Novel hybrid nanostructured materials of magnetite nanoparticles and pectin

Journal of Magnetism and Magnetic Materials 323 (2011) 980–987

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Novel hybrid nanostructured materials of magnetite nanoparticles and pectin Saurabh Sahu, Raj Kumar Dutta n Department of Chemistry, Indian Institute of Technology Roorkee, Roorkee 247 667, India

a r t i c l e i n f o

a b s t r a c t

Article history: Received 21 June 2010 Received in revised form 19 November 2010 Available online 27 November 2010

A novel hybrid nanostructured material comprising superparamagnetic magnetite nanoparticles (MNPs) and pectin was synthesized by crosslinking with Ca2 + ions to form spherical calcium pectinate nanostructures, referred as MCPs, which were typically found to be 100–150 nm in size in dried condition, confirmed from transmission electron microscopy and scanning electron microscopy. The uniform size distribution was revealed from dynamic light scattering measurement. In aqueous medium the MCPs showed swelling behavior with an average size of 400 nm. A mechanism of formation of spherical MCPs is outlined constituting a MNP–pectin interface encapsulated by calcium pectinate at the periphery, by using an array of characterization techniques like zeta potential, thermogravimetry, Fourier transformed infrared and X-ray photoelectron spectroscopy. The MCPs were stable in simulated gastrointestinal fluid and ensured minimal loss of magnetic material. They exhibited superparamagnetic behavior, confirmed from zero field cooled and field cooled profiles and showed high saturation magnetization (Ms) of 46.21 emu/g at 2.5 T and 300 K. Ms decreased with increasing precursor pectin concentrations, attributed to quenching of magnetic moments by formation of a magnetic dead layer on the MNPs. & 2010 Elsevier B.V. All rights reserved.

Keywords: Pectin Calcium pectinate Magnetite nanoparticle Superparamagnetic Zeta potential Saturation magnetization

1. Introduction Hybrid nanomaterials are of tremendous interest in biomedical applications due to the potential synergistic properties that may arise from the combination of two or more precursors. Two such precursors are pectin and magnetite nanoparticles (MNPs). Pectin is a biodegradable natural polymer consisting of linear anionic polysaccharide and is widely explored as a matrix for drug delivery due to its colon specificity [1]. This is mainly attributed to the two reasons: firstly, pectin is resistant to protease and amylase, which are active in upper gastrointestinal (GI) tract [2]. As a result, materials for colon specific delivery, if loaded in pectin could be protected from its dissolution in stomach environment. Secondly, pectin exhibits excellent controlled drug release properties in colon [3,4]. More over pectin cross linked with Ca2 + ions forming microbeads of calcium pectinate is reported to be effective formulation for certain colon specific drug molecules [5]. There has been significant advancement towards reducing the size of calcium pectinate to a few hundred nanometers which illustrated efficient loading and delivery of insulin, 5-Fluorouracil, genes [6–8]. The size reduction of the carrier matrix is encouraging as it might facilitate transport properties through biological pathways and barriers to enhance its bioavailability and functionalities. The second precursor, i.e., MNPs exhibits superparamagnetic property

n

Corresponding author. Tel.: + 91 1332 285280; fax: + 91 1332 286202. E-mail address: [email protected] (R.K. Dutta).

0304-8853/$ - see front matter & 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.jmmm.2010.11.085

