Neutral lipase from aqueous solutions on chitosan nano-particles
Introduction
Biopolymers, chitosan is the deacetylated form of chitin and composed of glucosamine, known as (1–4)-2-amino-2-deoxy-d-glucose. Chitosan has three types of reactive functional groups, an amino group as well as both primary and secondary hydroxyl groups at the C-2, C-3 and C-6 positions, respectively [1]. This special structure makes it exhibit chelation with various metal ions [2]. Muzzarelli [3] pointed out that chitosan combines with metal ions by three forms: ion exchange, sorption and chelation. Chitosan has been broadly used for the sorption of heavy metal ions [4], [5], [6]. Further physical and chemical modifications of chitosan have been made to improve the selectivities and the capacities for metals ions [7], [8], [9], [10]. Chitosan is also characterized by weak diffusion properties: long contact times are required to reach equilibrium. Sorption capacity can be controlled by sorbent particle size [8]. Due to the low porosity of chitosan, sorption performances are frequently controlled by mass transfer resistance. To reduce this resistance to mass transfer, chitosan gel beads have been developed to expand the polymer structure and reduce its crystallinity [11], [12], [13]. However, these treatments results in either a decrease of the number of available sorption sites (cross-linking treatment), or the volumetric sorption capacity (the percentage of water in the gel beads can reach 95%) [14]. Controlled drying can increase the volumetric sorption capacity [15]. Another possibility for increasing this volumetric sorption capacity is the grafting of supplementary functional groups [16].
A number of nano-scale inorganic particles offer favourable properties in regard to selective removal of target contaminants. For example, hydrated Fe(III) oxides particles can selectively adsorb dissolved heavy metals like zinc, copper or metalloids like arsenic oxyacids or oxyanions [17]. Amorphous and crystalline Fe(III) hydroxides have long been known to possess selective sorption properties toward arsenites and arsenates [18]. Polymer supported nano-particles have been prepared and used for selective removal of target arsenic compounds and heavy metals [17]. Chitosan nano-particles had been synthesized based on polymer and applied as drug carriers as reported in previous studies [19], [20], [21].
At present, most of chitosan carriers used as immobilized lipase enzyme were micro-sphere, film, micro-capsule, power, etc. [22], [23], [24], [25]. Researches about the sorption properties of chitosan nano-particles for enzyme/protein are rarely reported now. The unique character of nano-particles for their small size, great surface area and quantum size effect could make it exhibit higher capacities for neutral lipase. Chitosan nano-particles used as immobilized enzyme carriers had been studied by our group. The results about immobilized enzyme characters and comparison with other chitosan carriers had been published [26]. The work in this paper aimed to study its sorption kinetics and sorption mechanism for neutral lipase.
Section snippets
Materials
Neutral lipase (5000 U/g) was a commercial enzyme, of food-grade, obtained from Wuxi Enzyme Reagent Factory (Jiangshu, China). Chitosan was provided by Yuhuan Ocean Biochemical Co. Lit (Zhejiang, China), the molecule weight of which was 91,000. TPP abbreviated from sodium polyphosphate was purchased from Dongsheng Chemical Reagent Factory (Zhejiang, China). All other chemicals were of analytical grade and no further purification was required.
Preparation of chitosan nano-particles
Twenty milligram of chitosan was dissolved in 40 ml of
Size and morphology of chitosan nano-particles
The preparation of chitosan nano-particles was based on an ionic gelation interaction between positively charged chitosan and negatively charged tri-polyphosphate at room temperature [28], [29]. Chitosan nano-particles prepared in the experiment exhibited white powder shape, and were insoluble in water, dilute acid, and alkalescent solution. Results were followed in Fig. 1.
Surface functional groups of chitosan nano-particles
The sorption capacity of chitosan could be improved by the substitution of various functional groups, such as amino acid
Conclusion
Chitosan nano-particles have been prepared by ions gelation of chitosan and tri-polyphosphate. FT-IR spectra revealed the functional groups of chitosan nano-particles and the interaction with neutral lipase, the amine and hydroxyl group of nano-particles provided sorption sites for neutral lipase. The experiments results showed that chitosan nano-particles could adsorb neutral lipase from aqueous solution effectively. The experimental data of the sorption equilibrium from neutral lipase
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