本研究以雲芝 (Trametes versicolor LH1) 固態及液態發酵高粱酒糟 (sorghum distillery residue, SDR)，降低 SDR 之粗纖維含量，將雲芝酒糟 (f-SDR)取代魚粉作為肉食性魚飼料之配料，期能提升石斑魚攝食 f-SDR 之消化率及成長率。雲芝固態及液態發酵高粱酒糟於 28oC，7 天呈對數生長、7-28 天生長趨緩，進入平穩期，又以 21-28 天 f-SDR 之植物固醇及菌絲體生物量為最高。以液態發酵之木質纖維素酶活性，包括 cellulase (3.60 U/mg protein)、laccase (7.15 U/mg protein)、LIP (110.46 U/mg protein) 及 MNP (28.35 U/mg protein) 高於固態發酵。雲芝液態發酵高粱酒糟 7 天後，粗纖維含量由 11.95% 下降至 5.11 %，低於以固態發酵 28 天 (6.17%)，因此以雲芝液態發酵高粱酒糟 7 天為低纖維雲芝酒糟之最適發酵條件，其機能性成分含量為總多酚 (3093.22 g GAE/g)、總類黃酮 (122.22 g QE/g dw) 及三萜化合物 (31.18 g UAE/g dw)，但仍含抗營養因子含量：植酸 (130.09 g /g dw)、單寧 (0.53 mg CAE/g dw)，及胰蛋白酶抑制劑 (2.27 mg/g dw)。石斑魚攝食添加 20% 大豆粉發酵物 (soybean meal fermented products, f-SBM)、10、15、20% f-SDR 及20% SDR之飼料 8 週，以 20% SBM 及 15% f-SDR 二組之適口性、飼料利用率、比成長率皆無顯著差異 (p>0.05)，故高梁酒糟經雲芝菌發酵後，可提升其適口性及成長率並可發展為機能性飼料配方，15% 為雲芝高梁酒糟取代魚粉之最適比例。石斑魚分別攝食含高粱酒糟及雲芝菌發酵酒糟之飼料 8 週後，受低溫緊迫 (15oC) 維持 4 小時，以攝食 f-SDR 飼料，石斑魚之紅血球細胞膜較為完整、不易破裂，紅血球型態多呈橢圓狀，紅血球數、血比容積、血中溶氧、氧合紅血球及血色素較攝食 SDR 及 f-SBM 高，血漿葡萄糖及皮質醇濃度則顯著 (p<0.05) 低於攝食 SDR 及 f-SBM。綜合上述，高粱酒糟經雲芝液態發酵 7 天f-SDR 飼料之適口性與 f-SBM 相當可發展為抗低溫緊迫飼料配方，供石斑魚或其他養殖肉食性魚種之用。

This study examined the possibility of Trametes versicolor LH1 to reduce the crude fiber content of sorghum distillery residue (SDR) and the feasibility of replacing 20% fish meal in carnivorous fish diet by SDR fermented with T. versicolor LH1 (f-SDR). Submerged fermentation (SmF) and solid-state fermentation (SSF) of T. versicolor LH1 on SDR at 25oC showed a logarithmic growth phase during the first 7 days, followed by slow growth. The mycelium biomass and phytosterol content were the highest after 20 days of SmF fermentation. T. versicolor LH1 had high cellulolytic enzymes activities in SmF being higher than in SSF. SDR cultivated by SmF for 7 days, the crude fiber content reduced from 11.95% to 5.11%, which was lower than that by SSF for 28 days (6.17%). The optimal fermentation to obtain low-fiber f-SDR was obtained by SmF for 7 days. The bioactive compounds including total polyphenols (3093.22 g GAE/g), total flavonoids (122.22 g QE/g dw) and triterpenoids (31.18 g UAE/g dw) were obtained. The antinutritional factors of f-SDR including, phytic acid (130.09 mg /g dw), tannin (0.53 mg CAE/g dw) and trypsin inhibitor (2.27 mg/g dw) still remained. Five dietary feeds were separated into, 20% soybean meal fermented products (f-SBM) as control, 10% f-SDR, 15% f-SDR, 20 % f-SDR and 20% SDR fed grouper (Epinephelus coioides) for 8 weeks. At the end of feeding trail, there was no significant difference in palatability indicated by consuming time, feed utilization rate and specific growth rate between those groupers fed 20% f-SBM and 15% f-SDR groups. Therefore, 15% f-SDR was the optimal ratio of f-SDR to substitute for fish meal. Cold stress was performed at 15oC for 4 h on grouper previously fed diet containing 20% f-SDR showed higher hematocrit, red blood cell counts, hemoglobin concentrations, oxyhemoglobin and higher dissolved oxygen in blood than those fed 20% SDR or 20% f-SBM. The concentration of cortisol and glucose in plasma of the 20% f-SDR group was lower than those fed f-SBM group or SDR. Overall, the crude fiber content of SDR was significantly reduced by SmF for 7 days. Moreover, the reduction of crude fiber content improved the feed palatability and the growth rate, enhance anti-cold stress tolerance of grouper. These findings indicated that low-fiber f-SDR can be developed into functional feed for grouper or other carnivorous fish species.