[9–11]. Ideally, magnetization interference from domain wall is not expected especially when the particles consist of single magnetic domain in a matrix. In this regard, the MNPs of about 5–20 nm in diameter is considered to be a promising material for several biomedical applications, e.g., cellular imaging, targeted delivery, targeted chemotherapy, magnetic resonance imaging (MRI), hyperthermia [12–16]. These are attributed due to its superparamagnetic susceptibility, high saturation magnetization, biocompatibility and non-toxicity [17,18] and their relative ease of synthesis by co-precipitation method [19]. The hybrid nanomaterials of pectin and MNPs are thus reckoned to be magnetically responsive and are expected to exhibit the inherent controlled drug release properties. This will allow the investigation of new concepts like magnetic transportation and control release of drug molecules or other suitable substrate for colon specific sites. However, the hybrid nanomaterials if administered orally will transit through the stomach where typically the residence time is 2 h [20]. Gastric juice in the stomach consisting of pepsin, mucus and hydrochloric acid (HCl), constitutes pH  1.2 [21], and favors dissolution of MNPs [22]. Therefore, to retain the magnetic targeting efficiency of the hybrid nanomaterials for colon specific sites, it is very important to protect the MNPs of the hybrid system from acid dissolution in the gastric environment during its transit through stomach. We report here the synthesis of hybrid nanostructured materials consisting of MNPs encapsulated with calcium pectinate, referred to as MCPs. An optimum concentration of polyanionic pectin is in situ cross-linked with Ca2 + ions in the presence of MNPs

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dispersed with biodegradable tween-80. The current synthetic method does not involve any organic solvent. A mechanism is proposed regarding the formation of the MCPs based on several characterization methods. Besides their nanoscale dimensions, these hybrid nanostructures have the advantages of magnetic targeting, controlled release and are chemically resistant in stomach condition. Such hybrid nanomaterials will find various biomedical applications, especially in targeted drug delivery system [23–25].

2. Experimental 2.1. Materials Fe(NO3)3  9H2O, FeSO4  7H2O, liquid ammonia, anhydrous CaCl2 and other reagents for synthesis were all of analytical grade from Merck, India and were used without further purification. Pectin with 65–70% degree of esterification was procured from Hi-Media lab, India. Millipore water (resistivity of 18.1 MO cm at 25 1C) was used in all the experiments. 2.2. Synthesis of MNPs The MNPs were synthesized by rapidly adding liquid ammonia to a solution of a mixture of Fe(NO3)3  9H2O and FeSO4  7H2O with a molar ratio of 2:1, until the pH of the solution reached 10.0 70.1. The mixture was vigorously stirred for about 45 min as reported by Mikhaylova et al. [26]. However, the proposed method was modified by using biocompatible surfactant (0.2% v/v, tween-80), for dispersing the as-synthesized MNPs to achieve better particle size distribution. The excess tween-80 was magnetically decanted and washed with Millipore water followed by lyophilization, which resulted into black colored solid phase MNPs. 2.3. Synthesis of MCPs The pectin solutions of 0.2%, 0.4%, 0.6%, 0.8% and 1.0% w/v were prepared in Millipore water by continuously stirring for 24 h at room temperature. The 50 mL of aqueous dispersion of MNPs (pH was adjusted to about 4 with dilute HCl) was mixed with 50 mL pectin solutions of respective concentrations at pH 4 and this mixture was stirred vigorously for 1 h at constant pH 4. 50 mL of CaCl2 solution (Ca2 + /pectin mass ratio¼2:1) was then added drop wise to cross-link pectin by ionotropic gelation method [27] along with vigorous stirring for 6 h. Corresponding to the concentration of the pectin solution used (0.2–1.0% w/v), the samples of MCPs synthesized were referred to as MCP-0.2, MCP-0.4, MCP-0.6, MCP-0.8 and MCP-1.0, respectively. A fraction of each of these MCPs was isolated for their characterization by SEM, TEM, DLS and zeta potential measurements. The rest of the synthesized nanomaterials were magnetically separated and washed several times with Millipore water to remove excess pectin and were lyophilized. In addition, calcium pectinate nanomaterials without MNPs were synthesized by ionotropic gelation method. It was used as a reference sample for confirming the formation of calcium pectinate in the hybrid nanostructured materials of MCPs.

