Immobilized Enzymes On Modified Halloysite Nanotubes And Their Application In Phenolitic Wastewater Treatment

Phenolic compounds are common organic pollutants existing in wastewater of many chemical manufacturing industries. Enzymes, biological catalysts with high selectivities, have been studied as biocatalysts for wastewater treatment. However, the application of enzymes has been hampered by unstable nature and the resulting requirement of stringent conditions. The previous studies have revealed that enzyme immobilization can be utilized to overcome these problems and optimize various applications.Halloysite is a aluminosilicate clay mineral, which is available in abundance in China as well as other locations around the world. In contrast with other nano-sized materials, naturally occurring HNTs are readily obtainable and much cheaper than other nanoparticles such as carbon nanotubes. Their novel physical and chemical properties derived from the structural versatility provide opportunities for advanced applications in the fields such as electronics, catalysis, biological systems and functional materials. However, application of halloysite as support for enzymes immobilization is currently severely limited by its iorgnic properties, making surface modification of this clay of urgent interest. Three methods ((3-aminopropyl)triethoxysilane (APTES) modification, chitosan modification and self-assembly of HNTs into microgel) have been applied to modify hollaysite nanotubes in order to achieve high enzyme loading and high enzyme stability. The obtained hybrid materials were used as supports for enzymes (horseradish peroxidase and laccase). The application of the immobilized enzymes for the removal of phenolic compounds from aqueous solutions was also studied. With regard to the modification methods, the content could be divided into three parts as follows:(1) Immobilization of horseradish peroxidase on APTES modified halloysite clay nanotubes and application in phenol removalHorseradish peroxidase (HRP) was immobilized on (3-aminopropyl)triethoxysilane (APTES) modified halloysite clay nanotubes to obtain enzyme-clay composites for the treatment of phenol wastewater. Nitrogen adsorption-desorption measurement, fourier-transform infrared spectroscopy, transmission electron microscopy analysis have been employed to elucidate structure of the resulting. HRP were immobilized on the support through covalent bond and the maximum enzyme loading reached as large as 10.4 mg/g. High phenol removal efficiency can be obtained by immobilized HRP over a broader pH range from 6 to 9. With the increase of incubation time, the remain activity of immobilized HRP is about twice that of free HRP at both 45 and 55℃over 100 min incubation. After 1 month storage, the immobilized HRP retained about 61% of its original activity twice that of free HRP. The oxidation of phenol could be achieved within short time with the molar ratio of PEG/phenol and hydrogen peroxide/phenol around 0.01 and 1, respectively. Therefore, HNTs-APTES is suitable support for enzyme immobilization.(2) Halloysite-chitosan (HNTs-CTS) hybrid-nanotubes for horseradish peroxidase immobilization and applications in phenol removalChitosan-halloysite hybrid-nanotubes were based in the assembling of chitosan to halloysite, which is a natural aluminosilicate showing a network nanostructure. Nitrogen adsorption-desorption measurement, fourier-transform infrared spectroscopy and scan electron microscopy analysis have been employed to elucidate structure of the resulting. The results indicated that the nanocomposites were composed of chitosan-halloysite hybrid-nanotubes which exhbited microfibrous texture. As-prepared hierarchically structure showed excellent capacity for horseradish peroxidase (HRP) immobilization through cross-linking by glutaraldehyde. The maximum enzyme loading reached as large as 21.5 mg/g, much higher than that of raw halloysite. After 35 days storage, the immobilized HRP did not undergo any activity loss. And higher activity over a broad pH range than free HRP open the door for practical applications. Furthermore, phenol removal efficiency by the immobilized HRP was explored. The result showed the immobilized HRP exhibited overall high removal efficiency for phenol wastewater. Therefore, HNTs-CTS is suitable support for enzyme immobilization.(3) Mesoporous halloysite-chitosan microspheres as matrix for laccase immobilization and application in 2,4-dichlorophenol removalMesoporous halloysite-chitosan microspheres have been prepared facilely at room temperature by the assembly of chitosan modified halloysite nanotubes. Fourier-transform infrared spectroscopy, scan electron microscopy, laser particle size analyzer analysis have been employed to elucidate structure of the resulting. These new microspheres were hybrid-gel consisted of inorganic nanotubes and chitosan. The diameter of the microsphere dispersed in aqueous solution was in the range between 10-40μm. Laccase was immobilized onto the microspheres surface through physical adsorption. The maximum enzyme loading reached as large as 118 mg/g. And the immobilized laccase activity could achieve as large as 767 U/g. The thermal and the storage stability of immobilized laccases are higher than the free ones. Furthermore,2,4-dichlorophenol compound removal efficiency by the immobilized laccase was explored. The result (more than 80% after 6 h) showed the immobilized laccase exhibited overall high removal efficiency. Therefore, the hybrid porous microsphere, obviously improving the stabilities of the laccase adsorbed on their surface, was a promising support for enzyme immobilization.In this paper, HNTs were modified through various methods to obtain hybrid nanotubes, which hold the advantages of both inorganic and organic materials. Thanks to their biocompatible and non-toxic property, the hybrid-materials offer thus a new family for enzyme support, biocompatible delivery system, porous bio-support material, catalyst support, etc.