Improved catalytic performance of carrier-free immobilized lipase by advanced cross-linked enzyme aggregates technology

Abstract

The cross-linked enzyme aggregates (CLEAs) are one of the technologies that quickly immobilize the enzyme without a carrier. This carrier-free immobilization method has the advantages of simple operation, high reusability and low cost. In this study, ionic liquid with amino group (1-aminopropyl-3methylimidazole bromide IL) was used as the novel functional surface molecule to modify industrialized lipase (Candida rugosa lipase, CRL). The enzymatic properties of the prepared CRL-FIL-CLEAs were investigated. The activity of CRL-FIL-CLEAs (5.51 U/mg protein) was 1.9 times higher than that of CRL-CLEAs without surface modi cation (2.86 U/mg protein). After incubation at 60℃ for 50 min, CRL-FIL-CLEAs still maintained 61% of its initial activity, while the value for CRL-CLEAs was only 22%. After repeated use for ve times, compared with the 22% residual activity of CRL-CLEAs, the value of CRL-FIL-CLEAs was 51%. Further kinetic analysis indicated that the Km values for CRL-FIL-CLEAs and CRL-CLEAs were 4.80 mM and 8.06 mM, respectively, which was inferred that the a nity to substrate was increased after modi cation. Based on the above results, it was indicated that this method provided a new idea for the effective synthesis of immobilized enzyme. HighlightsNovel immobilized lipase was prepared with carrier-free immobilization method.The aggregates of ionic liquid and lipase proved to be an e cient catalyst.The activity and stability of carrier-free immobilized lipase were improved.The activity of the novel aggregates reached 1.9 times of conventional lipase.

Easy and Versatile Technique for the Preparation of Stable and Active Lipase-Based CLEA-like Copolymers by Using Two Homofunctional Cross-Linking Agents: Application to the Preparation of Enantiopure Ibuprofen

“…As commented in the introduction, the most used carrier-free immobilization method is the preparation of CLEAs; a plethora of publications and reviews have described CLEAs with multiple enzymes [12,42,57,58,67]. This methodology involves the precipitation of the enzyme molecules (with or without other reagents) and the ulterior crosslinking using mainly (but sometimes not only) GA [51].…”

“…Compared with the other latest examples of CLEAs with CRL found in the literature, our CLEA-LC seems to be advantageous; for instance, Jiaojiao et al [67] recently reported CRL-CLEAs using an expensive amine-containing ionic liquid (1-aminopropyl-3methylimidazole bromide, FIL) as the functional surface molecule with which to modify CRL. The activity of this CLEA was tested only with a chromogenic standard substrate without testing it on any racemic substrate to check its stereoselection capability.…”

“…As stated before, other CRL-CLEAs exhibited poorer behaviour, displaying low reusability of approximately 40% residual activity after five cycles [67]. On the other hand, Sampath et al [99] recently reported the preparation of a CLEA created using GA to cross-link CRL molecules, using bovine serum albumin (BSA) and polyethileneamine (PEI) as the co-feeders.…”

Lipases: Sources of Acquisition, Ways of Production, and Recent Applications

“…An advantage of this technique is its ability to make the enzyme more pH stable. JiaoJiao et al [178] showed that lipase from Candida rugosa could be immobilized by the CLEAs methodology and its behavior improved as the enzyme became more stable at a particular pH compared to the free enzyme.…”