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dispersive X-ray analyzer (FESEM–EDAX, FEI-Quanta 200F) operated at 20 kV. For TEM studies, the as-synthesized samples were highly diluted and a drop of this suspension containing nanomaterials was placed on a carbon coated 150 mesh copper grid and dried at room temperature. For SEM studies, diluted solution containing nanomaterials were sprayed on a clean glass plate, which was dried and coated with thin layer of Au to impart necessary electrical conductivity for the incident electrons. The DLS measurements were performed by using the Malvern Zetasizer Nano ZS90 instrument with a 4 mW He–Ne laser (633 nm wavelength) and a detector at a 1731 fixed angle. The size measurements were carried out in triplicate at 25 1C by transferring 1 mL of dust free sample suspension into fourclear-size disposable polystyrene cell (Malvern). In order to find evidence of the formation of nanostructured MCPs, the zeta potential measurements were carried out in triplicate at 25 1C by injecting 0.75 mL of dust free sample suspension into disposable folded capillary cells (Malvern). The molecular vibrations of MCPs were obtained by recording FT-IR (Nicolet, Nexus) spectra. Pellets of the dried samples were made with KBr and were scanned in the range of 500–4000 cm  1. The TGA measurements of samples were measured using Perkin Elmer, Pyris Diamond under a nitrogen flow (200 mL min  1) with a heating rate of 5 1C min  1 from ambient temperature up to 800 1C to ensure mass loss due to thermal degradation of the polymer and to minimize the increase in mass due to oxidation of iron in air. A superconducting quantum interference device (SQUID) magnetometer (MPMSXL, USA) was used to analyze the magnetic properties of hybrid nanomaterials. A known amount of lyophilized samples were packed in a diamagnetic capsule and were inserted in a polyethylene straw as a sample holder. The magnetization measurements were recorded from the hysteresis loop of M–H curve in the range 72.5 T at 300 K. The FC and ZFC measurements were recorded at an applied field of 200 Oe by scanning between 5 and 300 K. The XPS measurements were recorded on ESCA VSW scientific instruments Ltd., with AlKa as the source for excitation, operated at 10 kV with an emission current of 10 mA. The sample for XPS characterization was prepared by sprinkling dried lyophilized sample on silver paste applied on Cu holder. Quantitative analysis of the composition of the sample surface was performed by collecting the integrated intensities of C1s, O1s, Ca2p and Fe2p3/2 signals using the Wagner’s sensitivity factors. The dissolution studies of the MNPs and MCPs were conducted as per standard protocol of United State Pharmacopoeia in 900 mL of freshly prepared SGF solution, pH 1.2) at 3770.1 1C and 100 rpm for 120 min in order to mimic the physiological conditions similar to that of gastrointestinal tract of human body. A 5 mL of dissolution fluid was withdrawn at each specified time intervals and replaced with equal volume of fresh medium to mimic the sink conditions of the human body. The withdrawn fluid was filtered and iron content was estimated with Shimadzu 1600 UV Spectrophotometer using phenanthroline method by recording the absorbance at 510 nm [28].

3. Results and discussion 3.1. Synthesis and morphology of hybrid nanostructured MCPs

2.4. Characterization The X-ray diffraction measurements were performed with a powder diffractometer (Bruker AXS D8 Advance) using graphite monochromatized CuKa radiation source. The morphology of the as synthesized MNPs and the MCP hybrid nanostructured materials were studied using the transmission electron microscopy (TEM) operated at 200 kV FEI Technai-G2 microscope and the field emission scanning electron microscopy (beam resolution of 2 nm) with energy

The strategy to prepare the hybrid nanostructured MCPs involved the synthesis of stable MNPs by co-precipitation method followed by its in-situ encapsulation with pectin. The Ca2 + ions were further used for cross linking pectin to form nanostructured hybrid materials of MCPs. The above method was optimized to achieve uniform size distribution of these spherical nanostructures. The key parameters involved in the optimization process were pH 4, pectin precursor concentration (in % w/v), Ca2 + /pectin

