A novel polyurethane (PU) support with and without copper oxide nanoparticles (nCuO) was employed for immobilization of two chitinolytic enzymes, chitinase and N-acetyl β glucosaminidase (NAGase) to yield PU/nCuO/chitinase/NAGase and PU/chitinase/NAGase conjugates respectively. The surface morphologies, topologies and bonded interactions between different components of the immobilized enzyme conjugates were characterized using Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM) and Fourier Transform Infrared Spectroscopy (FTIR) respectively. High conjugation yield of 0.811±0.10mg/cm2 with 97.66±0.50% retention of specific activity of enzymes on PU/nCuO support vis-a-vis a conjugation yield of 0.531±0.50mg/cm2 with 75.23±0.60% retention activity on PU support was achieved. Additionally, increased pH and temperature tolerance, better kinetic parameters and enhanced stabilities of PU/nCuO/chitinase/NAGase conjugates established their superiority over the PU/chitinase/NAGase conjugates. However, both the conjugates were successfully employed for the determination of chitin contents in the apparently healthy stored rice grains. The methods were tested on analytical parameters such as linearity, limit of detection, the coefficient of variation and reproducibility of the externally added chitin, which were again found to be superior for PU/nCuO/chitinase/NAGase conjugates.
Keywords: Chitin determination; Chitinase; Copper oxide nanoparticles; Immobilization; N-acetyl β glucosaminidase; Polyurethane.
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