一般之in vitro終點培養法操作簡便迅速，但是無法瞭解瘤胃降解動態變化。In vitro氣體生成法能記錄不同培養時間點之氣體生成量，可用來預估芻料的降解動態變化。以百慕達草、梯牧草、不同割期狼尾草及盤固草(四、六及八週)，分別進行in vitro氣體生成及in situ降解率試驗，而in vitro氣體生成法除記錄各培養時間點之氣體生成量外同時測定其降解率。試驗結果經非線性模式分析顯示，百慕達草、梯牧草、狼尾草及盤固草之48小時潛在氣體生成量依序為33.76、49.76、40.76及44.18 mL，而氣體生成速率則分別為5.83、4.26、5.46及5.20 mL/h。在in vitro乾物質、有機物及中洗纖維之有效降解率方面，均顯示溫帶梯牧草較熱帶狼尾草、盤固草及百慕達草高。並以氣體生成量與in vitro及in situ降解率間進行迴歸分析，顯示經培養24小時後，氣體生成量與in vitro乾物質表面降解率（ADDM）、乾物質真降解率（TDDM）、有機物表面降解率（ADOM）及有機物真降解率（TDOM）均呈顯著相關(R2=0.58、0.65、0.85及0.82, P<0.05)，且48小時氣體生成量與48小時in vitro 之ADDM、TDDM、ADOM及TDOM亦呈顯著相關(R2=0.64、0.82、0.69及0.74, P<0.05)。在氣體生成量與in situ降解率間之相關方面，經培養16、24及48小時後，氣體生成量與乾物質降解量均呈顯著相關(R2=0.66、0.81及0.56, P<0.05)，而有機物降解量與氣體生成量在培養24及48小時後亦呈顯著相關(R2 =0.62及0.76,P<0.05)，此外氣體生成速率與in situ之乾物質及有機物降解速率也呈顯著相關(R2=0.57及0.76,P<0.05)。因此僅需要少量樣品且低成本之in vitro 5-6個時間點培養之氣體生成法可替代in situ尼龍袋及in vitro終點培養法做為快速評估芻料動態降解變化之方法，同時亦可獲得有關微生物產量及瘤胃發酵過程之能量利用效率之資訊。

In vitro digestion is a simple method which uses end point digestion measurement without evaluate dynamic changes during fermentation. In vitro gas production method however measured the gas production and changes of nutrient at different time points during incubation to predict dynamic changes in ruminal degradation. The aim of there studys was to predict forage degradation via method of in vitro gas production and of in situ nylon bag. Forage used in this trial including bermuda, timothy hay and local produced napier and pangola grass harvested at 4, 6, and 8-week of maturity. Result of gas production curve analyzed using Non-Linear Model show that potential gas production was in the order of 33.76, 49.76, 40.82, and 44.26 ml, the rate of gas production was 5.8, 4.3, 5.6, and 5.2 ml/h, respectively for the respective forage. The correlation of degradation using gas production, in vitro and in situ were apparent dry matter degradation (ADDM), true dry matter degradation (TDDM), apparent organic matter degradation (ADOM) and true organic matter degradation (TDOM) significantly correlated (R2=0.58、0.65、0.85及0.82, P<0.05) at 24 hour and 48 hour of incubation (R2=0.64、0.81、0.69及0.74, P<0.05). In addition, significantly correlated between gas production and in situ of DDM at 16, 24and 48h (R2=0.66、0.81及0.56, P<0.05), and of DOM at 24 and 48h (R2 =0.62及0.76, P<0.05). The rate of gas production and of in situ DM and OM degradation were also significantly correlated (R2=0.57 and 0.76, P<0.05). The small sample required and less cost in vitro gas production method can readily replace both in situ and in vitro methods in forage evaluation.

