水是人類最不可或缺的物質之一，而我們生活用水的來源：自來水其臭味問題是人們辨別飲用水質乾淨與否最直接之方法，也是最為大眾所詬病之處。淨化水質的各種物化處理方式對水庫中之臭味物質雖有或多或少的去除率，但其初設及維護成本過高亦為實際應用之考量。本研究利用本實驗室自行分離並經分子生物技術鑑定之Bacillus菌屬，添加不同共代謝基質濃度，探討實驗菌種對於水中臭味物質之降解效率，以求得最適當之共代謝基質添加比例。

本篇研究包含了菌種生長曲線以及從0.2%~1.0%五組不同乙醇添加比例的批次實驗，由生長曲線結果可知，二株菌屬生長情形相似，培養3小時後即大量生長，生長世代則為10小時。批次實驗結果發現，共代謝基質的添加確實可提高菌種對臭味物質的降解效率，且菌種對2-MIB之降解效率遠優於Geosmin。不論共代謝基質添加比例為何，降解反應之最佳期程皆為9天。當乙醇添加比例為0.6%時，菌種B-1與B-2對2-MIB之降解效率為所有添加比例中最佳之組別，分別可達21.38%與19.99%，二株菌種差異不大；效率其次為添加比例為0.8%之組別，降解效率分別為16.40%與14.71%；添加比例為1.0%之組別效果最差，整體效率不到5%。

由重鉻酸鉀密閉迴流法推算乙醇有效使用率時發現在乙醇添加比例為0.6%時效果最佳，菌種B-1與B-2乙醇有效使用率超過79%以上，此與共代謝降解結果相符，低添加比例下乙醇幾乎耗盡，對照其降解結果可發現，應是乙醇過早用完使降解反應停滯而導致最終效率不若高添加比例之組別。因此添加0.6%乙醇作為共代謝基質為兼具實質效益與成本考量之最適比例。

關鍵字：2-MIB、Geosmin、土霉臭味物質、生物降解、共代謝

Water is one of the most important necessities in human life. However, the order problem in the tap water has long become an annoying problem to solve, although order is a direct way to detect the quality of water. Various ways of water purification and deodorizations designed for reservoirs are proved to be effective but way too expensive in terms of installation and maintenance. The purpose of the study is to explore how Bacillus, separated voluntarily and identified through the molecular biological techniques in the laboratory, can help to degrade the order compounds in water when added with different substrate concentrations for co-metabolisms. The study is designed to find out the best proportion of the co-metabolism substrate addition.

The method of the study involved the bacteria culture (B-1 and B-2) and the five groups of different proportions of ethanol addition, from 0.2% to 1.0%. The research findings showed that two bacteria share similar growth cycle—growing massively 3 hours after being raised, with the growth generation for 10 hours. This results of the study indicated that adding the substrate for co-metabolism really enhanced the efficiency of order-compounds degradation by the test-bacterium. It is found that the degradation to 2-MIB is more efficient than to Geosmin. The best period of degradation is found to be 9 days, regardless the proportion of co-metabolism substrate. Moreover, when the ethanol proportion is 0.6%, the degradation reached the best effect for both bacteria (B-1 and B-2) to 2-MIB, with the efficiencies of B-1 and B-2 as 21.38% and 19.99%. No significant difference was found. The group with ethanol proportion as 0.8% reached less efficiencies, which are 16.40% and 14.71. The lowest efficiencies fell to less than 5% with the group that increased ethyl alcohol proportion to 1.0%.

Through the Potassium Dichromate closed reflux method, this researcher also found that ethanol reached the most effective utilization with proportion of 0.6% . In that proportion, B1 and B2 has utilized the ethanol up to 79%, which matched the effects of co-metabolism.

