Department of Food Science and Engineering, School of Food Science and Pharmaceutical Engineering, Nanjing Normal University, Nanjing, P.R. China.
Department of Food Science and Engineering, College of Food Science and Technology, Nanjing Agricultural University, Nanjing, P.R. China.
J Food Biochem. 2021 Feb;45(2):e13589. doi: 10.1111/jfbc.13589. Epub 2020 Dec 23.
β-glucosidase (BglA) immobilization from Thermotoga maritima on magnetic nanoparticles (MNPs) functionalized with chitosan (Cs) were efficiently investigated to improve lactose conversion and galactooligosaccharides (GOS) production. We used a batch method in order to improve the conversion of lactose to GOS. The efficiency and yield of immobilization were 79% and immobilized BglA was effectively recycled via a magnetic separation procedure through a batch-wise GOS with no activity lessening. Furthermore, analyses were done through screening kinetics of enzyme activity, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Fourier transform infrared spectroscopy (FT-IR), and transmission electron microscopy (TEM). Proposed methodology of immobilization shows a potential application as it is stable which was proved through many methods including pH, temperature, heat treatment, storage, and kinetics of the enzyme. GOS and residual enzyme activity showed to be 28.76 and 40.44%, respectively. However, free enzyme synthesis of GOS yield was just 24% after 12 hr. This study proposed applying magnet in the immobilization process of BglA on Cs-MNPs to produce GOS as new method for immobilizing enzyme in a biostable and cost-efficient way. PRACTICAL APPLICATIONS: This paper focus on immobilization of BglA from T. maritima onto MNPs functionalized with CS to investigate their further possibility improving lactose conversion and GOS production. Interestingly, a successful immobilization of Tm-BglA on the substrates were achieved in Cs-MNPs. The obtained results from enzyme activity, SDS-PAGE, FT-IR, and TEM showed that the high binding capacity of BglA to Cs-MNPs was successfully obtained. Furthermore, the binding efficiency calculation indicated that the immobilized BglA-Cs-MNPs conserved 40.44% of its native activity at the end of its 6th repeated use. In addition, magnetic separation technique was successfully employed for reuse of the immobilized BglA for repetitive batch-wise GOS without significant loss of activity.
β-葡萄糖苷酶（BglA）从海洋栖热菌（Thermotoga maritima）固定在壳聚糖（Cs）功能化的磁性纳米粒子（MNPs）上，可有效提高乳糖转化率和半乳糖低聚糖（GOS）产量。我们使用分批法来提高乳糖转化为 GOS 的效率。固定化效率和产率分别为 79%和 80%，并且通过磁分离程序可有效回收固定化 BglA，在批次 GOS 中没有活性损失。此外，还通过酶活性筛选动力学、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）、傅里叶变换红外光谱（FT-IR）和透射电子显微镜（TEM）进行了分析。所提出的固定化方法具有潜在的应用前景，因为它通过包括 pH 值、温度、热处理、储存和酶动力学在内的多种方法证明是稳定的。GOS 和残留酶活性分别为 28.76%和 40.44%。然而，游离酶合成 GOS 的产率仅为 12 小时后 24%。本研究提出在 Cs-MNPs 上固定化 BglA 过程中应用磁铁，以生产 GOS，这是一种在生物稳定和经济高效的方式固定化酶的新方法。
本文重点研究了将海洋栖热菌（Thermotoga maritima）的 BglA 固定在壳聚糖（CS）功能化的 MNPs 上，以研究其进一步提高乳糖转化率和 GOS 产量的可能性。有趣的是，在 Cs-MNPs 上成功实现了 Tm-BglA 的固定化。从酶活性、SDS-PAGE、FT-IR 和 TEM 获得的结果表明，成功获得了 BglA 与 Cs-MNPs 的高结合能力。此外，结合效率计算表明，在第 6 次重复使用结束时，固定化 BglA-Cs-MNPs 保留了其天然活性的 40.44%。此外，成功地采用了磁分离技术，在不显著损失活性的情况下，可将固定化的 BglA 重复用于批式 GOS。
