Imobilização da endoxilanase de Thermomyces Ianuginosus pc7s1t em quitosana magnética: caracterização e produção de xilooligossacarídeos a partir de resíduos agrícolas

Abstract

Xylanases are hydrolytic enzymes with great biotechnological potential and applications in several sectors, such as the food industry, animal feed, beverages, paper, biofuels, the production of xylooligosaccharides, and the bioconversion of agricultural waste. Their large-scale or industrial application is contingent upon parameters such as thermal stability, adaptability to various reaction conditions, and the feasibility of recovery and reuse post-process. Enzyme immobilization serves as a strategy to enhance biocatalysts by securing the enzyme within a matrix that imparts advantageous properties for its application. Chitosan is a non-toxic, low cost, renewable biopolymer with high protein affinity that stands out as a material for immobilization, allowing different mechanisms such as covalent binding and entrapment. This study aimed to characterize the endoxylanase from the filamentous fungus Thermomyces lanuginosus PC7S1T, immobilized in magnetic chitosan, and to explore its applicability in the production of xylooligosaccharides. The immobilization of endoxylanase on this support resulted in a yield of 90.65%, an efficiency of 73.66%, and a recovered activity of 66.77%. The enzyme demonstrated exceptional storage stability, maintaining 94% of its activity after 50 days at 4°C. The reuse investigation indicated that the immobilized enzyme preserved 54% of its relative activity over 10 cycles. At temperatures of 70°C and 75°C, the immobilized endoxylanase exhibited enhanced stability relative to its free form, and at pH 6.5, it retained 100% of its activity following 5 hours of incubation. The presence of Mn²⁺ ions resulted in enhanced activity in both the free (22.5%) and immobilized (25.3%) forms; whereas, exposure to ethanol and Triton X-100 led to a decrease in activity of 28.1% and 23.6% for the immobilized enzyme, respectively. The saccharification of hemicelluloses derived from agricultural residues (corn straw and cob, sugarcane bagasse, and sorghum biomass) demonstrated that the immobilized enzyme enhanced the production of xylooligosaccharides, particularly xylotriose (X3) and xylotetraose (X4), though with a slightly lower overall yield compared to the free enzyme. Scanning electron imaging demonstrated considerable depolymerization of these hemicellulose fibers by the immobilized enzyme. Toxicity evaluations revealed that the XOS generated by the immobilized enzyme displayed negligible toxicity to Artemia salina. The endoxylanase immobilized in magnetic chitosan exhibits novel and advantageous properties that satisfy the requirements for resilient biocatalysts appropriate for industrial applications, particularly in the degradation of agricultural residues and the synthesis of xylooligosaccharides with potential prebiotic uses.