Keywords: 2-MIB, biodegradation, co-metabolism, Geosmin, musty/earthy orders

參考文獻

行政院環境保護署環境檢驗所，「水中Geosmin及2-Methylisoborneol檢測方法建立專案計畫」，民國九十一年。

李俊德、陳是瑩、王月花、鄭明蕊、李明輝，「澄清湖水源臭味控制方法之研究」，國立成功大學環境工程研究報告第45 號，民國七十四年。

汪俊育、林財富，「鳳山與港西淨水場臭味問題之研究」，第十七屆自來水研討會論文集，pp.49-65，民國八十九年。

林財富等，「自來水配水管網中影響感官物質之調查與改善對策」，行政院環保署環境檢驗所期末報告，民國九十一年。

高小萍，「水中異臭味物質的分析」，國立成功大學環境工程研究所碩士論文，民國八十七年六月。

徐尉晏，「應用分生技術鑑定澄清湖水庫之放線菌及探討營養源氮磷之影響」，國立成功大學環境工程研究所碩士論文，民國九十四年六月。

翁玉芬，「水中超微量有機異臭味物質Geosmin 之分析及去除」，國立交通大學環境工程研究所碩士論文，民國八十二年六月。

陳是瑩、李俊德、曾怡禎，「澄清湖浮游生物與放線菌繁殖狀況對水源臭味之影響(一)」，國立成功大學環境工程研究報告第21 號，民國七十一年。

陳是瑩、李俊德、曾怡禎，「澄清湖浮游生物與放線菌繁殖狀況對水源臭味之影響(二)」，國立成功大學環境工程研究報告第21 號，民國七十二年。

陳重男、張家源、陳俊明、翁玉芬，「高級處理對水中微量有機物去除效能評估：子題(二)自來水中有機異臭味物質去除之研究」，行政院環保署(EPA-82-E3-J1-09-03)，民國八十二年。

許雅雯，「應用批次與連續流試驗探討實驗室分離之除臭菌株對水庫中臭味物質降解之能力」，國立成功大學環境工程研究所碩士論文，民國九十五年七月。

童淑珠，「水源中2-MIB與geosmin 鑑定與氧化之研究」，國立成功大學環境工程學系博士論文，民國九十五年六月。

劉韋廷，「水庫水質條件對放線菌產臭與臭味物質降解之影響」，國立成功大學環境工程學系碩士論文，民國九十七年七月。

網站，http://www.epa.gov.tw，行政院環保署，環境水質監測。

Allen, E. J., Taste and Odor Problem in New Reservoirs in Wooded Areas. J.AWWA, 52, 1027-1032, 1960.

Ashitani, K., Y. Hishida and K. Fujiwara. Behavior of Musty Odourous Compounds during the Process of Water Treatment. Wat. Sci. Tech., 20(8/9), 261-267, 1988.

Aoyama, K. Studies on the earthy-musty odors in natural water (IV).Mechanism of earthy-musty odor production of actinomycetes. Jour. Applied Bacteriol., 68, 405-410, 1990.

APHA, AWWA and WPCF. StaN.Dard Methods for the Examination of Water and Wastewater, 19th ed, Washington, D.C., USA., 1995.

APHA. Standard method for the examination of water and wastewater, 5520 chemical oxygen demand, pp.5-12~5-16., 1995.

Atasi, K. Z., M. Porter, T. Chen, J. I. Huddleston, C. C. Young and I. H.Suffet. Factor Screening for Ozonating the Taste- and Odor-Causing Compounds in Source Water at Detroit, USA. Wat. Sci. Tech., 40, 115, 1999.

Anselme, C., J. Mallevialle and I.H. Suffet. Removal of Taste and Odors by the Ozone-Granular Activated Carbon Water Treatment Processes.Presented at 7th Ozone World Congress, Intl. Ozone Assoc., Tokyo, Japan.

ASTM , Standard Test Methods for Chemical Oxygen Demand ( Dichromate Oxygen Demand )of Water., D1252-06, 2006

Bentley, R. and Meganathan, R. Geosmin and methylisoborneol biosynthesis in streptomycetes evidence for an isoprenoid pathway and its absence in non-differentiating isolates. FEBS lett., 125(2), 220-222, 1981.

Blevins, W. T., Schrader,K. K. and Saadoun, L. Comparative physiology of Geosmin production by Streptomyces HalstedII and Anabaena sp., Wat. Sci. Tech., 31(11),127-133, 1995.

Bartak, P. and L. Cap. Determination of Phenols by Solid-phase Microextraction. J. Chromatogr. A, 767, 171-175, 1997.

Carmichael, W. W., The Cyanotoxins. Adv. Botanical Res. 27, 211–256, 1997.

Chance V. L., H. C. Aldrich and A. S. Lindner. Isolation and Characterization of a Bacterium Capable of Removing Taste- and Odor-causing 2-methylisoborneol from Water. Water Research, 38, 4135-4142, 2004.

