چكيده به لاتين
Enzyme immobilization as a way to overcome challenges of using enzymes in industrial scale has been considered. In this research, the effects of coating and that of activation level on surface of magnetic ferric oxide nanoparticles (MNPs) on immobilization of amylase were investigated. Two MNPs with two different coating conditions -uncoated and silica coated- were subject to surface treatment using the amine groups and then activated with different concentrations of glutaraldehyde (GA). The GA activated carriers can take part in enzyme immobilization process through various physical, e.g. hydrophobic adsorption and ion exchange and chemical, e.g. covalent, adsorption methods. The ionic strength of the medium would have an impact on determination of the predominant method; whether it is dominated with a physical or a chemical mechanism. Therefore, the immobilization level and activity of the immobilized enzyme were studied for three low, moderate, and high ionic strengths of buffer solution. Furthermore, the type of the bond developed between enzyme and carrier was identified using amount of the immobilized enzyme release under different environments as well as adsorption kinetic studies and FTIR analysis. The results suggest that for most cases, the mechanisms for amylase immobilization on MNPs activated with GA occur in the following order: First physical adsorption occurs followed by forming covalent bond between enzyme and carrier. The effects of different operational conditions of α-amylase covalent immobilization on magnetic nanoparticles (MNPs), such as initial enzyme concentration, glutaraldehyde (GA) concentration, pH, ionic strength, were investigated using central composite design (CCD). Moreover, the two responses of biocatalyst activity and amount of immobilized enzyme were simultaneously studied by using Derringer’s desirability function. The optimum amount and activity of immobilized enzyme were determined as 24.83% and 556.41 mg/gMNP at 999.86 ppm initial enzyme concentration, solution pH of 4.6, 0.59% GA concentration, 99.99 mM ionic strength and 4 h process time. Kinetic parameters and stability of free and immobilized enzyme were studied in laboratory and industrial operational conditions. The results showed the significant enhancement in the performance of the immobilized enzyme with respect to the free enzyme in two mentioned conditions. The storage stability and reusability of immobilized biocatalyst were obtained about 50 and 40% of the initial activity after 12 days and 6 cycle uses, respectively in low concentration of substrate. Moreover, Immobilized enzyme showed only 40 % performance decrease in the case of industrial conditions.
