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Surface-functionalized magnetic nanoparticles are widely used in various fields of biotechnology. In this study, beta-cyclodextrin-conjugated magnetic nanoparticles (CD-APES-MNPs) are synthesized and the use of CD-APES-MNPs as a solid-phase artificial chaperone to assist protein refolding in vitro is demonstrated using carbonic anhydrase bovine (CA) as model protein. CD-APES-MNPs are fabricated by grafting mono-tosyl-beta-cyclodextrin (Ts-beta-CD) onto 3-aminopropyltriethoxysilane (APES)-modified magnetic nanoparticles (APES-MNPs). Results obtained from transmission electron microscopy (TEM) and vibrating sample magnetometery (VSM) show that the synthesized magnetic nanoparticles are superparamagnetic with a mean diameter of 11.5 nm. The beta-CD grafting is confirmed by Fourier transform infrared spectroscopy (FTIR) and elemental analysis. The amount of beta-CD grafted on the APES-MNPs is found to be 0.042 mmol g(-1) from elemental analysis. Our refolding results show that a maximum of 85% CA refolding yield can be achieved using these beta-CD-conjugated magnetic nanoparticles which is at the same level as that using liquid-phase artificial chaperone-assisted refolding. In addition, the secondary and tertiary structures of the refolded CA are the same as those of native protein under optimal conditions. These results indicate that CD-APES-MNPs are suitable and efficient stripping agents for solid-phase artificial chaperone-assisted refolding due to easier and faster separation of these nanoparticles from the refolded samples and also due to recycling of the stripping agents.

Adsorption of globular protein, lysozyme, on thermosensitive poly(N-isopropylacrylamide) coated nanomagnetic particles was studied at different temperatures and pHs. It was observed that a maximum amount of lysozyme was adsorbed at a temperature above the lower critical solution temperature (LCST) (32 degrees C ) of the polymer and at the isoelectric point (pI=11) of lysozyme. Desorption was carried out using either NaH2PO4 (pH 4) or NaSCN (pH 6) as the desorbing agents. Conformational changes in lysozyme on desorption from nanomagnetic particles was studied by circular dichroism and intrinsic fluorescence spectroscopy. Lysozyme desorbed by NaH2PO4 showed very little conformational changes while lysozyme desorbed by NaSCN showed significant conformational changes, and 87% enzymatic activity was retained in the desorbed enzyme for desorption by NaH2PO4.

Thermosensitive polymer coated nanomagnetic adsorbents were synthesized by seed polymerization using surface modified nanomagnetic particles as the seeds. The Fe3O4 nanomagnetic particles were prepared by chemical precipitation of Fe2+ and Fe3+ salts in the ratio of 1:2 under alkaline and inert condition. The surface of these particles was modified by surfactants to achieve stability against agglomeration. These stable particles were then polymerized using N-isopropylacrylamide (NIPAM) as the main monomer, methylene-bis-acrylamide as the crosslinker and potassium per sulfate as the initiator. The thermosensitive adsorbents were characterized by using transmission electron micrography (TEM) and vibrating sample magnetometer (VSM). TEM showed that the particle remained discrete with a mean diameter of 12 nm. Magnetic measurements revealed that the particles are superparamagnetic only with a decrease of magnetism after binding with the polymer due to the increase in surface spin disorientation. Pure Fe3O4 spinel structure of these nanoparticles was indicated by the X-ray diffraction (XRD) patterns. The polymerization of NIPAM with the surface modified nanomagnetic particles was confirmed by Fourier transform spectroscopy (FTIR), Thermogravimetric analysis (TGA) and X-ray photoelectron spectroscopy (XPS). In addition, the adsorption/desorption of BSA molecule on these thermosensitive nanoparticles was investigated as a function of temperature. More than 60% desorption efficiency was achieved under appropriate condition.

Adsorption and desorption behavior of Bovine Serum Albumin (BSA) on surface-modified magnetic nanoparticles covered with thermosensitive polymer (PNIPAM) was investigated as a function of temperature, pH, and ionic strength. Functionalization of surface-modified magnetic particles was performed by seed polymerization using N-isopropylacrylamide (PNIPAM) as the main monomer. Characterization of these particles was carried out using transmission electron micrography (TEM), and vibrating sample magnetometry (VSM). The adsorption results exhibited both pH and temperature sensitivity. The results showed that the temperature effect on adsorption/desorption behavior was mainly dependent on the properties of the particles' surface. The effect of pH was also investigated and it was observed that a smaller amount of protein was adsorbed at higher pH because of the electrostatic repulsive force between protein molecules and latex particles. The maximum amount of protein was adsorbed near the isoelectric point of BSA. Desorption results showed that more protein was desorbed when adsorption was done at lower temperatures and desorption efficiency was found to be higher than 80%.

