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首页> 外文学位 >Production of galacto-oligosaccharides from lactose by immobilized beta-galactosidase and posterior chromatographic separation.
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Production of galacto-oligosaccharides from lactose by immobilized beta-galactosidase and posterior chromatographic separation.

机译：通过固定的β-半乳糖苷酶和后色谱分离法从乳糖生产半乳糖低聚糖。

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摘要

Galacto-oligosaccharides (GOS) are non-digestible sugars containing two to five molecules of galactose and one molecule of glucose connected through glycosidic bonds. They are classified as prebiotic food because they can selectively stimulate the growth of bifidobacteria and lactobacilli in the lower intestine. The addition of GOS as a functional food ingredient has great potential to improve the quality of many foods. GOS can be produced from lactose, which is abundant in cheese whey, by enzymatic transgalactosylation with beta-galactosidase present in either free or immobilized form. The goal of this research was to evaluate the feasibility of using various microbial lactases immobilized on cotton cloth for GOS production from lactose and posterior purification by chromatographic technique using a commercial cation exchange resin Dowex 50W. The production of GOS from lactose was first studied with lactases from Aspergillus oryzae, Bacillus circulans, and Kluveromyces lactis. The total amount, types, and size of GOS produced were affected by the enzyme type and the initial lactose concentration in the reaction media. In general, more GOS can be produced when the initial lactose concentration was higher. With 400 g/L of lactose solution, a maximum GOS content of 40% (w/w) was achieved with B. circulans lactase, followed by K. lactis with 31% GOS and A. oryzae with 27% GOS. The addition of galactose inhibited the enzyme, reducing both GOS yield and the reaction rate.;The method of immobilizing enzyme on cotton cloth via adsorption with the binding support of polyethyleneimine (PEI) and cross linking with glutaraldehyde (GA) was developed for beta-galactosidase from B. circulans and A. oryzae. A high enzyme loading of 250 mg/g support with 35% and 90% activity yield was achieved for B. circulans and A. oryzae lactase, respectively. Thermal stability of B. circulans and A. oryzae lactase increased by 12 and 25-fold, respectively, upon immobilization on the cotton cloth. The immobilized enzyme showed the same GOS formation kinetics as that of the free enzyme, indicating that there was no diffusion limitation in the catalytic cotton cloth for the reaction.;Continuous production of GOS with immobilized enzyme in packed-bed reactors was studied. A high volumetric productivity of 4300 g/L/h was attained at 50% conversion with A. oryzae lactase and 180 g/L/h at 60% conversion with B. circulans lactase. Higher overall GOS productivity and conversion can be achieved with two enzyme reactors in sequence, with the first one containing A. oryzae lactase and second one containing B. circulans lactase.;Commercial gel-type cation exchange resins (Dowex 50W) with different salt forms (Na+, K+, and Ca++) and degrees of cross-linking (2%, 4%, and 8%) were evaluated for the separation of GOS from the multicomponent sugar mixture. Among all the factors studied, the degree of cross linking had the most significant effect on sugar adsorption capacity and selectivity, with 4% cross linking being the optimal. Other factors such as temperature and counter ion showed minimal effect. A high degree of removal of monosaccharides (95%) from the GOS mixture was achieved using an elution chromatographic column packed with Dowex 50W resins of 4% cross linking and in the Na+ form at room temperature.
机译：半乳糖低聚糖（GOS）是不可消化的糖，包含两到五个分子的半乳糖和一个分子的糖苷键连接的葡萄糖。它们被归类为益生元食品，因为它们可以选择性刺激下肠道中双歧杆菌和乳酸杆菌的生长。添加GOS作为功能性食品成分具有巨大潜力，可以改善许多食品的质量。通过用游离或固定化形式存在的β-半乳糖苷酶进行酶促半乳糖基化作用，可以从干酪乳清中丰富的乳糖中生产GOS。这项研究的目的是评估使用固定在棉布上的各种微生物乳糖酶从乳糖生产GOS以及使用市售阳离子交换树脂Dowex 50W通过色谱技术进行后纯化的可行性。首先用米曲霉，圆形芽孢杆菌和乳酸克鲁维酵母的乳糖酶研究了乳糖生产GOS的过程。产生的GOS的总量，类型和大小受酶类型和反应介质中初始乳糖浓度的影响。通常，当初始乳糖浓度较高时，可以产生更多的GOS。使用400 g / L的乳糖溶液，圆形双歧杆菌乳糖酶的最大GOS含量达到40％（w / w），其次是乳酸链球菌的GOS为31％，米曲霉的GOS为27％。半乳糖的添加抑制了该酶，降低了GOS的收率和反应速率。;开发了通过聚乙烯亚胺（PEI）的结合载体吸附并与戊二醛（GA）交联将酶固定在棉布上的方法。圆环芽孢杆菌和米曲霉的半乳糖苷酶。圆环芽孢杆菌和米曲霉乳糖酶分别达到250 mg / g的高酶负载量和35％和90％的活性收率。固定在棉布上后，B。circulans和米曲霉乳糖酶的热稳定性分别提高了12倍和25倍。固定化酶显示出与游离酶相同的GOS形成动力学，表明该反应在催化棉布中没有扩散限制。;研究了固定床反应器中固定化酶连续生产GOS。米曲霉乳糖酶在50％转化率下获得了4300 g / L / h的高生产率，而马氏芽孢杆菌乳糖酶在60％转化率下获得了180 g / L / h的高生产率。依次使用两个酶反应器可实现更高的总GOS生产率和转化率，其中第一个反应器包含米曲霉乳糖酶，第二个反应器包含圆环芽孢杆菌乳糖酶；具有不同盐形式的商业凝胶型阳离子交换树脂（Dowex 50W） （Na +，K +和Ca ++）（交联度）（2％，4％和8％）评估了多组分糖混合物中GOS的分离。在所有研究的因素中，交联度对糖的吸附能力和选择性影响最大，其中4％的交联为最佳。其他因素（例如温度和抗衡离子）显示的影响最小。使用装有4％交联的Dowex 50W树脂且在室温下呈Na +形式的洗脱色谱柱，可以从GOS混合物中高度去除单糖（95％）。

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