Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/98068
標題: 以液化澱粉芽孢桿菌發酵黑水虻蛹殼作為雞隻機能性飼料添加物之應用
Assessment of Hermetia illucens pupal exuviae fermented by Bacillus amyloliquefaciens as functional feed additive for chickens
作者: 張哲綸
Che-Lun Chang
關鍵字: 液化澱粉芽孢桿菌
黑水虻
幾丁質
幾丁寡醣
紅羽土雞
Bacillus amyloliquefaciens
Hermetia illucens
chitin
chitooligosaccharides
Red Feather Native chicken
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摘要: 動物糞便的處理一直是畜牧業迫切需解決的問題。近年,發展出一種依靠生物轉換的處理方法,藉由將昆蟲飼養於這些有機廢棄物中,不僅能重新轉換其中的營養物質,也能減少廢棄物對於環境的汙染。黑水虻 (Hermetia illucens, BSF) 為一種腐食性昆蟲,幼蟲能利用廚餘、動物糞便及稻稈等有機物生長,並且因其含有豐富的蛋白質及脂質,而被證實能作為動物飼料的蛋白質替代原料。因此,在循環經濟的意識抬頭下,黑水虻也逐漸成為熱門的選項並被規模化飼養。然而,在黑水虻被大量飼養情況下,於生長期間所蛻下的蛹殼 (BSFP) 便成為了無法被利用的廢棄物。本研究即是針對此蛹殼廢棄物藉以液化澱粉芽孢桿菌(Bacillus amyloliquefaciens, BA) 進行發酵,評估作為雞隻機能性飼料添加物之潛力。幾丁質為BSFP 中主要的多醣類物質,在經過微生物發酵後能被降解成分子較小且具有多種生物機能性的幾丁寡醣 (COS)為本研究評估與應用重點。試驗分為三部分,第一部分為篩選最適合的BA菌株作為後續發酵試驗的接種菌株。藉由測定從飼料原料中分離出之五隻BA菌株 (P2、Y2、T4、T6、T7) ,依其特性及酵素分泌能力,篩選出最適合的菌株。結果顯示,Y2菌株具有最佳的生長速度、胃液耐受性、耐熱性及酵素生產能力。另外,Y2菌株亦具有嗉囊上皮細胞之吸附能力,因此選用Y2菌株作為後續發酵BSFP菌株。研究第二部分為BSFP的發酵試驗,此過程為兩階段,第一階段先以蛋白酶處理後,再接種BA進行第二階段的發酵。測定發酵物中的總菌數、還原醣含量、纖維素酶活性及幾丁質酶活性作為指標,篩選出最適當的發酵條件。由結果顯示,以3000 U/g BSFP活性的蛋白酶處理,再接種BA發酵五天後,能夠明顯誘導生產纖維素酶與幾丁質酶,並大量釋出還原醣。在此最佳發酵條件發酵製程之發酵蛹殼 (FBSFP)中,測得具有少量的幾丁二糖、幾丁三糖、幾丁五糖及幾丁六糖等寡醣類,且在體外的抗菌試驗結果中顯示具有抑制大腸桿菌生長的效果。在發酵物抗氧化試驗中顯示,FBSFP相對於BSFP來說有更佳的總酚含量、還原力、自由基清除力。最後為評估以FBSFP添加於土雞飼糧之應用性。試驗將紅羽母土雞隨機分配至五組,分別為餵飼基礎飼糧的控制組 (Control); 基礎飼糧中添加1x107 CFU/kg BA (BA)、基礎飼糧中分別添加0.5% BSFP (BSFP2)、0.25% FBSFP (FBSFP1)及0.5% FBSFP (FBSFP2),試驗期70日。結果顯示,各組在生長表現上皆沒有顯著差異,而於腸道菌相結果,較於控制組,各處理組皆能提升雞隻迴腸中的乳酸菌數,其中BA組又能顯著降低迴腸中的大腸桿菌群數量。腸道型態則於FBSFP1組表現出較控制組高的空腸絨毛與腺窩深度比。另外,各處理組皆降低雞隻血清中的丙二醛含量,FBSFP2組則能顯著提升血清中的過氧化氫酶(CAT)及穀胱甘肽過氧化物酶 (GSH-Px)活性。飼糧中添加FBSFP的雞隻皆產生較高的血清中腫瘤壞死因子-α (TNF-α)含量。肉品質部分結果顯示,飼糧中添加FBSFP有助於降低雞胸肉的L*值,並在0.5% 的添加量下能顯著降低腿肉的蒸煮失重率。測定各組的墊料性狀,發現添加BA及FBSFP的處理組能顯著降低墊料中的大腸桿菌數及吲哚含量。綜合以上結果,選用適當的液化澱粉芽孢桿菌菌株配合蛋白酶發酵處理黑水虻蛹殼之發酵物具潛力作為雞隻之機能性飼料添加物以改善腸道菌相及型態,且對於墊料性狀亦具正面效益。
The recycling of animal manure is becoming the major concern of animal production. Recently, feeding insects on organic waste can greatly lower the environmental pollution and re-boost their value. Hermetia illucens, commonly known as black soldier fly (BSF), able to be reared on the organic waste such as food waste, manure and rice straw. BSF larvae are recognized as alternative protein due to the high content of protein and lipid in larvae and prepupae. Therefore, under the rising awareness of the circular economy, BSF has gradually become a popular insect and been raised on a large scale. However, the pupal exuviae (BSFP) left during the growth period became waste. This study was aimed at assessing the BSFP fermented by Bacillus amyloliquefaciens (BA) as a functional feed additive for chickens. Chitin is the main polysaccharide in BSFP. Chitooligosaccharides (COS), which can be degraded by microbial fermentation and possess many biological functions, are the emphasis of this research. The first part of the study was to select the best strain from five BA strains (P2, Y2, T4, T6, T7) isolated from feedstuff for