Correspondence,Email:hequanguo126.com Tel: +86-731-22182107Preparation and Characterization of Amino-coated Maghemite Nanoparticles Lei Zeng, Rong Hu, Zhaohui Wu and Quanguo He Green Packaging and Biological Nanotechnology Laboratory, Hunan University of Technology Zhuzhou 412008, China AbstractThe ferrous salt and ferric salt were used as raw materials to prepare magnetite (Fe3O4) nanoparticles as precursors, and then maghemite (-Fe2O3) nanoparticles were obtained from the precursors via high temperature pyrolysis for further oxidation，and corresponding amino-coated maghemite nanoparticles are also obtained via surface modification of 3-aminopropyltriethyloxy silane (APTES). The properties and structure of nanoparticles are characterized by transmission electron microscopy(TEM), energy dispersive spectroscopy (EDS), X-Ray photoelectron spectroscopy (XPS), ultraviolet visible spectra (UV-vis) , fourier transform infrared (FT-IR) spectrometer and magnetic property measurement system (MPMS) with superconducting quantum interference device (SQUID) magnetometry. The result reveals that the amino-coated maghemite particles have a slight dimensional increase in average diameter, and maintain almost original saturation magnetization that decreases from 109.2 emug-1 to 102.3 emug-1 after APTES coating. The finding is instructive for surface functionalization of MNPs related to sensing application. Keywords-Maghemite nanoparticles; Coprecipitation; Oxidation; Surface modification; Amino-functionalization I. INTRODUCTION In recent years, magnetic nanoparticles (MNPs) such as maghemite nanoparticles (-Fe2O3 NPs) have attracted a great deal of attention for their wide applications. MNPs not only have the properties of four basic effects of nanomaterials (i.e. surface effect, quantum dimensional effect, volume effect and macroscopic quantum tunnel effect), but also have the unusual magnetic properties dependent on the composition (such as superparamagnetism, high coercivity, low Curie temperature and high magnetic susceptibility, and so on) 1-5.MNPs have replaced many traditional micromaterials including the preparation of high-density magnetic recording materials 6. In biotechnology field, MNPs can accommodate biological molecules (such as DNA, protein, peptide, etc.) onto surface, and produce conjucted species, which often are used in active magnetic resonance imaging 7, targeted drug delivery 8, structural frame of tissue engineering 9, molecular recognition and labeling 10, DNA sensors 11 and biochips due to their similar size and dimension. With wide uses and potential applications in the nucleic acid analysis, clinical diagnosis, targeted drug, cell separation, enzyme immobilization, etc. MNPs have gained increasing research attractions 12-13. The biosensor progress by using MNPs indicates that the detection sensitivity and specificity can be significantly improved in terms of great reduction of biochemical reactions time, convenient manipulation and evident signal amplification. So how to introduce MNPs into various biochemical systems towards biosensor platform fabrication and establishment becomes a challenge focus that could greatly broaden their prospects 14-17. Generally speaking, the preparation methods of MNPs include physical, chemical and biological methods, etc 18-20. However, how to make biological molecules integrated onto the MNPs surface with high immobilization efficiency and good aqueous compatibility and stabilization has become a challenge from an interdisciplinary perspective between nanotechnology and biotechnology research. Therefore, the MNPs derivative or functionalized surface preparation by using appropriate physicochemical methods 21 is a necessary prerequisite and a key step for these purposes. In the paper, the -Fe2O3 NPs with an average diameter of ca. 20 nm were prepared through the precursors Fe3O4 NPs via further oxidation at high temperature and they display high-purity, high saturation magnetization and spherical distribution. The surface of -Fe2O3 NPs was modified with amino-groups (-NH2) by 3-aminopropyltriethoxysilane (APTES). The structure and properties of the bare -Fe2O3 NPs and amino-coated -Fe2O3 NPs were both clearly characterized by TEM, EDS, FT-IR, XPS, UV-vis, and SQUID. Such a success of amino-coated maghemite nanoparticles preparation has an instructive meaning in the field of surface functionalization associated with bimolecular conjugation, stable carrier, as well as sensors application. II. EXPERIMENTAL A. Reagents and instruments Ferric chloride (FeCl36H2O, 99.0%) was purchased from Tianjin Bodi Chemicals Co., Ltd. Ferrous chloride (FeCl24H2O, 99.0%) was purchased from Tianjin Shuangchuan Chemicals Co., Ltd. APTES was purchased from Sigma. Sodium hydroxide (NaOH) was purchased from Hunan Huihong Chemicals Co., Ltd. All of these chemicals were of analytical grade and used directly. Water (T= 25 , 18.2 M) was purified by SUPER ATER-II water purification systems, and ultra-pure water was used throughout the experiment. The NdFeB magnet, purchased locally, was used to separate magnetic particles in the washing and selection steps. 