在眾多生質能源原料中，木質纖維素被視為生質燃料和其他副產物主要源料，這是由於它們年產量大（4x109噸/年）的關係。狼尾草便是其中一種含有豐富纖維素的牧草，由於狼尾草具有容易種植、能在野生坡地等非種糧面積、生長速度快（一年可收割四次）、單位面積產量高、便宜、富含電子性（COD/TOC = 2.44 g COD/g TOC）的特點，所以我們以狼尾草作為這次產氫研究的主要基質，由於纖維素緊密且堅固的結構，加上其被半纖維素及木質素緊緊的包圍著，所以纖維素的水解成為整個生質能源的速率限制步驟。本研究分成兩個主要的部份，第一部份是開發一套生物系統---好氧式生物滴濾滲出床，以提高纖維素水解的效率。第二部份則是評估這套生物系統滲水液（產物）之厭氧生物產氫潛能。好氧式生物滴濾床為一多孔濾床固體醱酵反應器，用以承受高固體負荷及培養真菌優勢族群。另外，本研究加入廚餘作為輔助基質，用以提供額外容易利用的營養，如氮磷肥份，促進微生物生長。加入廚餘堆肥作為植種源，以提供大量及多樣性的、複合基質分解性的好氧微生物。利用自來水的沖洗，把狼尾草水解後的有機碳及流動性酵素沖出循環利用，這些有機碳帶著電子，供後續產氫程序產氫菌利用以促進產氫。
本研究先對狼尾草及廚餘的特性作分析及瞭解，狼尾草約有90%的固體物為有機體（TVS/TS=0.9）。固體態碳水化合物佔總乾重的70%，為一含固體碳水化合物豐富的基質，纖維素佔總乾重32%，COD/TOC為2.12，呈微氧化態碳，略低於狼尾草總質體比值。另一基質，廚餘為複雜有機廢棄物，富含高濃度的有機含量(VSS/SS)可達96%。總碳水化合物佔總乾重的40%，一半左右為可溶性的碳水化合物。總COD為1.53 g-O2/g-dry KW，溶解態的COD佔總COD的三分之一，COD/TOC為2.05，亦呈微氧化態有機碳，可被好氧菌分解。生物滴濾床植種源則為台南城西里好氧廚餘堆肥場，六槽廚餘堆肥中，各槽的總異營菌菌量，均在108 CFU/g-dry compost 左右。總異營真菌菌量則在106~108 CFU/g-dry compost。廚餘堆肥中有纖維素內切型水解酵素（Endo-β-1-4-glucanase）活性反應，隨著槽數（堆置時間）增加，酵素活性隨之增加，活性最高的為成品區腐熟廚餘堆肥，活性為1.4 U/g-dry compost。從DGGE分析中可見，廚餘堆肥中有多樣性的真菌族群，Candida, Fusarium, Trichoderma 均有在廚餘堆肥中出現。Aspergillus則沒有被發現。
本研究成功的把好氧式生物滴濾床從6 L放大至90 L（3年內），並進行半連續流自動化操作。在6 L好氧式生物滴濾床中，滲出混合液COD為49.5 g/L，主要成份為乳酸，約貢獻了49% COD，這些乳酸在後續的產氫潛能測試中，均被完全利用至低於儀器可偵測範圍。厭氧生物批次分解中，最高產氫速率及氫氣產率分別為2.13 L-H2 g-VSS-1d-1及0.04 L-H2 g-TCOD-1，可驗證廚餘經好氧分解之中間產物，可被厭氧產氫菌快速分解。經由T-RFLP的分生檢測分析結果得知，6 L 好氧式生物滴濾床出流水本身就富含產氫菌，且此產氫菌在批次馴養後會勝於連續流厭氧槽之植種菌源而成為優勢族群。經改良設計後，38 L 好氧式生物滴濾床成功的解決了濾床阻塞問題。在此廚餘混合狼尾草批分式好氧分解程序中，滲出液最高的總COD出現在第20天，為1,500 mg/L，滲出液淋洗出的總COD則佔原堆置物料總COD的1.2%。利用多因子實驗設計探討不同操作條件下，對好氧式生物滴濾床滲出液COD濃度之影響，發現在相同的出流水體積（即相同的水停留時間）下，增加沖水頻率，減少淋水量，比增加淋水量對沖出COD（在濃度及累積總量上）來得有效。最後改建的90 L好氧式生物滴濾床試程一中，為一次性流經濾床操作，水之停留時間為0.09天（2.16小時）。滲出液最高COD濃度出現在第7天，為1,200 mg/L。纖維素降解率為27%。試程二中，發現減少水固體塊狀物形成現象，並無助於滲出液中溶解態COD的增加。試程三中，增加水之停留時間至0.88天，滲出液最高總COD為1,200 mg/L，出現在批分式淋洗流程的第5天，纖維素的降解率為50%。試程四中，增加滴濾水之返送循環淋洗停留時間至8.85天，以促進混合基質之固態醱酵功能，滲出液最高總COD為1,500 mg/L，出現在第5天，纖維素的降解率為66%。因此，增加水之循環停留時間，能有效增加滴濾床內纖維素的降解率，但對於增加滲出液中總COD濃度，則沒有太大幫助。從DGGE分析中可見，從試程一到試程三，滴濾床內都有著十分多樣的真菌族群，Candida, Fusarium均有在廚餘堆肥中出現。Trichoderma , Aspergillus則沒有被發現。

Among potential bioenergy resources, lignocellulosics have been identified as the prime source of biofuels and other value-added products due to its enormous yield (about 4x109 tons/year). Napier Grass is a herbage which consists of abundant cellulose. Due to its advantages of easy planting, high-growing rate, high yield density per year, inexpensive and high electron density (COD/TOC = 2.44 g COD/g TOC), we decided to choose Napier Grass as the main substrate for this study. Cellulose, nevertheless, is difficult to hydrolyze due to its tight and rigid structure as well as the wrapping of hemicellulose and lignin. Therefore, cellulose hydrolysis is the rate limiting step in the whole bioenergy process. This study was divided into two main parts. The first part is to develop a biological system- Aerobic Bio-Leaching Bed (ABLB) for cellulose-hydrolyzing efficiency enhancement. The second part is to evaluate biohydrogen production potential utilizing the leachate of ABLB. Aerobic Bio-Leaching Bed is a solid-state fermentation bioreactor which can bear high-solid loading and is favorable for fungi cultivation. Kitchen waste is added as sub-substrate to provide extra-nutrient for encouraging microbial growth. Kitchen waste compost is added as seed to provide diverse and large quantity of microorganism. Tape water is added to flush out organic carbon which hydrolyzed from substrate and then the organic carbon which is carrying electron will go into the subsequent dark fermentation for hydrogen production.
