Novel polydopamine imprinting layers coated magnetic carbon nanotubes for specific separation of lysozyme from egg white

Highlights

• Novel imprinted magnetic carbon nanotubes for specific recognition of lysozyme.

• 10 nm of imprinting layer enabled the template access the binding cavities easily.

• Fast kinetic and high affinity were achieved.

• Successfully selective removal of Lyz from real egg white.

• A potential nanomaterial of electrochemical sensor for recognizing lysozyme.

Abstract

Novel core–shell nanocomposites, consisting of magnetic carbon nanotubes (MCNTs) core surrounded by a thin polydopamine (PDA) imprinting shell for specific recognition of lysozyme (Lyz), were fabricated for the first time. The obtained products were characterized and the results showed that the PDA layer was successfully attached onto the surface of MCNTs and the corresponding thickness of imprinting layer was just about 10 nm which could enable the template access the recognition cavities easily. The polymerization conditions and adsorption performance of the resultant nanomaterials were investigated in detail. The results indicated that the obtained imprinted polymers showed fast kinetic and high affinity towards Lyz and could be used to specifically separate Lyz from real egg white. In addition, the prepared materials had excellent stability and no obvious deterioration after five adsorption–regeneration cycles. Easy preparation, rapid separation, high binding capacity, and satisfactory selectivity for the template protein make this polymer attractive in biotechnology and biosensors.

Introduction

Lysozyme (Lyz) is a naturally occurring enzyme present in vegetables, milk, tears, and egg white, which has specific hydrolytic activity against bacterial cell walls and is nontoxic to humans. In addition, Lyz is also reported to have antiviral and anti-inflammatory activities [1], [2]. With the development of research for Lyz, it has become a commercially important enzyme, a potent antibacterial agent in food technology, and a drug for treatment of ulcers, infection, and cancer [3], [4]. To date, the commercial Lyz is obtained through isolation and purification from food by using ion exchange, chromatographic separation, and membrane treatment technologies [5], [6], [7]. However, low binding capacity, slow intraparticle diffusion of biomolecules, poor mechanical strength, and non-specificity limit the wide application of these methods. Therefore, more stable, specific, and economical strategies for the production of Lyz are desired.

Molecularly imprinted polymers (MIPs), man-made polymers with predetermined selectivity towards a given analyte or a group of structurally related species, have been demonstrated as one of the most potential materials in separation field because of its simplicity, special selectivity, chemical, mechanical, and thermal stability, and low cost of fabrication [8], [9]. Over the past decades, the MIPs have been successfully applied in small molecules, but imprinting of biomacromolecules still faces serious challenges due to the large molecular size, water-solubility, complex, and conformational flexibility of proteins [10], [11]. Some approaches are developed to deal with these issues, among which surface imprinting technology is considered to be an efficient way to build recognition sites on the surface of supporting materials for solving the mass transfer difficulty induced by large molecular size of proteins [12], [13].

Carbon nanotubes (CNTs), employed as supporting matrix to deal with protein adsorption or isolation, have gained much attention, especially in the field of surface imprinting [14], [15], [16], [17], [18]. Yuan et al. utilized multiwall CNTs as carrier and ionic liquid as functional monomer to prepare Lyz-imprinted polymers for separating Lyz from egg white [19]. Zhang et al. prepared bovine serum albumin-imprinted polymers based on the multiwall CNTs to successfully separate the target protein from a real blood sample [20]. However, the separation process of these MIPs from the solutions is tedious when CNTs were adopted as the supporter. It is generally known that Fe₃O₄ magnetic nanoparticles (MNPs) could be isolated easily by an external magnetic field owing to their super-magnetism. Therefore, the magnetic carbon nanotubes (MCNTs) combining the merits of CNTs and MNPs has faster separation speed than CNTs and larger surface area than MNPs, which is an appealing material for surface imprinting.

The functional monomer plays a vital role in the formation of imprinted polymers through giving assistance to creating the specific recognition cavities in an optimal position. Considering the solubility and complicacy of protein, dopamine (DA) with multifunctional groups, hydrophilicity, and biocompatibility is an ideal monomer for imprinting proteins [21]. Another distinct advantage of DA is excellent film-forming ability onto different kinds of materials by self-polymerization in weak base solution at room temperature [22], [23]. Some successful works using DA as the monomer and cross-linker for surface protein imprinting have been published up to now [24], [25], [26].

Herein, we fabricated core–shell nanocomposites consisting of a MCNTs core surrounded by a thin polydopamine (PDA) imprinting layer for specific recognition of Lyz for the first time. Transmission electron microscope (TEM), Fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD), and vibrating sample magnetometry (VSM) were used to characterize the synthesized products. The polymerization conditions and adsorption performance of the resultant nanomaterials were investigated in detail. The results indicated that the obtained imprinted polymers showed fast kinetic and high affinity towards Lyz and could be used to specifically separate Lyz from real egg white. The differences in binding capability of adopting MCNTs or MNPs as carrier were also evaluated to further demonstrate the superiority of the approach developed in this work.