초록 ▼

Ⅳ. 연구개발결과
□ Agarase 및 한천올리고당 생산 공정 최적화 연구
○ agarase 분리 정제 조건 및 최적의 효소 활성 조건 확립
○ 한천올리고당의 최적 생산 조건 설정 및 표준화 위한 효소반응조건 탐색
□ 한천올리고당의 안전성(독성)시험 통한 안전성 확인
○ Annexin V FITC Assay(BMDC, HepG2 세포독성 없음 확인)
○ 랫드를 이용한 독성 시험(단회경구 투여독성, 2주반복 투여독성시험)
○ 복귀돌연변이 시험
○ 염색체 이상 시험
○ 소핵시험
○

Ⅳ. 연구개발결과
□ Agarase 및 한천올리고당 생산 공정 최적화 연구
○ agarase 분리 정제 조건 및 최적의 효소 활성 조건 확립
○ 한천올리고당의 최적 생산 조건 설정 및 표준화 위한 효소반응조건 탐색
□ 한천올리고당의 안전성(독성)시험 통한 안전성 확인
○ Annexin V FITC Assay(BMDC, HepG2 세포독성 없음 확인)
○ 랫드를 이용한 독성 시험(단회경구 투여독성, 2주반복 투여독성시험)
○ 복귀돌연변이 시험
○ 염색체 이상 시험
○ 소핵시험
○ 장기투여 안전성 시험
□ 한천올리고당의 항비만 효능 검증
○ 3T3-L1 세포 분화 측정: NAOS가 지방전구세포 분화에는 영향을 미치지 않음.
○ 지질 미토콘드리아의 지방 축적 개선 효능 확인 및 신호전달기구 규명
○ 비만동물모델(DIO)에서 NAOS의 항비만 효능 확인
□ 한천올리고당의 대사질환 관련 면역기능 증강 효능 및 신호전달기구 확인
○ 면역세포(Raw264.7, THP-1, BMDC)에서의 사이토카인 발현 증가 확인
○ TLR2-/-, TLR4-/-, TLR9-/- 마우스 유래 수지상 세포 실험(NAO의 면역세포에서의 수용체가 TLR4임을 확인)

Abstract ▼

Agar is a major cell wall component of marine red algae. It is composed of heterogeneous polysaccharides (galactans) mainly consisting of D-galactose and 3,6-anhydro-L-galactose units (agarobiose) alternately linked by β-(1,4) and α-(1,3) linkages. Agar oligosaccharides have also been reported to ha

Agar is a major cell wall component of marine red algae. It is composed of heterogeneous polysaccharides (galactans) mainly consisting of D-galactose and 3,6-anhydro-L-galactose units (agarobiose) alternately linked by β-(1,4) and α-(1,3) linkages. Agar oligosaccharides have also been reported to have antioxidant activity, therapeutic activity in inflammatory disease, and antitumor activity, etc. which may broaden its application in the food, cosmetic, and pharmaceutical industries as well as biorefinement or biofuel industries. In this study, thermophilic bacteria having agarolytic activity were isolated and characterized for the production of agarase. The growth condition was also investigated and optimized to produce agarase. The genes of agarase were cloned from Gayadomonas jooniniege, Pseudoalteromonas hodoensis H7, and Pseudoalteromonas sp. H9. The agarase was expressed from E. coli and investigated to get the optimum condition for hydrolysis of agar. The hydrolyzed products of agar, neoagarooligosaccharides (NAOS), was purfied to identify its antiobesity effect and verify its safety for clinical trial approval application.
The nontoxicity of NAOS was verified in Raw264.7, THP-1, BMDC, and HepG2 cell. The safety of NAOS was confirmed by single dose oral toxicity test, 2-week repeated oral dose toxicity test, the micronucleus test using CHL(Chinese Hamster Lung) cells, a chromosome aberration test on cultured CHL cells, bacterial reverse mutation test.
In the DIO(Diet induced obesity) models reflecting characteristics to appear in obese patients, it showed the result that was decreased within the statistically significant level in weight, weight increment, feed conversion rate in comparison with HFD group by the group which took fodder contains NAOS for 22 weeks. And the fat cell weight around kidney and epididymis showed a tendency to decrease, the fat cell size decreased significantly. As a result, we determine the effect of antiobesity of NAOS. In the DIO models which is an obese animal model, the HFD group showed the tendency that diabetes-related indexes worsened, but was able to confirm what was made the good portion leaders by taking NAOS. Besides, the OGTT and hematological test showed a positive diabetes improvement effect in diet which took NAOS, it was proof in possibility as the antidiabetes functionality material of NAOS.
Furthermore, an immunomodulating effect related to metabolic syndrome and a signal pathway of NAOS were identified.