The characteristics of Napier Grass and kitchen waste were analyzed in this study firstly. Napier Grass possesses 90% of organic solid（TVS/TS=0.9）. Solid carbohydrate and cellulose was 70% and 32% of dry weight respectively, so Napier Grass was a solid carbohydrate-abundant substrate. Its COD/TOC was 2.12. Kitchen waste is a complex organic compound which contain up to 96% of VSS. Total carbohydrate is 40% of dry weight and half of them is soluble. Total COD is 1.53 g-O2/g-dry KW and one third of them is soluble. COD/TOC is 2.05. Seeding source came from kitchen waste compost in Cheng-Chi-Li kitchen waste collect center, Tainan. The average heterotrophic aerobic bacteria and total heterotrophic fungi was about 108 and 106~108 CFU/g-dry compost. There was Endo-β-1-4-glucanase activity on kitchen waste compost and the longer time the kitchen waste compost was piled, the higher the activity was. The highest activity which was 1.4 U/g-dry compost appeared in the mature compost pile. According to DGGE profile, there was diverse fungi community on kitchen waste compost. Candida, Fusarium, Trichoderma existed in kitchen waste compost but Aspergillus had not been found among six tanks of kitchen waste compost.
In this study, the scale up of Aerobic Bio-Leaching Bed from 6 L to 90 L and automated operation is successful. The leachate of 6 L Aerobic Bio-Leaching Bed contained COD 49.5 g/L and about 49% of COD was contributed by lactic acid. Lactic acid was utilized almost completely in the subsequent bio-hydrogen production test. The highest specific hydrogen production rate and hydrogen yield were 2.13 L-H2 g-VSS-1d-1and 0.04 L-H2 g-TCOD-1 respectively. According to the results of T-RFLP, the leachate of 6 L Aerobic Bio-Leaching Bed contained abundant hydrogen-producing bacteria and they became dominant after the bio-hydrogen production test. 38 L Aerobic Bio-Leaching Bed which bioreactor design was improved solved the blocking problem happened in 6 L Aerobic Bio-Leaching Bed. The highest total COD in its leachate was 1,500 mg/L on day 20. The total COD mass flushed out by leachate during the whole operation recovered about 1.2% of initial substrate COD. In the factorial experiment discussing the effect of different operational factors of Aerobic Bio-leaching Bed to leachate COD, the results showed that in the same leachate volume, i.e. the same passed time of water, increasing the flushing frequency and decreasing influent water volume is more efficient than increasing the volume of influent water for leachate COD enhancement (in both concentration and cumulative amount). In run 1 of Aerobic Bio-Leaching Bed, HRT was 0.09 day. The highest total COD in leachate was 1,200 mg/L on day 7 and cellulose degradation is 27%. In run 2, the results showed that reducing particle agglomeration had no influence on increasing soluble COD in leachate. In run 3, HRT was increased to 0.88 day. The highest total COD in leachate was 1,200 mg/L on day 5 and cellulose degradation is 50%. In run 4, HRT was increased to 8.85 days. The highest total COD in leachate was 1,500 mg/L on day 5 and cellulose degradation is 66%. Therefore, increasing HRT was favorable for increasing cellulose degradation percentage but had no influence on increasing the total COD in leachate. According to DGGE profile, there was diverse fungi community in Aerobic Bio-Leaching Bed from run 1 to 3. Candida and Fusarium, existed in it but Trichoderma and Aspergillus had not been found.

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