Cecilie Klausen, Mette H. Nicolaisen, Bjarne W. Strobel, Falk Warnecke, Jeppe L. Nielsen , Niels O.G Jorgensen(2004) Abundance of actinobacteria and production of geosmin and 2-methylisoborneol in
Danish streams and fish ponds , FEMS Microbiology Ecology.

Dionigi, C. P., Millie, D. F., Spanier, A. M. and Johnsen P. B. Spore and geosmin production by Streptomyces tendae on several media. J. Agric. Food Chem., 40, 122-125, 1992.

Dionigi, C. P. and Ingram, D. A. Effects of temperature and oxygen concentration on geosmin production by Streptomyces tendae and Pencillium expansum. J. Agric. Food Chem., 42, 143-145, 1994.

Duguet, J., J. Mallevialle, J. Ho and I. H. Suffet. Oxidation Processes:
Chlorine and Chloramines, In Advances in Taste-aN.D-Odor Treatment and Control. Ed. by Suffet, I. H., Mallevialle, J., and Kawczynski, American Water Works Association, Denver, Coloroda, USA., 1995.

Edward, G. and M. J. McGuire. An Early Warning System for Taste and Odor Control. J. AWWA, 1986.

Frauke, C. P. and Berger, R. G. Geosmin and related volatiles in bioreactor-cultured Streptmyces citreus CBC 109.60. Appl. Environ. Microbiol. 57, 3429-3432, 1996.

Ferreira V., R. López, A. Escudero and J. Cacho. Quantitative Determination of Trace and Ultra Trace Flavour Active Compounds in Red Wines through Gas Chromatographic-ion Trap Mass Spectrometric Analysis of Microextracts. J. Chromatogr. A, 806, 349-354, 1998.

Gerber, N. N. and H. A. Lechevalier. Geosmin, An Earthy-Smelling Substance Isolated from Actinomycetes. Appl. Microbiol., 13, 935-938, 1965.

Gerber, N. N., Volatile Substances from Actinomycetes－their Role in the Odor Pollution of Water. CRC Crit. Rev. Microbiol., 7 (3), 191–214, 1979.

Glaze, W. H. et al., Evaluating Oxidants for the Removal of Model Taste and Odor Compounds from a Municipal Water Supply. J. AWWA, 82, 79-84, 1990.

Gramith, J. T., Oxidation Processes: Ozone, In Advances in Taste-and-Odor Treatment and Control. Ed. by Suffet, I. H., Mallevialle, J., and Kawczynski, American Water Works Association, Denver, Coloroda, USA., 1995.

Gunten, U. V., Ozonation of Drinking Water: Part I. Oxidation Kinetics and Product Formation. Water Research, 37, Issue: 7, April, 1443-1467, 2003.

Hoehn, R. C., Biological Methods for the Control of Tastes and Odors.
Southwest Water Works J., 47, 26-28, 1965.

Herzing, D. R., V. L. Snoeyink and N. F. Wood. Activated Carbon Adsorption of the Odor Compounds 2-Methylisoborneol and Geosmin. J. AWWA, 69, 223-238, 1977.

Huck, P.M., S. L. Kenefick, S. E. Hrudey and S. Zhang. Bench-scale
Determination of the Removal of Odor Compounds with Biological
Treatment. Wat. Sci. Tech., 31(11), 203–209, 1995.

Izaguirre, G., C. J. Hwang, S. W. Krasner and M. J. McGuire. Geosmin and 2-methylisoborneol from Cyanobacteria in Three Water Supply Systems. Appl. Env. Microbiol., 43, 708-714, 1982.

Izaguirre, G. and W. D. Taylor. A Pseudanabaena Species from Castaic Lake, California, that produces 2-methylisoborneol. Water Research, 32 (5), 1673–1677, 1998.

Izaguirre, G., W. D. Taylor and J. Pasek. Off-flavor Problems in Two
Reservoirs, Associated with Planktonic Pseudanabaena Species. Wat. Sci. Tech., 40 (6), 85–90, 1999.

Jensen S. E., C. Anders, L. Goatcher, T. Perley, S. Kenefick and S. Hrudey. Actinomycetes as a Factor in Odour Problems Affecting Drinking Water from the North Saskatchewan River. Water Research, 28, 1393-1401,1994.

