Korean Journal of Food Science and Technology (한국식품과학회지)

Korean Society of Food Science and Technology (한국식품과학회)
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Enhancement of Immune Activities of Peptides from Asterias amurensis Using a Nano-encapsulation Process

나노 입자 불가사리 펩타이드의 면역 활성 증진

  • Jeong, Hyang-Suk (Division of Biomaterials Engineering, Kangwon National University) ;
  • Oh, Sung-Ho (Division of Biomaterials Engineering, Kangwon National University) ;
  • Kim, Seoung-Seop (Division of Biomaterials Engineering, Kangwon National University) ;
  • Jeong, Myoung-Hoon (Division of Biomaterials Engineering, Kangwon National University) ;
  • Choi, Woon-Yong (Division of Biomaterials Engineering, Kangwon National University) ;
  • Seo, Yong-Chang (Division of Biomaterials Engineering, Kangwon National University) ;
  • Choi, Geun-Pyo (Department of Food Processing & Bakery, Gangwon Provincial University) ;
  • Kim, Jin-Chul (Division of Biomaterials Engineering, Kangwon National University) ;
  • Lee, Hyeon-Yong (Division of Biomaterials Engineering, Kangwon National University)
  • 정향숙 (강원대학교 생물소재공학과) ;
  • 오성호 (강원대학교 생물소재공학과) ;
  • 김승섭 (강원대학교 생물소재공학과) ;
  • 정명훈 (강원대학교 생물소재공학과) ;
  • 최운용 (강원대학교 생물소재공학과) ;
  • 서용창 (강원대학교 생물소재공학과) ;
  • 최근표 (강원도립대학 식품가공제과제빵과) ;
  • 김진철 (강원대학교 생물소재공학과) ;
  • 이현용 (강원대학교 생물소재공학과)
  • Received : 2009.12.01
  • Accepted : 2010.03.02
  • Published : 2010.08.31
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Abstract

Immuno-modulatory activities of peptides from Asterias amurensis were investigated using a nano-encapsulation process. The molecular weights of the peptides in the range of 5-7 kDa were separated using Sephadex G-75 gel filtration. Eighty-five percent of the nano-particles were in the 300 nm range using dynamic light scattering. The cytotoxicity of the A. amurensis nano-particles against CCD-986sk human dermal fibroblast cells was 11.64% after adding 1.0 mg/mL of the samples, which was lower than that from the control (13.28% collagen). The secretion of $NO^-$ from macrophages was estimated as $40\;{\mu}M$ after adding 1.0 mg/mL of gelatin nano-particles, which was higher than the others. Prostaglandin $E_2$ production from UV-induced human skin cells decreased greatly to 860 pg/mL after adding 1.0 mg/mL of the samples. Confocal microscopy revealed that nano-particles effectively penetrated the cells within 1 hour. From these results, we consider that nano-encapsulation of the peptides from A. amurensis can improve their biological functions.

본 연구는 기존의 불가사리 콜라겐 유래 펩타이드, 저분자화된 펩타이드, 저분자화 된 펩타이드의 젤라틴 나노입자의 비교를 통해 면역 활성의 증진을 확인하였다. 불가사리 펩타이드는 Sephadex G-75 gel 크로마토그래피 분리를 통해 5-7 kDa 범위의 분자량 범위의 시료를 얻었고, MALDI-TOF MS 분석에 의해 펩타이드의 정확한 분자량을 측정하였다. 0.1% collagenase를 처리한 펩타이드의 분자량은 기존의 불가사리 콜라겐 유래 펩타이드에 비해 약 2,000 m/z까지 분자량이 감소되는 효과를 보였다. 이는 2개의 폴리펩티드결합으로 상호 결합되어 감겨있던 콜라겐형구조가 collagenase의 작용으로 풀어진 후 효소의 영향을 받아 가수분해되었기 때문인 것으로 사료된다. 이후 $20^{\circ}C$에서 젤라틴으로 시료를 나노입자화 하여 TEM 및 DLS로 분포와 크기를 각각 확인하였다. 이후 인간섬유아세포에 대한 세포독성 측정 결과, 나노입자화 한 ALPG가 최고 농도인 1.0 mg/mL의 농도에서 11.64%로 가장 낮게 나타났다. 각 시료의 면역물질의 분비 정도를 확인하기 위해 macrophage에서의 $NO^-$ 분비를 측정하였는데 역시 나노 입자화 한 ALPG가 $40\;{\mu}M$로 가장 높은 $NO^-$ 생성량을 보였다. 또한 불가사리 나노 시료가 저분자 추출물과 비교해 B cell 생육도를 10% 이상 향상시켰다. UV 자극에 의해 형성되는 염증물질로 잘 알려진 prostaglandin의 생성을 정량하기 위해 $PGE_2$ 방법을 이용하였는데, 나노입자를 가해준 모든 군에서 농도 의존적으로 $PGE_2$의 생성이 감소하였고 역시 ALPG가 860 pg/mL로 $PGE_2$를 가장 적은 농도로 생성하였다. 마지막으로 confocal 현미경을 이용해 인간 면역 세포에 나노입자가 얼마나 효과적으로 침투하는지를 관찰하였는데 나노 시료의 경우 입자의 크기가 50-200 nm로 분산됨에 따라 일반 불가사리 펩타이드에 비해 세포 침투의 용이성이 크게 증진되는 것을 확인하였다.

Keywords

References

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