978-1-4244-4713-8/10/$25.00 2010 IEEE Transmission electron microscopy (TEM) images were carried out on a Hitachi H-600 electron microscope (20 kV). Energy dispersive spectroscopy (EDS) was carried out on an EMAX of Hitachi 3000N microscope operated at 25 kV, where sample solutions were deposited onto an Al substrate at ambient temperature. Fourier transform infrared (FT-IR) spectrometer was performed on JEOL JIR-WINSPEC 50. X-ray photoelectron spectroscopy (XPS) was acquired on a Kratos Axis Ultra DLD. This system used a focused monochromatic Al x-ray (1486.6 eV) source for excitation and a spherical section analyzer. Ultravioletvisible spectra (UV-vis) were acquired on a Purkinje General T-1901 spectrophotometer. Superconducting quantum interference device (SQUID) magnetometry was performed on Quantum Design MPMS XL-7. B. Experimental Principle The first step was to synthesize the Fe3O4 NPs as previously reported 22. The chemical reactions were described as following equation (1). From equation (1), the optimized molar ratio of Fe2+：Fe3+：OH- was 1:2:8. The second step was to calcine the Fe3O4 NPs as precursors in muffle furnace at high temperature. Then the Fe3O4 precursors could be oxidized into -Fe2O3 NPs under atmosphere. The chemical reaction equation was as follows: C. Preparation of -Fe2O3 NPs 2 g FeCl24H2O, 5.2 g FeCl36H2O and 12.1 mol/L HCl 0.85 ml were dissolved into 200 ml H2O. The dissolved oxygen of the mixture was eliminated by ultrasound application. Then a solution of 250 ml 0.75 mol/L NaOH was added into the mixture. The temperature of mixed solution was maintained at 80 under N2 atmosphere. Dark precipitates were gradually produced during the reaction. The precipitates were separated by magnetic field and centrifugation. After that, the product was washed with ethanol two times and deionized water three times, then redispersed in ethanol to form 5 g/L solution. The dried product was collected after vacuum drying. The dried sample was precursors of magnetite Fe3O4. Then the dried precursors were oxidized in muffle furnace at 400 for 2 hours and cooled down to room temperature, the -Fe2O3 NPs were obtained finally and redispersed in absolute ethanol (5 g/L). D. Synthesis of APTES-coated -Fe2O3 NPs 25 ml -Fe2O3 ferrofluid (5 g/L) previously prepared was diluted in 150 ml absolute ethanol and sonicated for 30 min to further well disperse the -Fe2O3 NPs. The resulting colloidal solution was transferred to a three-neck flask. Then, 0.4 ml APTES was dropped into above colloidal solution and the mixture was stirred at room temperature and kept for about 7 h. The reactant mixture was centrifugally separated at 10000 r/min for 30 min and the APTES-coated -Fe2O3 NPs was thus gained. Then the NPs were washed with ethanol for five times and were redispersed in ethanol (1 g/L) prior to use. The solid sample was gained by drying in vacuum desiccator at 70. Fig. 1 shows the preparation principle of amino-coated MNPs. Figure 1. Illustration of Synthetic Chemistry of APTES-coated -Fe2O3 NPs III. RESULTS AND DISCUSSION A. Morphological analysis Figure 2. Representative TEM images of -Fe2O3 (a), and APTES-coated -Fe2O3 NPs(b) Fig. 2 shows the representative TEM images of -Fe2O3 (a) and APTES-coated -Fe2O3 NPs (b). Fig. 2 (a) depicts the TEM image of -Fe2O3 NPs, the average size is about 20 nm. The particles appear spherical with high monodispersity in size. Fig. 2 (b) shows the TEM image of amino-coated -Fe2O3 NPs and the average size has a slight increase and tends to agglomerate to some degree owing to the possible intra- and intermolecular crosslinking during amino-functionalization. However, Surface functionalization maintained almost the morphologies of the primary -Fe2O3 NPs. B. Composition characterization Figure 3. EDS profiles of -Fe2O3 (a), and APTES-coated -Fe2O3 NPs(b) The images of TEM show that these NPs morphology appears spherical distribution and some agglomeration. To confirm the composition of these NPs, EDS profiles in situ composition analysis were collected and the results are shown in Fig. 3. Fig. 3(a) is an EDS profile of the bare -Fe2O3 NPs, indicating the presence of iron, oxygen and carbon (the carbon was polluted from conductive adhesive). After amino-coating, the relevant profile in Fig. 3(b) shows different presence of nitrogen and silicon, while the oxygen peak is strengthened evidently. The ratio of N/Si (element percentage) is 6.59%/5.75% and it is close to 1:1, corresponding to the molecular composition ratio of APTES. The EDS profiles demonstrated both Fe and O existence in the modified NPs, so it confirms that the -Fe2O3 NPs are successfully coated and passivated by APTES. C. FT-IR characterization The FT-IR spectra of unmodified and APTES-modified NPs were determined with a JEOL JIR-WINSPEC 50 spectrometer. As shown in Fig. 4, the Fe-O stretching vibration at 581 cm-1, O-H stretching vibration at