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mass ratio of 2:1 and vigorous stirring for 6 h. The pectin concentration of 0.4% and 0.6% were found to be suitable for synthesizing spherical shaped MCP-0.4 and MCP-0.6, respectively. Pectin of 0.2% w/v was found insufficient for complete encapsulation of MNPs, while 1.0% w/v resulted into formation of polymeric matrix due to its cross linking with Ca2 + ions. In order to achieve maximum cross linking it might be desirable to prepare these hybrid nanostructured materials at pH45.5 where the pectin molecules exhibit very high zeta potential values (z ¼  52 to  56 mV) [29]. However, such higher pH was restricted due to the onset of de-polymerization effect of pectin at pH 45.5 [30]. On the other hand, lower pH (e.g., pH  2) tends to reduce the zeta potential of pectin [6] and hence may not be suitable for chemical cross-linking with Ca2 + ions. Besides, lower pH would favor dissolution of the MNPs encapsulated in the hybrid nanostructures of MCPs due to weakening of the Fe–O bond by protonation mechanism [31] and might lead towards loss of its magnetic property. As a compromise, pH  4 was chosen optimum for synthesis of MCPs which offered suitably high zeta potential of pectin (z ¼  35.7 mV, Table 1). The formation of as synthesized stable MNPs were confirmed by recording the position and relative intensities of diffraction patterns at 220, 311, 400, 511 and 440 planes (Fig. 1) which corroborated well with those of cubic magnetite structures as reported in JCPDS 01-11111 data. The average particle size (D) of the MNPs was found to be about 2 nm, using the Debye–Scherrer formula, i.e. D ¼(0.9l)/(D cos y), where l is X-ray wavelength, D is line broadening measured at half-height from the most intense peak of XRD (311 plane) and y is Bragg angle of the particles. The magnetite phase was also evident in all the samples of the MCPs as supported from respective diffraction patterns (Fig. 1). However, in MCP-1.0 the magnetite phase was weak, which might be attributed to the matrix effect or due to encapsulation of very small size of magnetite nanoparticles. The TEM measurements revealed that the particle sizes of the as-synthesized MNPs were in the range between 2 and 8 nm

(Fig. 2a), and the corresponding SAED image indicated its polycrystalline nature (Fig. 2b). After coating the MNPs with calcium pectinate the resulting nanostructured hybrid materials of MCP-0.4 and MCP-0.6 were found to be mostly spherical with size distribution in the range of 50–200 nm as evidenced from SEM study (Fig. 3a). Notably, the particles of sizes 100–150 nm were most frequently observed (inset of Fig. 3a). The co-localization of Fe and Ca in the EDAX analysis of a selected area of a representative spherical nanostructure (Fig. 3b) was characteristic of MNPs and calcium pectinate, respectively, and thus indicated the formation

Table 1 Zeta potential measurements of magnetite nanoparticles (MNPs), pectin solution and MNP coated calcium pectinate nanostructured hybrid materials (MCPs).

1200

440

 41.2 + 17.1  35.7  17.9  14.7  14.6

511

MNPs at pH  10 MNPs at pH  4 Pectin at pH  4 MCP-0.4 MCP-0.6 MCP-1.0

400

Measured zeta potential value (z) in mV

220 311

Samples

MNPs MCP-0.2

Intensity

900

MCP-0.4 MCP-0.6 MCP-0.8

600 300

MCP-1.0

0 20

30

40

50 60 2θ degree

70

80

Fig. 1. XRD of the MNPs and various compositions of MCPs.

Fig. 2. (a) TEM image of MNPs and (b) the SAED image corresponding to (a) confirmed polycrystalline MNPs.

