Preparation and characterization of polymer-coated mesoporous silica nanoparticles and their application in enzyme immobilization

The increasing application areas of enzymes made essential of production of the enzymes by more economical and efficient techniques. Subtilisin (also known as Alcalase) is a protease that one of the enzymes used for production of the cosmetic goods and detergents. The lack of the operational stability and high cost of the enzymes are resulted in low efficiency in the industrial applications. High temperatures or non-convenient pH values may lower enzyme activity which decrease reaction rate. Since its advantages like flexibility for the process conditions and increased stability of enzyme, immobilization is the key solution to overcome these obstacles [1]. Mesoporous silica nanoparticles (MSNs) have great potential as nanocarriers for biomolecules because of their biocompatibility, low toxicity, large surface area, high porosity, modifiable pore diameter, and adjustable surface properties. Chitosan (CS) has great potential as enzyme immobilization support because it is non-toxic, inexpensive and, user-friendly. The main reason of this great potential is the amino and hydroxyl groups in the chemical structure of chitosan [2]. For immobilization process, chitosan has the disadvantage of having a highly hydrophilic and swelling structure in aqueous media since high water content causes the polymer structure to lose its mechanical strength. To eliminate this obstacle, chitosan has been studied as a component of a polymer blend structure to obtain more strong and efficient matrices [3]. It is known that blending poly(acrylic acid) (PAA) and chitosan provides increased mechanical strength and pH stability. Also, linear polymer chains (such as poly(acrylic acid: PAA) grafted onto the surface of mesoporous silica nanoparticles (MSNs) show that pH-dependent behaviour [4]. Therefore, in the present study, acrylic acid and chitosan have blended onto mesoporous silica nanoparticles via in-situ polymerization technique. About this type of application, to the best of our knowledge, no study was reported in the literature about this type of polymer blend as a coating layer of mesoporous silica nanoparticles. On the other hand, nanobiocatalysis eliminates the activity-lowering effect of traditional immobilization methods by means of the stabilization of enzymes with nanoparticles which provides higher enzyme load per unit weight of the carrier since the nanostructured materials have the advantage of higher surface area. Therefore, subtilisin immobilization onto polymer-coated mesoporous silica nanoparticles (CS-PAA/MSNs) which were obtained via in-situ polymerization was investigated. Then, the optimal conditions for immobilization process were determined at pH 7 and 4 h of immobilization time at temperature of 25oC. Finally, the activity of enzyme immobilized at optimum conditions was compared with that of free enzyme, at different pH and temperature. Beside of this, the operational and storage stability of immobilized enzyme were also determined. The activity of immobilized enzyme reduced from 100% to 40% after 5 repeated use. According to storage stability experiments, the activity of immobilized enzyme reduced from 100% to 60% after 4 weeks of the storage time.