Crawford, S. A., Dagastine, R. R., Scales, P. J., & Martin, G. J. (2016). Nitrogen deprivation of microalgae: effect on cell size, cell wall thickness, cell strength, and resistance to mechanical disruption. Journal of industrial microbiology & biotechnology, 43(12), 1671-1680.
Yokoi, H., Mori, S., Hirose, J., Hayashi, S., & Takasaki, Y. (1998). H2 production from starch by a mixed culture of Clostridium butyricum and Rhodobacter sp. M [h] 19. Biotechnology letters, 20(9), 895-899.
Yoo, C., Jun, S.-Y., Lee, J.-Y., Ahn, C.-Y., & Oh, H.-M. (2010). Selection of microalgae for lipid production under high levels carbon dioxide. Bioresource technology, 101(1), S71-S74.
Zeng, X., Danquah, M. K., Zhang, S., Zhang, X., Wu, M., Chen, X. D., . . . Lu, Y. (2012). Autotrophic cultivation of Spirulina platensis for CO2 fixation and phycocyanin production. Chemical Engineering Journal, 183, 192-197.
Zhang, C., Yang, H., Yang, F., & Ma, Y. (2009). Current progress on butyric acid production by fermentation. Current microbiology, 59(6), 656-663.
吳僑鳴. (2014). 以廢水養殖藻類之研究. 中山大學海洋環境及工程學系研究所學位論文, 1-98.
施瑄育. (2016). 發展產氫菌微小化培養系統. 成功大學化學工程學系學位論文, 1-85.
蔡翊家. (2015). 微藻類於微型生物反應器之培養及分析. 成功大學化學工程學系學位論文, 1-97.