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Fig. 3. (a) Morphology and particle size distribution of MCP-0.4 by SEM, (b) EDAX of a selected nanostructure of MCP-0.4, marked in the inset, illustrated co-localization of Fe and Ca, (c) TEM of spherical shaped MCP-0.4 nanostructure (marked) of about 150 nm, and (d) detailed TEM of a part of MCP-0.4 nanostructure, as shown by an arrow in (c), illustrated the presence of multiple polycrystalline MNPs.

of MCPs. Further, the TEM analysis of the MCP-0.4 illustrated spherical hybrid nanostructures of 150 nm size which corroborated the SEM results (Fig. 3c). A detailed TEM study of a part of a representative nanostructure of MCP-0.4 revealed encapsulation of a large number of MNPs (Fig. 3d) and its SAED pattern signified polycrystalline nature (inset of Fig. 3d). Furthermore, DLS measurement of MCP-0.4 in aqueous medium at pH 4 exhibited unimodal size distribution in the range of 250–620 nm with maximum intensity at  400 nm (Fig. 4). This indicated that the method of synthesis offered a reasonably good

control over the size of these hybrid nanostructures. It may however be noted that the particle size measured by DLS were larger as compared to those measured by TEM and SEM. This might be attributed due to possible swelling behavior of calcium pectinate coating material in aqueous medium. Similar swelling effect was reported for calcium pectin hydrogels [32]. The DLS measurements of MCP-0.4 were carried out in aqueous media with pH ranging between 1 and 7, which showed quite similar size distribution as typically observed in Fig. 4, which might be due to saturation of swelling in aqueous media at various pH.

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Intensity (%)

25 20 15 10 5 0 0.1

1

10

100

1000

10000

Size (d.nm) Fig. 4. DLS measurement of MCP-0.4 in aqueous medium at pH 4.

cross linking

pH~4

Pectin

2+

Ca

MNPs

ions MCP hybrid nanostrucutres

COOHO

O H HO

H H

H

H OH

HO H O

OH H

-

O H

H

O

OOC

n

Structure of a pectin monomer unit Fig. 5. Schematic representation of the proposed mechanism for the synthesis of MCP hybrid nanostructures.

3.2. Mechanism of the formation of MCP hybrid nanostructured materials The zeta potential measurements (given in Table 1) offered an insight of the proposed mechanism towards formation of the MCPs as schematically represented in Fig. 5. Firstly, the highly stable MNPs, as confirmed from its zeta potential (  41.2 mV) measurement, were synthesized at pH  10. These stable MNPs were conditioned at pH 4 to interact with pectin. At this pH, the zeta potential of MNPs was found to be + 17.1 mV which was considered to be favorable for electrostatic interaction with polyanionic pectin molecules (z ¼  35.7 mV at pH  4) to form an MNP–pectin interface. The carboxylic groups of pectin molecules were cross-linked by Ca2 + ions to form spherical hybrid nanostructures, where the MNPs were encapsulated by calcium pectinate. The likelihood of formation of such spherical shape could be explained in terms of attaining stability by achieving minimum surface energy. The cross-linking of pectin might be interpreted from the lowering of zeta potential measurements of MCP-0.4 (  17.9 mV), MCP-0.6 (  14.7 mV) and MCP-1.0 ( 14.6 mV) as compared to that of pectin. The lowering of zeta potential might be attributed to shielding of charge density on the pectin molecules due to its electrostatic interactions with Ca2 + ions. The FT-IR and TGA analyses of the samples further supported the proposed mechanism. The FT-IR spectra (Fig. 6) of the pectin showed

Fig. 6. FT-IR spectrum of MNPs, MCP-0.4 and MCP-0.6, precursor pectin and calcium pectinate reference sample.

weak bands at 1690 cm  1 corresponding to COOH group while that of MCP revealed intense bands at 1685, 1395 and 1324 cm  1 which were characteristic of asymmetric and symmetric stretching of carboxylate groups (COO  ). Similar intense IR bands were also recorded for the synthesized calcium pectinate reference sample. These observations