around 3417 cm-1and O-H deformed vibration at 1631 cm-1 were observed both in Fig. 4 (curve 2), suggesting that -OH groups coating on the surface of -Fe2O3 NPs as reported 23-24. The intensity around 3417cm-1 band in Fig. 4 probably contributed to free amino groups on the APTES-modified -Fe2O3 NPs, which can be overlapped by the O-H stretching vibration band. The outstanding features in Fig. 4 (2) are the appearance of the peaks at 1105, 1382, and 2912 cm-1 respectively. The Si-O stretching vibration was observed at 1105 cm-1 significantly reveals that the covalent bonds of Fe-O-Si are formed after APTES modification. The bands around 2912 and 1385 cm-1 were assigned to -CH2 and C-N stretching vibration, respectively. The results provided the evidences again that APTES was bonded on the -Fe2O3 NPs surface through silanization reaction with -OH groups. Figure 4. FTIR Spectra comparison of -Fe2O3 (a), and amino-coated -Fe2O3 NPs (b) D. XPS characterization Figure 5. XPS wide-scan profiles of -Fe2O3 (1), and APTES-coated -Fe2O3 NPs(2)(a), and the region for (b)Fe(2p),(c) C(1s), (d)Si(2p), (e) N(1s) The surface composition of -Fe2O3 and APTES-coated -Fe2O3 NPs was further analyzed by XPS, and the representative profiles are shown in Fig. 5. A very high concentration of carbon (284.6 eV) is found on the surface of -Fe2O3 NPs in Fig. 5 (a) due to the contaminative carbon. The major characteristic peaks are iron and oxygen, which is accorded with the EDS analysis. The XPS of Fe from the -Fe2O3/APTES NPs as shown in Fig. 5 (b) demonstrates existence of Fe2p1/2 (710.8 eV) which is close to the standard data of -Fe2O3 (Fe2p1/2, 710.4 eV), and it is assigned to the character band energy of -Fe2O3 structure. After amino-coating, there are new peaks of Si2p (d) appearing at 102.5 eV and N1s (e) appearing at 400.8 eV, which elucidated that the particles have been covalently coated by APTES. Meanwhile, the Fe2p peak intensity of the amino-coated sample decreased obviously as compared with that of the bare -Fe2O3 sample. The XPS results are consisted with the results of TEM and EDS, indicating that the magnetic -Fe2O3 core has been fully coated in APTES shell format and greatly reducing the intensity signals of the element inside. E. Optical properties Figure 6. Uv-vis spectra of -Fe2O3(1), and APTES-coated -Fe2O3 NPs(2) Fig. 6 shows UV-vis spectrum of the freshly prepared -Fe2O3 and the amino-coated -Fe2O3 NPs (both dispersed in ethanol). The -Fe2O3 NPs (dispersed in 5 g/L ethanol) have a strong absorption in UV region when the wavelength is less than 400 nm and a weak absorption in visible region. The similar optical properties have been observed in that of the amino-coated -Fe2O3 (dispersed in 1 g/L ethanol). The UV absorption of amino-coated sample is lower than that of uncoated sample; it may be contributed to a lowered dispersed concentration. The visible absorptions of both have no distinguished peaks, which are different with metal coated nanoparticals, such as gold NPs which can appear a strong absorption peak as a result of surface plasmon resonance (SP) 25. F. Magnetic properties A B Figure 7. Hysteresis loops of -Fe2O3 and APTES-coated -Fe2O3 NPs measured at T=300 K and T=5 K(A), Magnetic isolation photo of -Fe2O3 (a) and APTES-coated -Fe2O3 NPs (b)(B) SQUID magnetometry reveals the effect on the saturation magnetization (Ms) by comparing the amino-coated MNPs with the bare -Fe2O3 NPs. Fig. 7 shows the hysteresis loops measured at T = 300 K (close to room temperature) and T = 5 K for -Fe2O3 NPs and amino-coated -Fe2O3 NPs, and the Ms of -Fe2O3 NPs and amino-coated -Fe2O3 NPs was found to be 87.2 emug-1 at 300 K and 102.3 emug-1 at 5 K, respectively. SQUID magnetometry reveals that overlaying -Fe2O3 NPs surface with a shell of APTES has a negligible decrease on magnetic behavior, and both particles are closed to superparamagnetic at 300 K. Obviously, the Ms at 300 K is lower than that at 5 K, which agrees with the formula that increasing the temperature would cause the Ms to decrease 26. The above interpretation in terms of a surface anisotropy, as a result of the interaction with ligands or outer layers like the APTES coating, indicates that the surface anisotropy probably also affects the moment of the inner -Fe2O3 NPs via the exchange interaction with those at the outer surface. Thus, the Ms of -Fe2O3 NPs was decreased by the immobilization of the APTES shell on their surfaces. Its significant to realize the amino particles application in conjugation with biomolecules, target carrier, and sensors. Fig.7 B shows the -Fe2O3 and the amino-coated -Fe2O3 NPs dispersed in ethanol solution. Both of them exhibited a strong magnetization in the presence of a magnetic field. They presented a good magnetic response. IV. CONCLUTION The amino-functionalized -Fe2O3 NPs were successfully prepared from maghemite NPs (which were oxidized by magnetite precursors at high temperature) and further surface APTES modification. 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