Lipase and Laccase Encapsulated on Zeolite Imidazolate Framework: Enzyme Activity and Stability from Voltammetric Measurements

Abstract

Lipase (Pseudomonas fluorescens) and laccase (Trametates versicolor) were encapsulated on two zeolite imidazolate framework, ZIF‐8 and ZIF‐zni, materials using a one‐pot synthesis‐immobilization method in aqueous solution at room temperature. The synthesized immobilized biocatalysts (Lip@ZIF‐8, Lip@ZIF‐zni, Lac@ZIF‐8, and Lac@ZIF‐zni) were characterized by X‐ray diffraction, scanning electron microscopy, Fourier transform infrared spectroscopy, and thermogravimetric analysis. The enzymatic activities of the four immobilized biocatalysts were characterized via the electrochemical detection of the substrates, p‐nitrophenyl butyrate and 2,2‐azinobis‐3‐ethylbenzthiazoline‐6‐sulfonic acid. For Lip@ZIF‐8 the specific activity was 91.9 U mg−1 and 123.1 U mg−1 for Lip@ZIF‐zni, while for Lac@ZIF‐8 and Lac@ZIF‐zni, the activity was 51 U mg−1 and 163 U mg−1, respectively, confirming that laccase retains a higher level of activity when immobilized onto ZIF‐zni than on ZIF‐8. Lac@ZIF‐8 was the most stable system on storage (15 days at 5 °C), retaining 94 % of initial activity, while Lip@ZIF‐zni biocatalyst had the optimal level of reusability, retaining 40 % of initial activity after five reaction cycles.

Laccase Encapsulation in ZIF‐8 Metal‐Organic Framework Shows Stability Enhancement and Substrate Selectivity

“…Different to the work of Patil et al. and Naseri et al., the reaction time was cut from 20 min and 12 h, respectively, to only 5 min in our work . Therefore, we provide the fastest in situ encapsulation of laccase in ZIF‐8 to this day, resulting in a composite with enhanced solvent and temperature stability and novel size selective specificity against large substrates.…”

A Systematic Study of the Stability of Enzyme/Zeolitic Imidazolate Framework‐8 Composites in Various Biologically Relevant Solutions

“…The conditions used in the synthesis (i. e. in aqueous solution, at mild pH and ambient temperature and for short periods of time) are also compatible with enzymes. A wide range of enzymes have been encapsulated in ZIF-8, including cytochrome c, [5][6][7] horseradish peroxidase, [5,6,[8][9][10] lipase, [5,6,[11][12][13] glucose oxidase (GOx), [9,14] laccase, [12,15,16] catalase, [17] urease, [18] glucoamylase, [19] chloroperoxidase [20] and phenylethanol dehydrogenase. [21] It is also possible to encapsulate multiple enzymes in the same ZIF-8 for cascade reactions.…”

“…[9,21,22] The encapsulated enzymes have been used in colorimetric or electrochemical detections [5,7,14,22] or in repeated cycles of catalysis. [12,13,16] Despite such extensive studies on the encapsulation of enzymes in ZIF-8, the structural stability of the enzyme@ZIF-8 composites has, to our best knowledge, been rarely reported. Structural stability is a critical consideration because the enzymes will leach out unless the integrity of the composites is maintained during their use or storage in solutions.…”

Biomimetic Mineralization of Cytochrome c Improves the Catalytic Efficiency and Confers a Functional Multi-Enzyme Composite

“…Thermal gravimetric analysis (TGA) under nitrogen atmosphere also confirmed the embedment of protein into the Cu(Im)2 (Figure 2b). In the range of less than 200 °C, the mass loss is mainly due to the loss of water and unreacted reagent [14]. The second decomposition stage was observed for all enzyme@Cu(Im)2 samples, 25.39% (Cyt c@Cu(Im)2), 25.76% (GOx-Cyt c@Cu(Im)2) between 200 °C and 370 °C ( Table 1).…”