Abstract
Recombinant Bacillus subtilis lipase was immobilised on magnetic nanoparticles by a facile covalent method and applied to fish oil hydrolysis. High loading of enzyme to the functionalised nanoparticle was achieved with a protein binding efficiency of 95%. Structural changes of the confined enzyme on the surface of the nanoparticles was investigated using transmission electron microscopy and spectroscopic techniques (attenuated total reflectance-Fourier transform infrared and circular dichroism). The biocatalytic potential of immobilised lipase was compared with that of free enzyme and biochemically characterised with respect to different parameters such as pH, temperature, substrate concentrations and substrate specificity. The thermal stability of functionalised nanoparticle bound enzyme was doubled that of free enzyme. Immobilised lipase retained more than 50% of its initial biocatalytic activity after recyclability for twenty cycles. The ability to the immobilised thermostable lipase to concentrate omega-3 fatty acids from fish oil was investigated. Using synthetic substrate, the immobilised enzyme showed 1.5 times higher selectivity for docosahexaenoic acid (DHA), and retained the same degree of selectivity for eicosapentaenoic acid (EPA), when compared to the free enzyme.
Highlights
Enzymes have been used as food additives or in processing raw material in food biotechnology for many years
The magnetic hysteresis loop of the doped nanoparticles showed a zero coercivity, indicative of a superparamagnetic nature, that enable the quick enzyme separation via a magnet as well as high surface area of the nanoparticle being stable dispersal for higher enzyme loading without a magnet [12]
Recombinant lipase 6B sourced from Bacillus subtilis was produced and purified in our laboratory as per our previous optimised protocol [37]
Summary
Enzymes have been used as food additives or in processing raw material in food biotechnology for many years. Enzymes labelled as GRAS (Generally Recognised As Safe), are regulated for use in the US food industry as either ingredients or processing aids [1,2]. The number of these is limited and so there are efforts to discover novel enzymes and optimisation of their inherent properties such as stability, reaction rate and specificity, to enable the development of new green industrial processing methods. Uses of enzymes in food processing strictly follows green chemistry approaches, including avoiding use of costly harsh chemicals in various food processing activities [3,4]. Exploring novel robust enzymes sourced from recombinant/wild microbial strains can provide new industrially useful catalysts for food or industrial processes.
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