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標題: Screening of lactic acid bacteria with activities to inhibit the enteropathogenic bacteria and to lower the cholesterol level as well as elucidation of the possible mechanisms and industrial application
作者: 林文鑫
Lin, Wen-Hsin
關鍵字: lactic acid bacteria;乳酸菌;pathogenic bacteria;cholesterol;病原菌;膽固醇
出版社: 食品暨應用生物科技學系
Lactic acid bacteria (LAB) have been extensively applied in food and feed field for a long time. The work of this study included evaluation of the potential effect of LAB on anti-Salmonella and anti-Helicobacter, and the function of LAB with active bile salt hydrolase (BSH) to improve the liver and plasma lipid profile in hamster. In addition, the optimal conditions of potential LAB fermentation and freeze-drying were investigated for assessing the industrialization.
For screening the mixed LAB with anti-Salmonella activity, Int-407 cells were used to estimate the protective effect against Salmonella invasion. The evaluation was further advanced to validate the protective effect against infection in vivo. Results showed that two group of mixed LAB have inhibited Salmonella invasion to mice. To study the mechanism of immune response in vitro, the results showed LAB has promoted phagocytosis and bactericidal activity etc. In SPF chicks test, we found that LAB population in the crop, ileum and cecum have increased after feeding Lactobacillus acidophilus LAP5 strain. In addition, growth properties of chicken have been improved slightly. Productions of IgA antibody in serum and intestine have also increased significantly in field trial. Therefore, there are many factors of LAB to protect mice from pathogenic infection including the LAB barrier to protect the intestinal cell, and enhance the immune system both humoral immunity and cellular immunity.
Helicobacter pylori (H. pylori) have developed strategies to colonize the mucous layer of human gastric epithelium. H. pylori have been thought a major causative factor of peptic ulcer disease, gastric adenocarcinoma and chronic gastritis in humans. Spent culture supernatant (SCS) of LAB was used to investigate the bactericidal activity to Helicobacter pylori. Results demonstrated organic acids in SCS are major bactericidal factors. Several reports indicated cholesterol play an important role in cardiovascular diseases. Probiotic LABs are defined as the microbial food supplements, which beneficially affect the host by improving its serum lipids level. In this study, the LAB strains with bile salt hydrolase (BSH) in vitro were screened and the efficiency to reduce hypercholesterolemia induced by fed high cholesterol diet in hamster was estimated. Results showed L. plantarum BV6 isolated from infants has the strongest BSH activity. In vivo test, this strain has reduced both cholesterol and triglyceride concentration in plasma and liver, and this effect is better than L. acidophilus ATCC 43121. Therefore, BV6 strain should be worthy to perform the clinical trail to assess the beneficial effect.
Finally, the optimal conditions of fermentation and freeze-drying for potential probiotics were studied. Afterwards, the fermented conditions of L. plantarum BV6 were carried into 300 liter bioreactor. The preliminary results showed the BV6 strain could meet the requirement for the industrial process.

乳酸菌為腸道中主要的有益菌,被廣泛應用於食品或飼料中。本研究內容主要包括篩選具有抗沙門氏菌及幽門桿菌感染,以及篩選可表現膽鹽水解酵素(Bile salts hydrolase, BSH)活性的乳酸菌,並以動物體內分析其降低膽固醇能力。論文最後對潛力菌株的發酵及冷凍乾燥條件進行探討,評估其產業化之可行性。
在篩選抗沙門氏菌之乳酸菌研究方面,本試驗以Int-407腸道細胞模式分析混合乳酸菌抑制沙門氏菌侵入細胞之能力,並進一步餵食小鼠以探討乳酸菌之保護效果,結果顯示篩選出的兩組混合乳酸菌有減少沙門氏菌侵入的效果。以體外試驗探討免疫機制,證實乳酸菌有明顯提升巨噬細胞的吞噬能力及殺菌作用等。在餵食無特定病原菌(SPF)的小雞實驗中,發現餵食Lactobacillus acidophilus LAP5菌株後嗉囊、迴腸及盲腸之乳酸菌數有明顯提高,當進行田間試驗餵食肉雞時,發現LAP5菌株有略微提升肉雞之生長性狀,並明顯提高腸道中IgA抗體濃度。因此,整體上來說,乳酸菌降低宿主遭受感染的因素可能有多項因子,包括形成障壁保護腸道細胞,以及增強宿主的體液免疫(humoral immunity)及細胞免疫(cellular immunity)反應。
幽門桿菌(Helicobacter pylori)為造成胃潰瘍和胃癌的重要原因,在乳酸菌抗幽門桿菌感染的研究上,發現乳酸菌所產生的有機酸為主要殺死幽門桿菌的原因。許多研究報告指出,膽固醇為造成心血管疾病的重要因子,而乳酸菌改善血脂質的效果也相當受到重視,本研究以體外試驗篩選具有表現膽鹽水解酵素活性的菌種,結果發現篩選自嬰兒的L. plantarum BV6乳酸菌具有高度膽鹽去結合的能力,餵食倉鼠證實所篩選的BV6菌株具有降低血液及肝臟中的膽固醇和三酸甘油酯(Triglycerides)的能力,且降低的效果較L. acidophilus ATCC 43121佳。因此,此菌株值得進一步進行人體試驗評估其在臨床上的有效性。
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