Gluconobacter oxydans의 당 전이효소 활성을 이용한 다당류의 제조 및 배양액의 품질 특성
Synthesis and physicochemical properties of polysaccharides by Gluconobacter oxydans with glycosyltransferase activity
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포도당의 주 사슬결합이 α-1,6 형태로 이루어져 있는 동종 다당체인 dextran은 유산균뿐만 아니라, 초산균에서 유래하는 dextran dextrinase(DDase)라는 당전이 효소작용에 의해 dextrin으로부터 합성되는 미생물 유래의 biopolymer이다. 본 연구에서는 DDase에 의해 당전이 활성이 있는 초산균을 다양한 배지 조성에 따라 배양함으로써 dextran 합성에 효과적인 배지 구성 조건을 탐색하고, 배지 내 생성된 다당류의 특성을 조사하고자 하였다. 즉, glycerol 첨가(2%, v/v) 유/무, maltodextrin 중합도 및 농도(1, 5%)에 따른 G. oxydans ATCC 19357의 생육곡선, pH, 산도, 환원당 및 점도 변화를 통하여 이화학적 특성을 분석하고, 각각의 배지에서 회수한 다당류의 구성당 패턴(HPAEC-PAD)및 결합형태(1H-NMR)를 조사하였다. 그 결과, glycerol 및 maltodextrin 첨가군에서 초산균 7일째 배양 배지의 OD값이 3배 가량 증가하여 배지조성에 따른 생육 증진효과를 확인하였다. G. oxydans ATCC 19357의 배양 7일째 배지의 pH는 2.97-3.45, 산도는 0.39-0.69로 나타났으며, 환원당은 1일차에 급격히 감소한 후 지속적으로 감소하였다. dextrin 5% 첨가 배지(G(+)AAB+ 5%MD1, G(+)AAB+5%MD2)의 점도는 배양 기간에 따라 증가하여 pseudoplastic한 성질을 나타내어 고분자 다당류가 생성되었음을 유추할 수 있었다. 특히 G(+)AAB+5%MD1 배지 배양액의 겉보기 점도는 5.53±0.05 mPa․s로 20% dextran 수용액과 유사한 수준이었다. 배양액 내 생성된 구성 당류의 패턴을 분석한 결과, dextrin 첨가 배지의 구성당 패턴과 달리 dextran과 유사한 패턴의 중합도가 높은 물질 peak가 관찰되었다. 이러한 다당류의 분자 간 결합 형태는 α-1,6 glycosidic bond를 형성한 물질에서 나타나는 4.9 ppm 위치에 chemical shift가 우세한 것으로 관찰되어, 첨가한 dextrin 이 초산균에 의해 dextran으로 생물 전환된 것을 확인하였다. 따라서 당전이 활성이 있는 초산균 배양 시 배지 구성으로 glycerol 및 5% maltodextrin(D.E. 4.0-7.0, MD2)의 첨가는 G. oxydans ATCC 19357의 증식 및 dextran으로 전환을 촉진함으로써 생물전환을 통한 dextran 제조에 효과적인 것으로 사료된다.
Abstract
Dextran, a homologous polysaccharide with the main chain of glucose units composed by an α-1,6 glycosidic bond, is synthesized from dextrin by dextran-dextrinase (DDase), a transglucosidase, derived from acetic acid bacteria Gluconobacter oxydans and lactic acid bacteria (LAB). The effective culture conditions were examined for producing dextran via bio-conversion with acetic acid bacteria (AAB) demonstrating DDase activity in various medium compositions during 0 to 7 days with or without glycerol addition (2%, v/v) and different degrees of dextrin polymerization (D.E.) based on the addition level (1,5%, w/v). On day 7, the G. oxydans growth was almost tripled in presence of glycerol as observed via a cell growth curve (OD). After culturing for 7 days, the pH decreased from 6 to 3.1-3.5, and the acidity increased from 0.12% to 0.4-0.62%, depending on the dextrin D.E. and the addition level. The reducing sugar decreased continuously. The medium containing 5% dextrin showed shear-thinning characteristics. The apparent viscosity of the 5% dextrin DE4-7 culture solution was 5.6 mPa·s, which was similar to that of the 20% dextran aqueous solution. The analysis of constituent saccharides contained in the culture medium (HPAEC-PAD) showed a substance with a high degree of polymerization. 1H-NMR analysis showed that α-1,6 glycosidic bond existed as the intermolecular bond of this substance. Therefore, efficient production of dextran was possible by culturing in a medium containing 5% dextrin and glycerol during culture of AAB.
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