subsequent BSFP fermentation. The selection conducted through several in vitro assays. Results showed that the strain Y2 possesses best growth rate, gastrointestinal juice tolerance, heat tolerance, cellulase, protease and chitinase production ability. In addition, the possibility of Y2 adhering to the crop epitheliums of broiler was also discovered. The second part of the study was to discuss the BSFP fermentation. The fermentation was carried out in two steps. BSFP were treated by protease first and then inoculated BA Y2 to start fermentation. The total bacterial count, reducing sugar content, cellulase activity, and chitinase activity of the fermented product were analyzed as indicators for the best fermentation. Results showed that reducing sugar contents, cellulase activity and chitinase activity of FBSFP reached its peak after 3000 U/g protease treatment and after five days of fermentation. COS had been measured out in BSFP, while N-acetyl-chitobiose, N-acetyl-chitotriose, N-acetyl-chitopentaose and N-acetyl-chitohexaose contents were higher in FBSFP. FBSFP showed great antibacterial effect against E.coli and better total phenol contents, reducing power, DPPH radical-scavenging activity and ferrous chelating capacity than BSFP. The last part was to evaluate the application of FBSFP to chickens. Red Feather Native chickens were allocated into five groups and fed as follows: basal diet (Control), basal diet supplemented with 1x107 CFU/kg BA (BA), 0.5% BSFP (BSFP2), 0.25% FBSFP (contains 5x106 CFU/kg BA, FBSFP1), and 0.5% FBSFP (contains 1×107 CFU/kg BA, FBSFP2). The experiment lasted for 70 days. Although there were no differences on the growth performance (1-70 d) among all groups, the lactic acid bacteria counts in ileum was significantly increased in all groups and the coliform counts in ileum was significantly decreased in BA compared to the Control group. FBSFP1 resulted in higher jejunal villi height and crypt depth ratio. All treatments significant decrease the serum MDA contents of chickens. However, the serum CAT and GSH-Px activities of FBSFP2 were significantly higher than there of other groups. Supplementation with FBSFP could significantly increase serum TNF-α of chickens and significantly decreased L* of breast muscle, and meanwhile significantly decrease cooking loss of thigh muscle at the 0.5% supplementation. Except BSFP, supplementation of both BA and FBSFP (0.25% and 0.5%) in diets decreased the counts of coliform bacteria and indole concentration in the litter. In conclusion, selecting appropriate strain of BA to ferment BSFP with protease treatment might be a great way to utilize it. The final product of fermentation performed great potential as feed additive for chickens to improve the intestinal microflora, morphology as well as antioxidant capacity of chickens, and possessed positive benefits for litter properties.
URI: http://hdl.handle.net/11455/98068
文章公開時間: 2021-08-22
Appears in Collections:動物科學系

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