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Language: korean

Conflict of Interest: In relation to this article, we declare that there is no conflict of interest.

Publication history: Received September 6, 2013
Accepted October 17, 2013

This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/bync/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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이온교환크로마토그래피에서 라이소자임 분리에 미치는 pH와 온도 영향

The Effect of pH and Temperature on Lysozyme Separation in Ion-exchange Chromatography

고관영 김인호^†

Kwan-Young Ko In-Ho Kim^†

충남대학교 화학공학과, 305-764 대전광역시 유성구 궁동 220

Department of Chemical Engineering, Chungnam National University, 220 Gung-dong, Yuseong-gu, Daejeon 305-764, Korea

ihkim@cnu.ac.kr

Korean Chemical Engineering Research, February 2014, 52(1), 98-105(8), 10.9713/kcer.2014.52.1.98 Epub 3 February 2014

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Abstract

라이소자임은 용균 작용, 조직 회복 과정의 촉진 등의 작용을 하며 난백 중에 0.3% 함유되어 있다. 난백에서 라이소자임을 분리하는 방법으로 친화성 크로마토그래피, 이온교환크로마토그래피, 한외여과법 등이 있는데 이 중 이온교환크로마토그래피가 가장 많이 사용된다. 라이소자임을 양이온 젤이 충전된 유리 칼럼에서 분리·정제할 때 최적의 pH와 온도 조건을 찾는 것을 실험 목표로 하고, ASPEN Chromatography 전산 모사의 결과와 비교하였다. 실험에 사용되는 완충용액은 인산완충용액이었고 pH를 5~8로 변화를 주어 상온에서 실험하였고, 가장 분리가 잘 이루어진 pH에서 온도를 5 ℃ 간격으로 25~40 ℃로 변화시켜 실험하였다. RP-HPLC (Reversed phase High Performance Liquid Chromatography) 분석을 통해 라이소자임의 체류 시간을 확인하였고, OriginPro 8을 이용해 용출 단계에서 크로마토그램의 면적을 비교하여 라이소자임의 양을 정량분석하였다. 결과를 분석한 결과, pH 5일 때, 온도가 25 ℃에서 가장 많은 양의 라이소자임이 분리되었다.

Lysozyme amounts to 0.3% in egg white and functions as an agent of cell lysis and activator of tissue reconstruction. Ion exchange chromatography is the most useful method of separation among affinity chromatography, ion exchange chromatography, and ultra-filtration. The aim of present study is to find the optimum pH and temperature for the separation of lysozyme in egg white within cation exchange gel filled glass column. And we compared results of experiments with those of simulations. Phosphate buffer was used, and pH and temperature were varied as 5~7 and 25~40 ℃ respectively. RP-HPLC was the tool for the retention time identification and quantitative analysis of lysozyme. OriginPro 8 measured the peak area of lysozyme chromatogram and quantified the eluted lysozyme. Largest amount of lysozyme was separated under the conditions of pH 5 and T 25 ℃.

Keywords

Lysozyme Ion Exchange Chromatography Aspen Simulation

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