Adsorptions of bovine serum albumin (BSA) on nano-sized magnetic particles with and without the presence of carbodiimide were studied. Desorption of BSA from magnetic particles were carried out in either NaOH or Na 2 HPO 4 solutions. The structures of native BSA, adsorbed BSA on magnetic particles, and desorbed BSA were studied by several methods, circular dichroism (CD), fluorescence spectroscopy and differential scanning calorimetry (DSC). The magnitude of conformational changes of protein was determined by calculating the α-helix content from the circular dichroism (CD) spectra and by evaluating fluorescence spectrum and DSC thermograms. Adsorbed BSA on magnetic particles shows no thermal transition with respect to the native BSA. The structural change of BSA when desorbed by Na 2 HPO 4 solution is much smaller in comparison to that when desorbed by NaOH solution. Hence, this indicates that BSA could be desorbed from nano-sized magnetic particles using Na 2 HPO 4 without much conformational change.

Chiral resolution aromatic amino acids, dl-tryptophan (dl-Trp), dl-phenylalanine (dl-Phe), dl-tyrosine (dl-Tyr) from phosphate buffer solution was achieved in present study employing the concept of selective adsorption by surface functionalized magnetic nanoparticles (MNPs). Surfaces of magnetic nanoparticles were functionalized with silica and carboxymethyl-β-cyclodextrin (CMCD) to investigate their adsorption resolution characteristics. Resolution of enantiomers from racemic mixture was quantified in terms of enantiomeric excess using chromatographic method. The MNPs selectively adsorbed l-enantiomers of dl-Trp, dl-Phe, and dl-Tyr from racemic mixture and enantiomeric excesses (e.e.) were determined as 94%, 73% and 58%, respectively. FTIR studies demonstrated that hydrophobic portion of enantiomer penetrated into hydrophobic cavity of cyclodextrin molecules to form inclusion complex. Furthermore, adsorption site was explored using XPS and it was revealed that amino group at chiral center of the amino acid molecule formed hydrogen bond with secondary hydroxyl group of CMCD molecule and favorability of hydrogen bond formation resulted in selective adsorption of l-enantiomer. Finally, stability constant (K) and Gibbs free energy change (−ΔG°) for inclusion complexation of CMCD with l-/d-enantiomers of amino acids were determined using spectroflurometry in aqueous buffer solution. Higher binding constants were obtained for inclusion complexation of CMCD with l-enantiomers compared to d-enantiomers which stimulated enantioselective properties of CMCD functionalized magnetite silica nanoparticles.

A novel magnetic nanoadsorbent (CMCD-APTS-MNPs) containing the superparamagnetic and molecular recognition properties was synthesized by grafting carboxymethyl-β-cyclodextrin (CM-β-CD) on 3-aminopropyltriethoxysile (APTS) modified Fe(3)O(4) nanoparticles. The feasibility of using CMCD-APTS-MNPs as magnetic nanoadsorbent for selective adsorption of adenosine (A) and guanosine (G) based on inclusion and molecular recognition was demonstrated. The as-synthesized magnetic nanoparticles were characterized by TEM, FTIR and TGA analyses. The effects of pH and initial nucleoside concentrations on the adsorption behavior were studied. The complexation of CMCD-APTS-MNPs with both nucleosides was found to follow the Langmuir adsorption isotherm. The CMCD-APTS-MNPs showed a higher adsorption ability and selectivity for G than A under identical experimental conditions, which results from the ability of selective binding and recognition of the immobilized CM-β-CD towards G. The driving force of the separation between G and A is through the different weak interaction with grafted CM-β-CD, i.e., hydrogen bond interaction, which is evidenced by different inclusion equilibrium constants and FTIR analyses of inclusion complexes between grafted cyclodextrin and the guest molecules. Our results indicated that this nanoadsorbent would be a promising tool for easy, fast and selective separation, analysis of nucleosides and nucleotides in biological samples.

