다양한 HPLC Column에서의 IgY(Immunoglobulin Yolk) 분리특성

Abstract

동물 혈청 중의 IgG (Immunoglobulin G)에 해당되는 난황에 포함된 면역 단백질 IgY (Immunoglobulin Yolk)는 식품 단백질로 장내 면역 물질로 중요하다. IgY를 정제하기 위해 신선란의 노른자에 카리지난이나 아라빅검을 전처리 물질로 사용하였다. 전처리 후 FPLC (Fast Protein Liquid chromatography)의 DEAE (Diethylaminoethyl) Sepharose 칼럼에서 이온교환법에 의해 불순물을 제거하여 IgY를 얻고, GF HPLC (Gel Filtration High Performance Liquid Chromatography)로 IgY의 분자량을 측정하고 표준 IgY와 비교하여 IgY 단백질을 동정하였다. GF HPLC에서 IgY의 다양성을 발견하였고 IgY 단백질 군의 다양성을 IE HPLC (Ion Exchange High Performance Liquid Chromatography)에서 AX, CX, SCX 칼럼을 사용하여 pH, NaCl 농도를 바꾸어 조사하였다. AX를 사용하여 0.5M NaCl, pH=8 조건에서 3개의 IgY 피크를 분리하였고, SCX를 이용했을 때 0.5M NaCl, pH=5 조건에서도 3개의 IgY 피크를 분리할 수 있었다. IgY (Immunoglobulin Yolk) in egg yolk corresponds to IgG (Immunoglobulin G) in animal serum and plays an important role as immunological proteins in intestines. Carrageenan and Arabic gum were used as pretreatment agents to purify IgY from fresh egg yolk. DEAE (Diethylaminoethyl) Sepharose column in FPLC (Fast Protein Liquid chromatography) was an ion exchange tool to remove contaminants as well as to elute IgY from the column. GF HPLC (Gel Filtration High Performance Liquid Chromatography) enables to measure the molecular weights of IgY and to identify the purified IgY by comparing the molecular weight of standard IgY with the purified one. IgY is a heterogeneous group of different molecular weight and ionic properties, which was investigated with various IE HPLC (Ion Exchange High Performance Liquid Chromatography) columns such as AX, CX and SCX. Three peaks of IgY were separated in the AX column under the conditions of 0.5 M NaCl and pH=8. The SCX column also gave the three peaks of IgY at 0.5 M NaCl and pH=5.