Oxidation of phenolic compounds catalyzed by immobilized multi-enzyme systems with integrated hydrogen peroxide production

Abstract

Suicide inactivation of peroxidases by hydrogen peroxide is the major deterrent to using such biocatalysts in oxidative processes. This has been successfully addressed by the in situ generation of H 2 O 2 . In this study, we have developed a novel multi-enzyme biocatalyst that has been immobilized on agarose-type carriers to oxidize phenols using oxygen and formic acid as indirect oxidants. This original system couples the in situ production of H 2 O 2 to the phenol oxidation via an enzymatic cascade that involved three different enzymes (formate dehydrogenase, NADH-oxidase and peroxidase) and two different redox cofactors: nicotinamide and flavin derivatives. The cascade reaction only works when enzymes are immobilized on the solid support since soluble enzymes are dramatically inactivated under the reaction conditions. The highest oxidation efficiency was achieved by combining two different solid biocatalysts:(1) formate dehydrogenase and NADH-oxidase co-immobilized onto agarose beads activated with glyoxyl groups and (2) peroxidase immobilized onto agarose beads as well but activated with boronate groups. Unlike conventional peroxidase-mediated oxidations with exogenous H 2 O 2 , this novel system enables the quantitative oxidation of phenol without the addition of H 2 O 2 . Furthermore, this novel system can use a broad range of redox cofactors with similar oxidative effectiveness. Therefore, this novel immobilized trienzyme system removes important pollutants such as hydroxylated aromatic derivatives ( phenol, 4-aminophenol, 2,4-dichloro-phenol or α-naphthol) using formic acid and molecular oxygen as substrates. In addition, this system generates CO 2 as waste beyond the oxidized phenols that can be easily separated from the aqueous solution. The sustainability of this system is supported by an E-factor of 1.3 and an atom economy of 43%. † Electronic supplementary information (ESI) available. See

Immobilizing Systems Biocatalysis for the Selective Oxidation of Glycerol Coupled to In Situ Cofactor Recycling and Hydrogen Peroxide Elimination

“…[10][11][12] The co-immobilization of multienzyme systems can improve: 1) the kinetics of the chemical cascade because of the spatial localization of the different biocatalytic modules, which avoids the accumulation of intermediates and increases the cofactor recycling efficiency [10] and 2) the stability of the biocatalyst because of the in situ elimination of toxic byproducts. [13] Nevertheless, to yield water and O 2 as innocuous products, which avoided the spontaneous DHA oxidation triggered by H 2 O 2 . The co-immobilization of the three enzymes on the same porous carrier allowed the in situ recycling and disproportionation of the redox cofactor and H 2 O 2 , respectively, to produce up to 9.5 mm DHA, which is 18-and 6-fold higher than glycerol dehydrogenase itself and a soluble multienzyme system, respectively.…”

“…We have recently reported two examples in which the optimal design of the immobilization protocols allowed the co-immobilization of several enzymes on the same carrier through their optimal immobilization chemistries, which preserved both the global activity and stability of the multienzyme systems. [10,13] The success of this approach relies on the versatility of the surface chemistry given by the agarose beads that allows us to synthesize carrier surfaces activated with different reactive groups that specifically attach each enzyme through its optimal immobilization chemistry.…”

The E Factor and Process Mass Intensity

“…Striving for better environmental and economic syntheses, chemists had achieved the optimization of many processes, including those of sildenafil citrate (Viagra) [68] and sertraline [69]. Today, innovative waste reduction methods are numerous and diverse, appearing in such areas as catalysis [70,71], catalyst recovery [72], solvent recovery [73] and optimization of work-up conditions [74]. Without the E factor metric, the field of green chemistry would not be where it currently is.…”

Specific immobilization ofd-amino acid oxidase on hematin-functionalized support mimicking multi-enzyme catalysis

“…By measuring the 13 adsorption at 412 nm, the standard curve for SH groups was obtained as shown in Fig. 14 S1. 38 Herein, ELP-DAAO was 24 purified by using inverse transition cycling method using NaCl to trigger the phase 25 groups of hematin, as illustrated in Scheme 1, the CNTs were functionalized with 1 hematin. XPS spectra were acquired using a Thermo VG 6 ESCALAB250 X-ray photoelectron spectrometer, which was operated at the pressure 7 of 2 × 10 -9 Pa using Mg Ka X-ray as the excitation source.…”

“…The covalent attachment of ELP-DAAO onto the24 hematin-functionalized CNTs (hematin-CNTs) was based on the method previously , the catalysis was carried out at a constant temperature of 40 0 C. After17 5 min of the Ellman reaction, the solution was filtered to remove the MPTS-coated 18 CNTs, and the adsorption at 412 nm was measured for the filtrate.…”