壹、前言 1
貳、文獻回顧 2
一、低溫環境對石斑養殖之影響 2
二、酒糟乾物 (Distillers Dried Grains with Solubles, DDGS) 3
1. 酒糟乾物之功能特性 3
2. 營養價值 4
3. 飼料消化率 5
4. 酒糟乾物作為飼料配料之限制 5
三、高粱酒糟 6
1. 一般成分及營養價值 6
2. 機能性成分 7
2.1. 多酚化合物 7
2.2. 類黃酮 7
2.3. 脂肪醇、植物固醇 8
3. 抗營養因子 9
四、雲芝 (Trametes versicolor) 10
1. 雲芝之一般成分及營養成分 11
2. 雲芝多醣之生理活性 11
3. 雲芝三萜化合物 12
4. 雲芝降解纖維素之作用 12
5. 雲芝降解木質素之作用 14
參、實驗架構 15
一、高粱酒糟以雲芝固態及液態發酵 28 天，f-SDR 之成分分析 15
二、石斑攝食 f-SDR 飼料之養殖試驗 16
三、石斑攝食 f-SDR 飼料之低溫緊迫試驗 17
肆、材料方法 18
一、材料 18
1. 高粱酒糟 18
2. 雲芝 (Trametes versicolor LH1) 18
3. 點帶石斑 (grouper, Epinephelus coioides) 18
4. 試驗飼料 19
二、方法 21
1. 雲芝菌培養 21
2. 雲芝發酵高粱酒糟 (Trametes versicolor fermented-sorghum distillery residue, f-SDR) 22
3. 雲芝菌絲體含量測定 23
4. 木質纖維素酶活性測定 25
5. 一般成分分析 29
6. 纖維素、半纖維素及木質素含量測定 30
8. 抗營養因子含量分析 33
9. 體外試驗 35
10. 石斑養殖試驗 37
11. 降溫試驗 39
伍、結果與討論 44
一、雲芝降解高粱酒糟粗纖維之最適條件 44
1. 雲芝液態預活化 44
2. 菌絲體生物量之變化 49
3. 木質纖維素酶活性之變化 54
4. 產物之粗纖維含量 56
二、雲芝酒糟之機能性成分及抗營養因子 59
1. 總多酚、總類黃酮及三萜化合物含量 59
2. 植酸、單寧及胰蛋白酶抑制劑含量 61
三、低纖雲芝酒糟作為石斑養殖飼料之應用 63
1. 石斑之食性 63
2. 體外消化率 64
3. 適口性及飼料利用率 65
4. 成長率 67
四、石斑魚攝食低纖雲芝酒糟飼料於低溫緊迫之生理變化 69
1. 受低溫緊迫後水質之變化 69
2. 血漿生化指標 70
3. 紅血球形態之變化 75
4. 血液與血球參數 77
陸、結論 79
柒、參考文獻 82

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