Enhanced Performance of Immobilized Xylanase/Filter Paper-ase on a Magnetic Chitosan Support

Abstract

Enzyme immobilization on different supports has emerged as an efficient and cost-effective tool to improve their stability and reuse capacity. This work aimed to produce a stable immobilized multienzymatic system of xylanase and filter paper-ase (FPase) onto magnetic chitosan using genipin as a cross-linking agent and to evaluate its biochemical properties and reuse capacity. A mixture of chitosan magnetic nanoparticles, xylanase, and FPase was covalently bonded using genipin. Immobilization yield and efficiency were quantified. The activity of free and immobilized enzymes was quantified at different values of pH, temperature, substrate concentration (Km and Vmax), and reuse cycles. The immobilization yield, immobilization efficiency, and activity recovery were 145.3% ± 3.06%, 14.8% ± 0.81%, and 21.5% ± 0.72%, respectively, measured as the total hydrolytic activity. Immobilization confers resistance to acidic/basic conditions and thermal stability compared to the free form. Immobilization improved 3.5-fold and 78-fold the catalytic efficiency (Kcat/Km) of the xylanase and filter paper-ase activities, while immobilized xylanase and FPase could be reused for 34 min and 43 min, respectively. Cross-linking significantly improved the biochemical properties of immobilized enzymes, combined with their simplicity of reuse due to the paramagnetic property of the support. Multienzyme immobilization technology is an important issue for industrial applications.

Enzyme Storage and Recycling: Nanoassemblies of α-Amylase and Xylanase Immobilized on Biomimetic Magnetic Nanoparticles

“… 28 This method is well established in the area, and many other authors have used the DNS reaction to evaluate the activity of immobilized glycoside hydrolases. 29 − 31 …”

Magnetic Nanoparticle-Containing Supports as Carriers of Immobilized Enzymes: Key Factors Influencing the Biocatalyst Performance

“…The magnetic recovery of immobilized enzymes is normally realized by tethering enzymes to magnetic NPs via some linkers such as polymers, difunctional molecules, etc. [ 31 , 32 , 33 , 34 , 35 , 36 , 37 ]. To create an appreciable magnetic moment for a successful separation, magnetic NPs are often organized into clusters [ 34 , 38 ], imbedded in gels [ 36 ], other polymer or inorganic matrices [ 13 , 39 , 40 , 41 ], or self-assembled into larger structures [ 42 , 43 ].…”

“…Furthermore, it modified the secondary lipase structure and allowed for the exposure of the enzyme active site. Crosslinking of the chitosan-magnetic NP support with immobilized xylanase and filter paper-ase using genipin allowed for a significant enhancement of activity compared to a non-crosslinked biocatalyst [ 31 ]. This was attributed to a higher local concentration of the substrate in the confined space, resulting in a boosted number of interactions between the substrates and immobilized enzymes.…”

“…The magnetic recovery of immobilized enzymes is normally realized by tethering enzymes to magnetic NPs via some linkers such as polymers, difunctional molecules, etc. [31][32][33][34][35][36][37].…”

“…Clustering of the NPs surrounded by the thermoresponsive polymer allows for easy magnetic separation. [31][32][33][34][35][36][37]. To create an appreciable magnetic moment for a successful s NPs are often organized into clusters [34,38], imbedded in gels [36], o organic matrices [13,[39][40][41], or self-assembled into larger structures interparticle interactions and increases magnetic attraction between m external magnetic field.…”