臺灣博碩士論文加值系統

許多人爲了顧及體態、健康、環保等因素轉而增加食用素食飲食，使得素食飲食被掀起一股熱潮。長期的素食飲食面對的一大問題便是蛋白質不足，原因主要有二，一是植物蛋白胺基酸組成不完全，二是植物蛋白較難消化吸收。對此，益生菌的添加便成爲解決的辦法之一。本研究將以特性分析實驗、體外培養實驗和體内動物實驗等多個角度，探討植物乳桿菌Lactobacillus plantarum-1 (Lp1)與Lactobacillus plantarum-2 (Lp2)在生物體内促進植物蛋白水解並促進胺基酸吸收的能力。
在特性分析實驗中，益生菌Lp1無論是生長或代謝能力、耐受酵素性和耐膽鹽性皆略優於Lp2。於體外實驗，可觀察到Lp1及Lp2對大豆水解蛋白(SPI)與豌豆水解蛋白(PPI)皆具備基本的水解能力。但深入探討時，發現同樣是培養4小時後，與控制組相比，益生菌Lp1能顯著(p < 0.05)提高SPI中13種胺基酸的濃度，而Lp2只能提高(p < 0.05) 6種胺基酸的濃度。將益生菌與PPI共同培養4小時後，Lp1顯著(p < 0.05)提高PPI中6種胺基酸的濃度，Lp2僅對3種胺基酸的濃度有提升(p < 0.05)效果。由此初步判斷Lp1對植物蛋白之水解能力凌駕於Lp2之上。
在短期動物飼養模式中，大鼠在同時攝食含有1 mL濃度為Log 9 CFU/mL的益生菌與高蛋白飲食，益生菌Lp1可提高實驗動物血液必需胺基酸(EAA)濃度約82%，而Lp2只能提高約69.4%。在長期的動物飼養模式下，給予動物1 ml濃度為Log 7 CFU/mL和1 ml濃度為Log 9 CFU/mL兩種劑量的益生菌，觀察大鼠在不同飲食下長期攝取益生菌對其血液中胺基酸濃度的影響。結果顯示無論是Chow diet (CD)或高蛋白飲食組(HPD)，攝取低劑量益生菌Lp1或Lp2對胺基酸的影響主要為非必需胺基酸，而高劑量的Lp1或Lp2 則能顯著(p < 0.05)增加EAA的濃度。
綜合以上結果，益生菌Lp1不論是在特性還是在功效上皆展現出較佳的結果。兩株益生菌對蛋白質都具備一定的水解效果，而經過益生菌的代謝後，Lp1能較為有效的促進較多胺基酸在濃度上的提升。透過動物實驗，發現兩株菌在高劑量下有助餐後血液中EAA濃度的提高，但相比之下，Lp1比Lp2為血液EAA濃度帶來更明顯的提升效果，基於植物蛋白並非完全蛋白質，常見會有一至二種必需胺基酸含量略低，綜合考量下，將植物蛋白搭配益生菌Lp1同時食用有助改善植物蛋白的營養價值。

Many people have started adopting a vegetarian diet due to factors like maintaining body shape, health benefits, and environmental considerations, leading to a surge in the popularity of vegetarianism. A major challenge faced by long-term vegetarian diets is the lack of sufficient protein, mainly due to two reasons: the incomplete amino acid composition of plant proteins and the difficulty in digesting and absorbing plant proteins. To address this issue, the addition of probiotics has become one of the solutions. This study aims to investigate the ability of Lactobacillus plantarum-1 (Lp1) and Lactobacillus plantarum-2 (Lp2) to promote the hydrolysis of plant proteins and the absorption of amino acids in the body from various perspectives, including characteristic analysis experiments, in vitro culture experiments, and in vivo animal experiments.
In the characteristic analysis experiments, Lp1 demonstrated slightly superior growth or metabolic capacity, enzyme tolerance, and bile salt tolerance compared to Lp2. In the in vitro experiments, both Lp1 and Lp2 exhibited basic hydrolysis ability on soy protein isolate (SPI) and pea protein isolate (PPI). However, upon further investigation, it was found that after 4 hours of cultivation, Lp1 significantly (p < 0.05) increased the concentration of 13 amino acids in SPI compared to the control group, while Lp2 only increased the concentration of 6 amino acids (p < 0.05). After 4 hours of co-culturing probiotics with PPI, Lp1 significantly (p < 0.05) increased the concentration of 6 amino acids in PPI, whereas Lp2 only improved the concentration of 3 amino acids (p < 0.05). From this preliminary assessment, it was determined that Lp1 has a superior hydrolysis ability on plant proteins compared to Lp2.
In the short-term animal feeding model, rats were fed a high-protein diet with 1 mL of probiotics at a concentration of Log 9 CFU/mL. Probiotic Lp1 increased the concentration of essential amino acids (EAA) in the blood of experimental animals by approximately 82%, while Lp2 increased it by about 69.4%. In the long-term animal feeding model, animals were given two doses of probiotics: 1 mL at a concentration of Log 7 CFU/mL and 1 mL at a concentration of Log 9 CFU/mL. The effect of long-term probiotic intake on amino acid concentration in the blood of rats under different diets was observed. The results showed that whether under a Chow diet (CD) or high-protein diet (HPD), the intake of low-dose probiotics Lp1 or Lp2 primarily affected non-essential amino acids. However, high-dose Lp1 or Lp2 significantly (p < 0.05) increased the concentration of EAAs.
In summary, probiotic Lp1 showed better results both in characteristics and efficacy. Both probiotics demonstrated a certain degree of protein hydrolysis effect, and after probiotic metabolism, Lp1 was more effective in promoting the increase in the concentration of more amino acids. Through animal experiments, it was found that both strains at high doses contributed to the rise in postprandial blood EAA concentration. However, Lp1 showed a more significant effect on increasing blood EAA concentration than Lp2. Given that plant proteins are not complete proteins and usually have one or two essential amino acids in slightly lower quantities, it is concluded that consuming plant proteins with probiotic Lp1 can help improve the nutritional value of plant proteins.