Juttner F., Physiology and Biochemistry of Odourous Compounds from Freshwater Cyanobacteria and Algae. Wat. Sci. Tech., 31(11), 69-78, 1995.

Jia M., H. Zhang and D. Min. Optimization of Solid-phase Microextraction Analysis for Headspace Flavor Compounds of Orange Juice. J. Agric. Food. Chem., 46, 2744-2747, 1998.
Jelen H., K. Wlazly, E. Wasowicz and E. Kaminski. Solid-phase
Microextraction for the Analysis of Some Alcohols and Esters in Beer: Comparison with Static Headspace Method. J. Agric. Food. Chem., 46, 1469-1473, 1998.

Krasner S. W., C. J. Huang and M. J. McGuire. A Standard Method for
Quantification of Earthy-Musty Odorants in Water, Sediments, and Algal Cultures. Wat. Sci. Tech., 15(6/7), 127-138, 1983.

Kaas, H. and P. Henriksen. Saxitoxins (PSP toxins) in Danish Lakes. Water Research, 34 (7), 2089–2097, 2000.

Lalezary, S., M. Pirbazari and M.J. McGuire. Oxidation of Five Earthy-Musty Taste and Odor Compound. J. Am. Water Works Assoc., 78, 62, 1986.

Louch D., S. Motlagh and J. Pawliszyn. Dynamics of Organic Compound Extraction from Water Using Liquid-coated Fused Silica Fibers. Anal. Chem., 64, 1187-1199, 1992.

Lin, T. F., and C. Y. Fu. Origin and Control of Odors in Cheng-Chin Lake Water Treatment Plant. In Proceeding of 3rd International Workshop on Drinking Water Quality Management and Treatment Technology, 127-137, Taiwan, R.O.C., March 5-6, 1997.

Lloyd S. W., J. M. Lea, P. V. Zimba and C. C. Grimm. Rapid Analysis of Geosmin and 2-methylisoborneol in Water Using Solid Phase
Microextraction Procedures. Water Research, 32, 2140-2146, 1998.

Llompart M., K. Li and M. Fingas. Solid-phase Microextraction and
Headspace Solid Phase Microextraction for the Determination of
Polychlorinated Biphenyls in Water Samples. Anal. Chem., 70, 2510-2515, 1998.

Medsker, L. L., D. Jenkins, and J. F. Thomas. An Earthy-Smelling Compound Associated with Blue-Green Algae and Actinomycetes. Envir. Sci. Technol., 2, 461-464, 1968.

Martin, J. F. and Demain A. L. Control by phosphate of candicidin production. Biochem. Biophys Res Commun., 4, 1103-1110, 1976.

Madry, N. and Pape, H. Formation of secondary metabolism enzymes in the tylosin producer Streptomyces T59-235. Arch Microbiol., 131, 170-173, 1982.

Mallevialle J. and I. H. Suffet. Identification and Treatment of Tastes and Odors in Drinking Water. American Water Works Association Research Foundation/Lyonnaise des Eaux, AWWA, Denver, CO., 1987.

Oikawa, E., A. Ishibashi and Y. Ishibashi. 2-Methylisoborneol Degradation by the Cam Operon from Pseudomonas putida. Wat. Sci. Tech., 31 (11), 79–86, 1995.

Persson P. E., Off-flavours in Aquatic Ecosystems an Introduction. Wat. Sci. Tech. 15(6/7), 1-11, 1983.

Palmentier J. P., V. Y. Taguchi, S. W. Jenkins, D. T. Wang, K. P. Ngo and D. Robinson. The Determination of Geosmin and 2-methylisoborneol in Water Using Isotope Dilution High Resolution Mass Spectrometry. Water Research, 32, 287-294, 1998.

Paul, W., M. Rodriguez-Hernandez, L. Baker and M. Sommerfeld. Seasonal Occurrence and Degradation of 2-methylisoborneol in Water Supply Reservoirs. Water Research, 39, 4899-4912, 2005.

Silvey, J. K. G., D. R. Redden and W. C. McCormick. Actinomycetes and ommon Tastes and Odors. J. AWWA, 50(8), 1018-1026, 1959.

Suffet, I. H., GC/MS Identification of Trace Organics in Philadelphia
Drinking Water During a Two Year Period. Water Research, 14, 853, 1980.