Surface of magnetic silica nanoparticles is modified by grafting with carboxymethyl-β-cyclodextrin (CM-β-CD) via carbodiimide activation. The functionalized magnetic core–shell nanoparticles (MNPs) are characterized by Transmission Electron Microscopy (TEM), Fourier Transform Infra Red (FTIR) spectroscopy, X-ray Diffraction (XRD), X-ray Photoelectron Spectroscopy (XPS) and Vibrating Sample Magnetometer (VSM). These nano-sized particles are scrutinized for adsorption of certain chiral aromatic amino acid enantiomers namely, d - and l -tryptophan (Trp), d - and l -phenylalanine (Phe) and d - and l -tyrosine (Tyr) from phosphate buffer solutions. Adsorption capacities of the coated magnetic nanoparticles toward amino acid enantiomers are in the order: l -Trp > l -Phe > l -Tyr and under the same condition, adsorption capacities are higher for l -enantiomers than the corresponding d -enantiomers. All the equilibrium adsorption isotherms are fitted well to Freundlich model. FTIR studies depict significant changes after adsorption of amino acids onto nanoparticles. The stretching vibration frequencies of N H bonds of the amino acid molecules are changed with complex formation through host–guest interaction. The structure and hydrophobicity of amino acid molecules emphasize the interactions between amino acid molecules and the nano-adsorbents bearing cyclodextrin, thus play important roles in the difference of their adsorption behaviors.

Magnetic particles about 10 nm in size were prepared by chemical precipitation under nitrogen and used for the selective and sequential adsorption of bovine serum albumin (BSA) (pI = 4.7) and lysozyme (LSZ) (pI = 1.1) under different conditions, such as pH and initial protein concentration. The separation ratio of BSA over LSZ at pH 4.6 is about 5, which is about 1.5 times the separation ratio of LSZ over BSA at pH 11.0. Only 10% of the preadsorbed BSA could be displaced by the sequential adsorption of LSZ at pH 11.0. On the other hand, 60% of the preadsorbed LSZ was desorbed due to the sequential adsorption of BSA at pH 4.6. Over 50% desorption of BSA or LSZ could be achieved either by 0.5 M Na(2)HPO(4) or 0.5 M NaH(2)PO(4) after 2 h. Over 80% of the enzymatic activity of LSZ was preserved when it was desorbed from magnetic particles.

Adsorption and desorption of lysozyme on nano-sized magnetic particles and its conformational change were studied in this work. Adsorption of lysozyme on nano-sized magnetic particles (Fe 3 O 4 ) was carried out at different pH. Maximum adsorption of lysozyme (4.65 mg/m 2 ) occurred at its isoelectric point (p I =11.1). Differential scanning calorimetry (DSC) results show that the lysozyme adsorbed on magnetic particles did not show any thermal transition over the range 20–100 °C. High desorption of lysozyme from magnetic particles was achieved using NaH 2 PO 4 (pH 4.0) (90%) and NaSCN (pH 6.0) (97%) as desorbents. The conformational change of the lysozyme desorbed by NaH 2 PO 4 was small, while the lysozyme desorbed by NaSCN underwent a significant conformational change as measured by the intrinsic fluorescence. Eighty-eight and 82% activity was retained in the desorbed enzyme for desorption by NaH 2 PO 4 and NaSCN, respectively.

Adsorption of bovine serum albumin (BSA) on nanosized magnetic particles (Fe(3)O(4)) was carried out in the presence of carbodiimide. The equilibrium and kinetics of the adsorption process were studied. Nanosized magnetic particles (Fe(3)O(4)) were prepared by the chemical precipitation method using Fe2+, Fe3+ salts, and ammonium hydroxide under a nitrogen atmosphere. Characterizations of magnetic particles were carried out using transmission electron microscopy (TEM) and a vibrating sample magnetometer (VSM). Fourier-transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) were used to confirm the attachment of BSA on magnetic particles. Effects of pH and salt concentrations were investigated on the adsorption process. The experimental results show that the adsorption of BSA on magnetic particles was affected greatly by the pH, while the effect of salt concentrations was insignificant at a low concentration range. The adsorption equilibrium isotherm was fitted well by the Langmuir model. The maximum adsorption of BSA on magnetic particles occurred at the isoelectric point of BSA. Adsorption kinetics was analyzed by a linear driving force mass-transfer model. BSA was desorbed from magnetic particles under alkaline conditions, which was confirmed by SDS-PAGE electrophoresis and FTIR results.