Abstract Cross-linked enzyme aggregates (CLEAs) are considered as an effective tool for the immobilization of enzyme. The ionic cross-linking agent-sodium tripolyphosphate (TPP) was first used in preparing CLEAs. Aspergillus niger lipase was precipitated with ammonium sulfate and further cross-linked by TPP. The factors including enzyme concentration, pH of cross-linking medium, TPP dosage and cross-linking time were optimized. Maximum recovery activity (99.5 ± 0.634 %) and cross-linking yield (88.4 ± 0.46 %) can be obtained under the optimal process conditions, which can illustrate TPP had little effect on enzyme activity. CLEAs showed improved activity over broad pH and temperature range compared to the free enzyme. The thermal stability was obviously improved compared to free enzyme under the optimal temperature (40℃) and the half-life was 7.5-fold higher than that of free enzyme. Moreover, scanning electron microscopy (SEM) revealed that CLEAs had a cavity with porous structure and the particle size was 249 ± 3.98 nm. X-ray diffraction (XRD) showed the crystallinity of the CLEAs decreased. The changes in secondary structures of CLEAs revealed the increment in conformational rigidity. Such results suggested that the CLEAs has ideal application prospects.

中文翻译：

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脂肪酶与三聚磷酸钠交联制备纳米酶聚集体

摘要 交联酶聚集体（CLEAs）被认为是一种有效的酶固定工具。离子交联剂-三聚磷酸钠 (TPP) 首先用于制备 CLEA。黑曲霉脂肪酶用硫酸铵沉淀，并通过 TPP 进一步交联。对酶浓度、交联介质pH值、TPP用量和交联时间等因素进行了优化。在最佳工艺条件下可获得最大回收活性（99.5±0.634%）和交联产率（88.4±0.46%），说明TPP对酶活性影响不大。与游离酶相比，CLEA 在较宽的 pH 值和温度范围内显示出更高的活性。在最适温度（40℃）下，热稳定性较游离酶有明显提高，半衰期为7。比游离酶高5倍。此外，扫描电子显微镜 (SEM) 显示 CLEA 具有多孔结构的空腔，粒径为 249 ± 3.98 nm。X 射线衍射 (XRD) 显示 CLEA 的结晶度降低。CLEA 二级结构的变化揭示了构象刚性的增加。这样的结果表明CLEAs具有理想的应用前景。