Abstract
CNTs need to be dispersed in aqueous solution for their successful use, and most methods to disperse CNTs rely on tedious and time-consuming acid-based oxidation. Here, we report the simple dispersion of intact multi-walled carbon nanotubes (CNTs) by adding them directly into an aqueous solution of glucose oxidase (GOx), resulting in simultaneous CNT dispersion and facile enzyme immobilization through sequential enzyme adsorption, precipitation, and crosslinking (EAPC). The EAPC achieved high enzyme loading and stability because of crosslinked enzyme coatings on intact CNTs, while obviating the chemical pretreatment that can seriously damage the electron conductivity of CNTs. EAPC-driven GOx activity was 4.5- and 11-times higher than those of covalently-attached GOx (CA) on acid-treated CNTs and simply-adsorbed GOx (ADS) on intact CNTs, respectively. EAPC showed no decrease of GOx activity for 270 days. EAPC was employed to prepare the enzyme anodes for biofuel cells, and the EAPC anode produced 7.5-times higher power output than the CA anode. Even with a higher amount of bound non-conductive enzymes, the EAPC anode showed 1.7-fold higher electron transfer rate than the CA anode. The EAPC on intact CNTs can improve enzyme loading and stability with key routes of improved electron transfer in various biosensing and bioelectronics devices.
Highlights
Carbon nanotubes (CNTs) need to be dispersed in aqueous solution for their successful use, and most methods to disperse CNTs rely on tedious and time-consuming acid-based oxidation
According to 3D structural analysis, glucose oxidase (GOx) has a hydrophobic patch on its surface that can interact with the hydrophobic side wall of intact CNTs, resulting in facile GOx adsorption onto the surface of CNTs (Fig. S1)
Hydrophilic interactions between water molecules and hydrophilic side chains on the surface of GOx lead to highly-dispersed intact CNTs in aqueous enzyme solution (Figs S1 and 1b)
Summary
The maximum power density ratio between EAPC-based and CA-based biofuel cells (7.2) is higher than their enzyme activity ratio (4.5) (Table 1) This difference can be explained by the difference in their electron transfer rate constants. In the present work we demonstrated the successful dispersion of CNTs in GOx solutions with simultaneous adsorption of the enzyme onto the CNTs followed by precipitation and crosslinking Such EAPCs of GOx resulted in highly active and stable form of immobilized enzyme preparations at high enzyme loading. The EAPC of GOx showed improved electron conductivity in a biofuel cell operation when compared to conventional enzyme immobilization of covalent enzyme attachment using acid-treated and oxidized CNTs. The versatile uses of CNTs are well established and still growing, and it is anticipated that the simple CNT dispersion in enzyme solutions can find additional applications of biomolecules in the development of biosensing and bioelectronics devices
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