Design of a 3D Amino-Functionalized Rice Husk Ash Nano-Silica/Chitosan/Alginate Composite as Support for Laccase Immobilization

Abstract

Recycling of agro-industrial waste is one of the major issues addressed in recent years aimed at obtaining products with high added value as a future alternative to traditional ones in the per-spective of a bio-based and circular economy. One of the most produced wastes is rice husk and it is particularly interesting because it is very rich in silica, a material with a high intrinsic value. In the present study, a method to extract silica from rice husk ash (RHA) and to use it as a carrier for the immobilization of laccase from Trametes versicolor was developed. The obtained mesoporous nano-silica was characterized by X-ray diffraction (XRD), ATR-FTIR spectroscopy, Scanning Elec-tron Microscopy (SEM), and Energy Dispersive X-ray spectroscopy (EDS). A nano-silica purity of about 100% was found. Nano-silica was then introduced in a cross-linked chitosan/alginate scaffold to make it more easily recoverable after reuse. To favor laccase immobilization into the composite scaffold, functionalization of the nano-silica with (γ-aminopropyl) triethoxysilane (APTES) was performed. The APTES/RHA nano-silica/chitosan/alginate (ARCA) composite al-lowed to obtain under mild conditions (pH 7, room temperature, 1.5 h reaction time) a robust and easily reusable solid biocatalyst with 3.8 U/g of immobilized enzyme which maintained 50% of its activity after six reuses. The biocatalytic system, tested for syringic acid bioremediation, was able to totally oxidize the contaminant in 24 h.

Keywords: APTES; composite; immobilized laccase; nano-silica; rice-husk ash; syringic acid removal.

Figure 1

Chemical reaction of the nano-silica extraction process from RHA.

Figure 2

EDS analysis (a) and X-ray diffraction graph (b) of rice-husk nano-silica. The X-ray spectrum was obtained using Mo as a target material.

Figure 3

SEM morphological analysis of rice husk (a,b) and rice-husk nano-silica (c,d) at different magnification (1, 2, 2, and 100 KX, respectively).

Figure 4

ATR-FTIR spectra (A) of chitosan/alginate scaffold (CA) (a), RHA nano-silica/chitosan/alginate (RCA) scaffolds with nano-silica:scaffold ratio of 1:1 (w:w) (b) and 2:1 (w:w) (c), and RHA nano-silica (d) and their magnification (B).

Figure 5

Thermogravimetric curves of CA (grey line), RCA 2:1 (w:w) (black line), and RCA 1:1 (w:w) (dark grey line) scaffolds.

Figure 6

EDS spectra (a) and SEM micrograph (b) of APTES/RHA nano-silica/chitosan/alginate scaffold obtained using an APTES:nano-silica ratio of 5:1 (w:w).

Figure 7

ATR-FTIR spectra (A) of APTES (a), RCA scaffold (b), ARCA scaffold (APTES:nano-silica:scaffold ratio of 10:2:1 (w:w:w) (c), and their magnification (B).

Figure 8

Operational stability of the laccase-APTES/RHA nano-silica/chitosan/alginate scaffold. Experimental conditions: 0.18 mM ABTS, 30 °C, 0.1 M citrate—0.2 M PBS, pH 3.

Figure 9

UV-Vis spectra of the syringic acid in the reaction medium at different times of incubation with the ARCA scaffold-optimized biocatalyst. Experimental conditions: starting biocatalyst activity 0.013 U, analysis time 24 h (1440 min), syringic acid concentration 50 mg/L, pH 5.