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the binding energy of the photoelectron peaks corresponding to C1s, O1s, and Ca2p3/2, which were the signatures of calcium pectinate (Fig. 8). It also showed weak binding energy peak corresponding to Fe2p3/2 (inset of Fig. 8) and the concentration of Fe was calculated to be 1.670.6%. 3.3. Stability of MCPs in simulated gastric fluid Considering the potential applications of using the hybrid nanostructured MCPs for colon specific delivery via oral administration, it is essential for the MCPs to be protected from gastric degradation during its transit through stomach (pH 1.2) to avoid loss of magnetic materials. The non-degradability of MCPs in gastric condition would further ensure protection of loaded materials (e.g., drugs) for colon specific delivery. This aspect was studied by treating MCP-0.4 and equivalent amount of MNPs present in MCP-0.4 for a total of 120 min in simulated gastric fluid (SGF). The cumulative Fe release in SGF from MCP-0.4 in sink condition was found to be only 1.9% for a period of 120 min (Fig. 9a), while that of MNPs was 21.3% (Fig. 9b). The low Fe release from MCP-0.4 in SGF corroborated well with the Fe concentration at the surface of MCP-0.4 calculated from the XPS measurement. This indicated that most of the MNPs were encapsulated within the MCP nanostructure and were well protected from gastric environment. Degradation of MCPs would have otherwise caused significant release and dissolution of MNPs in SGF. It may therefore be

1300

Intensity

3000

Intensity

2500 2000

Fe2p3/2

1200 1100 1000 900 700

1500

710 720 730 740 Binding Energy(eV)

O1s 1000

C1s

500

Cl1s

Ca2p

0 0

200

400 600 800 Binding Energy (eV)

1000

Fig. 8. XPS of MCP-0.4 (full scan) and detailed scan for Fe analysis in the inset.

b % Cumulative Fe Release

indicated the formation of calcium pectinate in the hybrid nanostructures of MCPs. However, the IR bands corresponding to carboxylate group (1390 and 1620 cm  1) were also observed for the assynthesized MNPs which were attributed to the carboxylate groups of tween-80 used for stabilizing the MNPs. In addition, the IR signatures for pectin and the MCPs were found similar in the fingerprint region of 900–1250 cm  1 which might be due to their similar molecular structures. However, it was not possible to rule out the presence of certain amount of pectin which was not modified to calcium pectinate during the synthesis of MCPs. The formation of MCP-0.4 and MCP-0.6 was further confirmed from TGA studies (Fig. 7), which illustrated nearly 5% higher mass loss for MCP-0.6 as compared to MCP-0.4. This was attributed to higher pectin precursor concentration for synthesis of MCPs. In addition, their TGA thermogram comprised with three distinct thermal events. The first event was recorded in the temperature range between ambient temperature and 110 1C corresponding to a mass loss of about 11% which was due to desorption of water molecules. The second event showed a gradual mass loss of about 17% in the temperature range of 200–450 1C, which was comparable with the thermal degradation pattern of the calcium pectinate. The third thermal event corresponding to nearly 11% mass loss in the temperature range of 550–775 1C did not concur with that of calcium pectinate. However, this thermal event appeared to be correlated with the thermal decomposition pattern of precursor pectin corresponding to the temperature range of 320–455 1C. From these observations, it may be inferred that the hybrid nanostructured MCPs contained both pectin as well as calcium pectinate components. Most likely, calcium pectinate constituted the periphery of the MCPs that enclosed pectin interfaced with MNPs. According to this proposed MCP nanostructure, the thermal decomposition of pectin would be expected after the thermal degradation of calcium pectinate at the periphery. This was perhaps the reason why the onset of thermal decomposition of pectin in the MCPs occurred at much higher temperature than that of the precursor pectin. Furthermore, the formation of calcium pectinate at the periphery of the MCPs could be further deduced by comparing the mass loss phenomenon due to desorption of water at 110 1C for MCP with that of precursor pectin and calcium pectinate reference sample. For MCPs this mass loss was much higher (11%) as compared to that of pectin (4%) but less than that of calcium pectinate (33%). The higher affinity for calcium pectinate to adsorb water was also correlated from the broad IR band observed in the range 3000–3600 cm  1. The nature of mass loss at 110 1C for MCPs therefore supported the formation of calcium pectinate at the periphery of MCP. This was confirmed from the surface analysis by X-ray photoelectron spectroscopy studies of MCP-0.4. It revealed

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20

15

10

5

a 0 0

Fig. 7. TGA analysis of MNPs, MCP-0.4, MCP-0.6, pectin and calcium pectinate.

