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본 연구는 기능성 발효유인 케피어로부터 박테리오신 생산 유산균을 분리하고 이 균주의 배양학적 특성, 생산 박테리오신의 물리화학적 및 생화학적 특성을 분석하였다.
분리된 유산균주(CJNU 2008)의 16S rRNA 유전자 염기서열 분석결과 Enterococcus faecium으로 동정되어 E. faecium CJNU 2008로 명명하였다.
E. faecium CJNU 2008 균주의 항균스펙트럼을 확인한 결과 병원성 미생물(Listeria monocytogenes, Staphylococcus aureus)을 포함한 일부 균주를 억제하는 것이 확인되었다.
E. faecium CJNU 2008 균주의 박테리오신 생산 최적조건은 MRS broth, 30℃ 및 pH 7.0으로 나타났다.
부분 정제된 박테리오신은 산과 알칼리에서 내성을 보이나 중성 부근에서 가장 높은 활성을 보이며 열에 대한 안정성(100℃에서 30분간 활성 유지)이 우수하였다. 또한 유기용매(Ethanol, Methanol,Acetonitrile, Acetone)에 대한 안정성이 확인되었고 Lipase와 α-Amylase 처리 시 활성이 유지된 반면 Protease 처리 시 활성이 소실되는 것으로 보아 본 항균물질이 단백질성의 박테리오신임을 알 수 있다. L. monocytogenes에 대한 박테리오신의 항균작용기작을 분석한 결과 살균작용(bactericidal)을 하는 것을 확인하였다. 배양 상등액의 acetone 추출을 통한 부분 정제된 박테리오신은 HPLC를 통해 최종 정제하였으며 향후 생화학적인 특성을 규명할 예정이다.
분리된 유산균주(CJNU 2008)의 16S rRNA 유전자 염기서열 분석결과 Enterococcus faecium으로 동정되어 E. faecium CJNU 2008로 명명하였다.
E. faecium CJNU 2008 균주의 항균스펙트럼을 확인한 결과 병원성 미생물(Listeria monocytogenes, Staphylococcus aureus)을 포함한 일부 균주를 억제하는 것이 확인되었다.
E. faecium CJNU 2008 균주의 박테리오신 생산 최적조건은 MRS broth, 30℃ 및 pH 7.0으로 나타났다.
부분 정제된 박테리오신은 산과 알칼리에서 내성을 보이나 중성 부근에서 가장 높은 활성을 보이며 열에 대한 안정성(100℃에서 30분간 활성 유지)이 우수하였다. 또한 유기용매(Ethanol, Methanol,Acetonitrile, Acetone)에 대한 안정성이 확인되었고 Lipase와 α-Amylase 처리 시 활성이 유지된 반면 Protease 처리 시 활성이 소실되는 것으로 보아 본 항균물질이 단백질성의 박테리오신임을 알 수 있다. L. monocytogenes에 대한 박테리오신의 항균작용기작을 분석한 결과 살균작용(bactericidal)을 하는 것을 확인하였다. 배양 상등액의 acetone 추출을 통한 부분 정제된 박테리오신은 HPLC를 통해 최종 정제하였으며 향후 생화학적인 특성을 규명할 예정이다.
목차
Ⅰ. 서 론 ················································································1Ⅱ. 실험재료 및 방법 ···························································61. 실험 재료 ··········································································61) 박테리오신 생산 균주 선발을 위한 재료 ····················62) 사용 균주 ·········································································63) 배지 ··················································································72. 실험 방법 ·········································································71) 박테리오신 생산 유산균주 분리 ··································72) 항균스펙트럼 ··································································93) 박테리오신 생산 최적 조건 조사 ·································94) 박테리오신 안정성 조사 ················································115) 박테리오신 작용 양상 ····················································146) 박테리오신 정제 ·····························································14Ⅲ. 연구 결과 ·······································································171. 박테리오신 생산 유산균주 분리 ··································172. 항균 스펙트럼 ·································································183. 박테리오신 생산 최적 조건 조사 ·································214. 박테리오신 안정성 조사 ················································265. 박테리오신 작용 양상 ····················································326. 박테리오신 정제 ·····························································34Ⅳ. 결 론 ···············································································38참고문헌 ···············································································40Abstract ················································································45
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