Sivone, K. Influencing odor production by actinomycetes. Hydrobiology, 86, 165-170, 1982.
Suffet, I. H., J. Ho, D. Chou, D. Khiari, and J. Mallevialle. Taste and Odor Observed during Drinking Water Treatment, In Advances in
Taste-and-Odor Treatment and Control”, Ed. by Suffet, I.H., Mallevialle, J., and Kawczynski, American Water Works Association, Denver, Coloroda, USA., 1995.

Suffet, I. H. and O. Wable. Removal of Taste-and-Odor Compounds by
Activated Carbon. In Advances in Taste-and-Odor Treatment and Control, Ed. by Suffet, I.H., Mallevialle, J., and Kawczynski, American Water Works Association, Denver, Coloroda, USA., 1995.

Suffet, I. H., J. Duguet, and J. Mallevialle. Oxidation Processes: Potassium Permangnate, In Advances in Taste-and-Odor Treatment and Control, Ed. by Suffet, I.H., Mallevialle, J., and Kawczynski, American Water Works Association, Denver, Coloroda, USA., 1995.

Suffet, I. H., D. Khiari, A. Bruchet. The Drinking Water Taste and Odor Wheel for the Millennium: Beyond Geosmin and 2-methylisoborneol. Wat.Sci. Tech. 40 (6), 1–13, 1999.

Saito A., T. Tokuyama, A. Tanaka, T. Oritani and K. Fuchigmmi. Microbiological Degradation of (-)-Geosmin. Water Research, 33,3303-3036, 1999.

Susan, B. W., B. Brownlee, T. Satchwill and E. F. Hargesheimer. Quantitative Analysis of Trace Levels of Geosmin and MIB in Source and Drinking Water Using Headspace SPME. Water Research, 34, 2818-2828, 2000.

Schrader, K. K. and Blevins, W. T. Effect of carbon source, phosphorus concentration, and several micronutrients on biomass and geosmin production by Streptomyces halstedii. J. of Industrial Microbiology & Biotechnology. 26, 241-247, 2001.

Trudgill, P., Microbial Metabolism of Monoterpenes－Recent Developments. Biodegradation 1, 93–105, 1990.
Terauchi, N., T. Ohtani, K. Yamanaka, T. Tsuji, T. Sudou and K. Ito. Studies on a Biological Filter for Musty Odor Removal in Drinking Water Process. Wat. Sci. Tech., 31(11), 229–235, 1995.

Tanaka A., T. Oritani, F. Uehara, A. Saito, H. Kishita, Y. Niizeki, H. Yokota and K. Fuchigami. Biodegradation of a Musty Odour Compent, 2-Methylisoborneol, Wat. Res. Vol. 30, No.3, pp.759-761, 1996

Weete, J. D., Blevins, W. T., Wilt, G. R. and Durham, D. Chemical, biological and environmental factors responsible for the earthy odor in Auburn City water supply. Bull. Agr. Exp. Sta. Ala. 490, 1-46, 1977.

Wood, S., William, S. T., White, W. R. and Jones F. Factors influencing geosmin production by a streptomycete and their relevance to the occurrence of earthy taints in reservoirs. Wat. Sci. Tech., 15, 191-198, 1983.

Wnorowski A. U., Taste and Odours in the Aquatic Environment: a Review. Water SA, 18, 203-214, 1992.

Watson S., B. Brownlee, T. Satchwill and E. McCauley. The Use of Solid Phase Microextraction (SPME) to Monitor for Major Organoleptic Compounds Produced by Chrysophytes in Surface Waters. Wat. Sci. Tech., 40(6), 251-256, 1999.

Yagi, O., Sugiura, N. and Sudo, R. Chemical and physical factors in the production of musty odor by streptomyces spp. Isolated from lake Kasumigaura. Agric. Biol. chem., 51, 2081-, 1987.

Yagi, M., S. Nakashima and S. Muramoto. Biological Degradation of Musty Odor Compounds, 2-methylisoborneol and Geosmin in a Bio-Activated Carbon Filter. Wat. Sci. Tech., 20(8/9), 225-260, 1988.

Young W. F., H. Horth, R. Crane, T. Ogden and M. Arnott. Taste and Odour Threshold Concentrations of Potential Potable Water Contaminants. Water Research, 30, 331-340, 1996.

Zhang Z. and J. Pawliszyn. Headspace Solid-phase Microextraction. Anal. Chem., 65, 1843-1852, 1993.