目錄 頁次
壹、中文摘要----------------------------------------------01
貳、前言--------------------------------------------------02
參、文獻檢討----------------------------------------------03
一、比較in vivo與in vitro瘤胃氣體生成原理-----------------03
1.瘤胃內氣體來源------------------------------------------03
2.In vitro之氣體生成--------------------------------------04
3.瘤胃氣體生成量與微生物合成量之關係----------------------05
二、氣體生成法之應用--------------------------------------06
1.預估芻料降解率及代謝能----------------------------------06
2.芻料採食量----------------------------------------------07
3.預估抗營養因子之影響------------------------------------08
三、影響in vitro氣體生成之因子及測定條件------------------09
1.溫度----------------------------------------------------10
2.pH值----------------------------------------------------10
3.含氧濃度------------------------------------------------11
4.瘤胃液來源----------------------------------------------11
5.氨濃度--------------------------------------------------12
四、比較in vitro氣體生成法與in situ尼龍袋法---------------13
肆、材料與方法--------------------------------------------14
Ⅰ、試驗一、in vitro氣體生成法----------------------------14
一、試驗材料----------------------------------------------14
二、瘤胃液之收集------------------------------------------14
三、測定分法與分析----------------------------------------17
(一)降解率之測定步驟--------------------------------------17
(二)氣體測定及樣品收集------------------------------------18
(三)分析方法----------------------------------------------18
1.樣品之成分分析------------------------------------------18
2.總揮發性脂肪酸之測定------------------------------------18
3.個別揮發性脂肪酸之測定----------------------------------18
4.二氧化碳濃度之測定--------------------------------------19
5.烷濃度之測定--------------------------------------------19
Ⅱ、試驗二、in situ尼龍袋法-------------------------------20
一、試驗材料----------------------------------------------20
二、試驗動物及飼養管理------------------------------------20
三、原位評估程序------------------------------------------20
四、分析項目----------------------------------------------21
Ⅲ、數據分析與計算----------------------------------------21
Ⅳ、統計分析----------------------------------------------22
伍、結果與討論--------------------------------------------23
Ⅰ、牧草之化學組成----------------------------------------23
Ⅱ、In vitro氣體生成法評估牧草降解率----------------------25
一、牧草之氣體生成----------------------------------------25
二、牧草降解率--------------------------------------------27
三、揮發性脂肪酸------------------------------------------32
四、牧草之中洗纖維降解率----------------------------------41
五、In vitro微生物產量------------------------------------43
六、In vitro氣體生成與降解率之關係------------------------46
Ⅲ、化學計量法與實際測定牧草降解特性之比較----------------48
一、化學計量法--------------------------------------------48
二、化學計量法與實際測定牧草降解特性之比較----------------52
Ⅳ、In situ尼龍袋法評估牧草降解率-------------------------56
一、牧草之乾物質降解率------------------------------------56
二、牧草之有機物降解率------------------------------------58
三、牧草之中洗纖維降解率----------------------------------60
四、牧草之蛋白質降解率------------------------------------62
五、In vitro氣體生成法與in situ尼龍袋法之相關-------------65
陸、結論--------------------------------------------------68
柒、參考文獻----------------------------------------------69
捌、英文摘要----------------------------------------------76
表次 頁次
表1. In vitro培養微生物發酵葡萄糖生成酸產物及氣體---------05
表2. 試驗用乾乳期瘤胃開窗荷蘭牛之試驗日糧組成及營養成分---15
表3. 人工唾液組成-----------------------------------------16
表4. 牧草之化學組成---------------------------------------24
表5. 牧草之in vitro氣體生成量及非線性參數值---------------26
表6. 牧草之in vitro乾物質表面降解率、降解參數值及有效降解率-28
表7. 牧草之in vitro乾物質真降解率、降解參數值及有效降解率-29
表8. 牧草之in vitro有機物表面降解率、降解參數值及有效降解率-33
表9. 牧草之in vitro有機物真降解率、降解參數值及有效降解率---34
表10. In vitro培養24小時後牧草發酵產生之揮發性脂肪酸量------39
表11. In vitro培養48小時後牧草發酵產生之揮發性脂肪酸量------40
表12. 牧草之in vitro中洗纖維降解參數值及有效降解率----------42
表13. 牧草之in vitro微生物量及微生物氮量--------------------45
表14. In vitro氣體生成量與乾物質表面降解率、乾物質真降解率、有機物表面降解率及有機物真降解率之關係--------------------------47
表15. 化學計量方法計算牧草於24小時之氣體量、基質消耗量、ATP及partitioning factor (PF)------------------------------------49
表16. 化學計量方法計算牧草於48小時之氣體量、基質消耗量、ATP及partitioning factor (PF)------------------------------------50
表17.牧草於 in vitro培養24及48小時後，測定二氧化碳、甲烷、真降解量、氣體生成量及partitioning factor (PF)-----------------55
表18. 牧草之in situ乾物質降解率、降解參數值及有效降解率-----57
表19. 牧草之in situ有機物降解率、降解參數值及有效降解率-----59
表20. 牧草之in situ中洗纖維降解參數值及有效降解率-----------61
表21. 牧草之in situ蛋白質降解參數值及有效降解率-------------63
表22. In vitro氣體生成量及生成速率(mL / 200mg or % / h)與in situ乾物質及有機物降解率(%)及降解速率(% / h)之相關-------------67
圖次 頁次
圖1. In vitro培養狼尾草發酵生成之總揮發性脂肪酸-------------36
圖2. In vitro培養盤固草發酵生成之總揮發性脂肪酸-------------37
圖1. In vitro培養狼尾草、百慕達草及梯牧草發酵生成總揮發性脂肪酸-------------------------------------------------------------38

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