Colorimetric quantification of galactose using a nanostructured multi-catalyst system entrapping galactose oxidase and magnetic nanoparticles as peroxidase mimetics

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Abstract
A colorimetric method for quantification of galactose, which utilizes a nanostructured multi-catalyst system consisting of Fe3O4 magnetic nanoparticles (MNPs) and galactose oxidase (Gal Ox) simultaneously entrapped in large pore sized mesocellular silica, is described. Gal Ox, immobilized in a silica matrix, promotes reaction of galactose to generate H2O2 that subsequently activates MNPs in silica mesopores to convert a colorimetric substrate into a colored product. By using this colorimetric method, galactose can be specifically detected. Along with excellent reusability via application of simple magnetic capturing, enhanced operational stability was achieved by employing a cross-linked enzyme aggregate (CLEA) method for Gal Ox immobilization. This protocol leads to effective prevention of enzyme leaching from the pores of mesocellular silica. The analytical utility of the new colorimetric biosensor was demonstrated by its use in diagnosing galactosemia, a genetic metabolic disorder characterized by the inability to utilize galactose, through analysis of clinical dried blood spot specimens. A microscale well-plate format was employed that possesses a multiplexing capability. The multi-catalyst system entrapping Gal Ox and MNPs represents a new approach for rapid, convenient, and cost-effective quantification of galactose in human blood and it holds promise as an alternative method for galactosemia diagnosis, replacing the laborious procedures that are currently in use.

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