摘要 i
Abstract iii
目錄 v
表次 ix
圖次 xi
1. 前言 1
1.1 益生菌 1
1.1.1 定義及歷史 1
1.1.2 益生菌對腸道的健康效益 2
1.1.3 益生菌的分類 3
1.1.4 植物乳桿菌 5
1.1.5 益生菌分泌的酵素類型 8
1.2 胺基酸的代謝 9
1.2.1 常見胺基酸的代謝路徑 9
1.2.2 胺基酸的相互代謝作用 9
1.2.3 益生菌在胺基酸代謝作用中所扮演的角色 11
1.3 胺基酸於生物體内所扮演的角色 12
1.3.1 血液胺基酸變化之意義 12
1.3.2 蛋白質對人體的重要性 13
1.4 蛋白質營養不良 14
1.4.1 蛋白質營養不良之定義 14
1.4.2 植物性蛋白與蛋白質營養不良 15
1.4.3 益生菌與植物蛋白之間的聯繫 17
2. 研究目的 19
3. 材料與方法 20
3.1 實驗模式 20
3.2 樣品來源 20
3.3 特性分析實驗 22
3.3.1 益生菌之生長曲線與產酸能力 22
3.3.2 益生菌於腸道模擬環境中的存活能力 22
3.3.3 益生菌之耐膽汁鹽能力 25
3.4 體外實驗 27
3.4.1 水解植物蛋白活性 27
3.4.2 植物蛋白水解液混合培養實驗 27
3.4.3 胺基酸相互代謝分析試驗 29
3.5 體内實驗 30
3.5.1 動物飼養 30
3.5.2 短期動物實驗 30
3.5.3 長期動物實驗 31
3.5.3.1 長期動物實驗分組 31
3.5.3.2 飼料與樣品的給予 32
3.6 液相層析系統 36
3.6.1 樣品上清液製備 36
3.6.2 柱前衍生化 36
3.6.3 色譜系統 37
3.6.4 溶劑選擇以及洗脫條件 37
3.7 糞便短鏈脂肪酸萃取與定量 38
3.8 統計分析 40
4. 結果與討論 41
4.1 活性與耐受性檢測試驗 41
4.1.1 生長曲線與產酸能力 41
4.1.2 耐酸、耐酵素試驗 44
4.1.3 耐膽鹽試驗 46
4.2 體外試驗 50
4.2.1 培養基蛋白質水解試驗 50
4.2.2 植物蛋白混合培養試驗 54
4.2.2.1 益生菌Lp1及Lp2水解SPI的潛力評估 55
4.2.2.2 益生菌Lp1及Lp2水解PPI的潛力評估 58
4.2.2.3 Lp1及Lp2對SPI和PPI胺基酸濃度提升程度之比較 62
4.2.3 胺基酸轉換能力分析 62
4.2.3.1 益生菌Lp1及Lp2與NEAAsl溶液共同培養分析 64
4.2.3.2 益生菌Lp1及Lp2與EAAsl溶液共同培養分析 67
4.3 體内實驗 71
4.3.1 大鼠體重、攝食量及飲水量之變化 71
4.3.2 短期動物實驗 72
4.3.3 長期動物實驗 75
4.3.3.1 益生菌Lp1搭配一般飲食對大鼠血清胺基酸的影響 80
4.3.3.2 益生菌Lp1搭配高蛋白飲食對大鼠血清胺基酸的影響 80
4.3.3.3 益生菌Lp2搭配一般飲食對大鼠血清胺基酸的影響 81
4.3.3.4 益生菌Lp2搭配高蛋白飲食對大鼠血清胺基酸的影響 82
4.3.4 大鼠糞便短鏈脂肪酸含量 83
4.3.4.1 益生菌Lp1對大鼠糞便短鏈脂肪酸的影響 88
4.3.4.2 益生菌Lp2對大鼠糞便短鏈脂肪酸的影響 88
5. 總結 90
6. 參考文獻 92

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