异麦芽寡糖（isomaltooligosaccharides，IMO）是葡萄糖以α-1，6糖苷键为主连接而成的低聚糖，具有低血糖指数特征和益生菌增殖功能。目前商业化IMO主流制备工艺得率较低，且含大量异麦芽糖，而IMO的益生功能主要归功于三糖及以上的高聚合度组分。利用黑曲霉（Aspergillus niger）来源的α-葡萄糖苷酶和嗜热脂肪芽孢杆菌（Bacillus stearothermophilus NO2）来源的环糊精葡萄糖基转移酶（CGTase）进行复配制备IMO，以获得得率和聚合度提升的IMO（命名为IMOH）。通过单因素实验，考察了反应pH、温度、时间、底物种类及加酶（α-葡萄糖苷酶）量对产物得率的影响，随后探究了复配酶（GTase）加酶量及反应时间的影响，结果表明在pH 5.5、50 ℃的反应条件下，以300 g/L的DE值为15~20的麦芽糊精为底物，当α-葡萄糖苷酶加酶量为5 U/g（以底物质量计）、CGTase加酶量为15 U/g（以底物质量计）时，IMOH产物得率达到64.27%，其中三糖及以上的高聚合度产物质量分数为71.57%。在对产物进行结构分析后发现，产物α-1，6糖苷键键型占比为70.67%，平均相对分子质量为706，高于市售IMO50。通过肠道微生物体外发酵实验，IMOH与市售IMO50相较，对青春双歧杆菌更具有增殖效果，同时可以进一步抑制肠道条件致病菌大肠杆菌的生长，具有一定的体外益生功能。该研究提出了一种双酶复配制备IMO的新工艺，为工业制备高得率、高聚合度的IMO提供了新思路。
Isomaltooligosaccharides (IMO) are oligosaccharides mainly composed of α-1,6 glycosidic bonds linked glucoses, which have physiological characteristics of hypoglycemic index and promoting probiotics proliferation. The current commercial IMO production has low yield and high isomaltose content in the product, while the probiotic function of IMO is mainly attributed to the high polymerization fractions like disaccharides and above. In this study, α-glucosidase from Aspergillus niger and cyclodextrin glucosyltransferase (CGTase) from Bacillus stearothermophilus NO2 were combined for synthesis of IMO with improved yield and polymerization (named IMOH). In this study, the effects of reaction pH, temperature, time, substrate type and enzyme addition on the yield were investigated by single-factor experiments, and the effects of compound enzyme amount and reaction time were subsequently explored. The results showed that the yield of IMOH reached 64.27%, and the high polymerization content of trisaccharides and above was 71.57% under pH 5.5 at 50 ℃, using 300 g/L maltodextrin with DE values ranges from 15 to 20 as the substrate, and with α-glucosidase of 5 U/g substrate (calculated by substrate mass) and CGTase of 20 U/g substrate (calculated by substrate mass). The structural analysis of IMOH products showed that the ratio of α-1,6 bond ratio was 70.67%, and the average molecular weight of products was 706, which was higher than that of commercial IMO50. The in vitro fermentation experiments of intestinal microorganisms suggested that compared with commercial IMO50, the IMOH had a better proliferative effect on Bifidobacterium adolescentis, and could further inhibit the growth of opportunistic bacteria Escherichia coli in intestinal tract, showing a certain in vitro probiotic effect. This study proposed a new process of the IMO synthesis by the dual-enzyme compounds, providing a new idea for the industrial synthesis of IMO with high yield and high degree of polymerization.
XU Ying, WANG Lei, WEI Beibei, SU Lingqia, CHEN Sheng, WU Jing. Synthesis of Isomaltooligosaccharides with High Degree of Polymerization and Its in vitro Probiotic Function Evaluation[J]. Journal of Food Science and Biotechnology,2023,42(1):66-74.