A novel method for preparation of MNP@CS-tethered coenzyme for coupled oxidoreductase system

• A novel MNP@CS with azelaic acid spacer and active amine group was prepared.
• The NADH and NAD+ could be easily covalently linked to particle with high loading.
• The prepared nanoparticle tethered NAD(H) could be regenerated in coupled oxidoreductase system.

The immobilized cofactor NAD(H) is easily recovered from the reaction bulk, which is essential for repeated use of NAD(H) in the bioprocess catalyzed by NAD(H)-dependent oxidoreductase. Here, a magnetic nanoparticle platform was designed to immobilize both of the NADH and the NAD+. The design was based on chitosan-coated magnetic nanoparticles (MNP@CS) which was activated by the EDC/NHS with the aid of azelaic acid as spacer. Interestingly, the succinimide group at the end of spacer arm catalyzed direct coupling of a carboxyl-terminal to the 6-amino group of the adenine residue of NAD(H). Our results indicated that 150 μmol NADH and 50 μmol NAD+ was effectively attached to 1 g MNP@CS at 25 °C in 120 min and the prepared MNP@CS–NAD(H) showed good activity according to the coupling reaction of benzyl alcohol and acetaldehyde catalyzed by alcohol dehydrogenase.