대두의 Flatulence Factor 제거를 위한 α-Galactosidase 효소제의 특성에 관한 연구

Abstract

본 연구는 大豆중에 존재하는 腸內 가스發生因子(flatulence factor)인 raffinose와 stachyose의 除去에 酵素劑를 이용하기 위해 시도되었다. 그리하여 6종의 市販用 酵素劑들의 α-galactosidase 力價를 측정하고 그 중에서 제일 높은 力價를 나타낸 Aspergillus niger에서 얻은 酵素劑의 特性을 조사하였다. 그리고 p-nitrophenyl-α-D-galactoside, raffinose, sucrose에 대한 加水分解 限度를 측정한 다음, raffinose, stachyose에 대한 加水分解 산물을 thin-layer chromatography로 살펴 보았다. 그 결과를 요약하면 다음과 같다. 1) 市販되는 6종의 酵素劑에 대하여 α-galactosidase 力價를 측정한 결과 Aspergillus niger에서 얻은 pectinase 제제가 가장 강력하였다. 2) 선정된 酵素劑의 特性을 조사한 결과 作用最適 pH는 4.0-4.5, 安定度最適 pH는 4 - 5 ,作用最適 온도는 45℃였다. 合成基質인 p-nitrophenyl -α-D-galactoside에 대한 Michaelis constant(Km)는 2.08mM, 最大反應速度(Vmax)는 435 μmoles substrate/minute/g enzyme preparation이었으며 阻害劑의 실험 결과 SH基를 필수로 하는 酵素로 밝혀졌다. 3) 本酵素劑는 3가지 基質인 raffinose, sucrose, p-nitrophenyl -α-D-galactoside를 거의 완전하게 加水分解하였으며, raffinose와 stachyose의 分解과정을 thin-layer chromatography로 조사한 결과 2시간 후에는 완전히 分解되어 단당류의 spot만이 나타났다. 4) 따라서 大豆食品중의 腸內 가스發生因子는 α-galactosidase 力價를 가지는 酵素劑의 이용에 의하여 쉽게 제거될 수 있을 것으로 생각한다.;This study was carried out to use commercial enzymes for the removal of raffinose and stachyose related to flatulence in soybean. Thus α-galactosidase activity of six commercial enzyme preparations was compared and enzymatic characteristics were investigated for the pectinase from Aspergillus niger which was shown the most potent α-galactosidase activity. The selected enzyme preparation was further investigated about the extent of hydrolysis toward p-nitrophenyl-α-D-galactoside, raffinose and sucrose, and for the hydrolysis products of raffinose and stachyose by means of thin-layer chromatography. The results are summarized as follows: 1) Among the commercially available enzyme preparations, a product from Aspergillus niger was shown to be the most potent, in α-galactosidase activity. 2) The enzyme characteristics of the selected preparation were shown to be pH 4.0-4.5 for optimum activity, pH 4-5 for optimum stability and 45℃ for optimum activity. Upon reaction on a synthetic substrate p-nitrophenyl-α-D-galactoside, Michaelis constant was 2.08 mM and maximum velocity was 435 micromoles of substrate/minute/g enzyme preparation. The enzyme was proved to be essential for SH group for its activity. 3) The selected enzyme preparation was capable of hydrolyzing three substrates raffinose, sucrose and p-nitropheyl-α-D-galactoside almost completely. Thin-layer chromatographic analysis exhibited that the enzyme treatment of raffinose and stachyose was resulted to produce only monosaccharides in 2 hours of hydrolysis. 4) It was, therefore, concluded that the flatulence factor in soybean foods can be easily removed by the use of enzyme preparations showing α-galactosidase activity.