20

60 40 80 Time (Minutes)

100

120

Fig. 9. Dissolution profile of Fe release from MNPs (a) MCP-0.4, (b) in SGF for a total period of 120 min.

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interpreted that the MCP-0.4 was impermeable to acidic solution and ensured minimal loss of magnetic materials.

80 MNPs MCP-0.4

The superparamagnetic behavior of the hybrid nanostructures of MCP-0.4 was studied using the temperature dependence magnetization between 5 and 300 K of zero-field cooling (ZFC) and that of field cooling (FC) in an applied magnetic field of 200 Oe (Fig. 10). At low temperatures (for FC curve), an externally applied magnetic field energetically favored the reorientation of the individual magnetic moment and hence resulted in the observed magnetization along the direction of the applied field, which decreased with increasing temperature. On the other hand, in the absence of applied magnetic field and when the sample was cooled the total magnetization was zero (as shown in ZFC curve), due to the random orientation of the magnetic moments of individual particles. As the temperature was increased more particles reoriented their magnetic moment (magnetization) along the external applied field. Due to this the magnetization increased till it reached a maximum value at 93.3 K, referred as blocking temperature of the synthesized MNPs, as shown in the ZFC curve (Fig. 10) and then its profile was similar to that of the FC curve. These features are typically observed for small grain sizes of magnetite nanoparticles [33] which are reported to exhibit superparamagnetism for particles of sizes less than 20 nm [34] due to lack of well defined domain structure. However, from the FC–ZFC curve shown in Fig. 10, it may be argued that both ferromagnetic and superparamagnetic regimes coexisted owing to the distribution of grain size of magnetite nanoparticles in the batch of MCP. In that case, the magnetic behavior exhibited by the MCP could be attributed to partial volume of the magnetite nanoparticles with sizes corresponding to these two regimes. It was noted that the magnetization curves (M–H) of different compositions of MCPs recorded at 300 K from the hysteresis loop in SQUID measurements were similar to that of the as-synthesized MNPs, and exhibited negligible coercivity and remanence magnetization (Fig. 11). This property was attributed to superparamagnetic behavior of the magnetite nanoparticles, expected for sizes less than 20 nm and hence consistent with our synthesized magnetite nanoparticles which are 2–8 nm in sizes. It may be inferred that the magnetic nanoparticles in the synthesized batches of MCP-0.4 are predominantly superparamagnetic in nature which further corroborated the ZFC and FC studies.

18

M (emu/g)

40 3.4. Magnetic properties of hybrid nanostructures of MCP

MCP-0.6 0

MCP-1.0

-40 -80 -20000

0 H (Oe)

20000

Fig. 11. Magnetization for dry samples of MNPs and MCPs at 300 K.

As the precursor pectin concentration was increased from 0.4% to 1.0% w/v the saturation magnetization (Ms) of their corresponding hybrid nanostructures of MCP decreased. The measured Ms at 300 K and 2.5 T were 46.21, 38.70 and 3.05 emu/g for MCP-0.4, MCP-0.6 and MCP-1.0, respectively. These values were however smaller than that of the as-synthesized MNPs (52.16 emu/g) and notably the Ms reduced significantly, when 1.0% precursor pectin concentration was used for synthesis of MCP-1.0. The drastic reduction of saturation magnetization due to increase in pectin concentration could be attributed to various reasons. It might be due to effect of small particle size owing to non-collinear spin arrangement at the surface [35], or due to the formation of magnetic dead layer by pectin at the domain boundary wall of MNPs [36,37]. The particle size effect on reduction of Ms may be ruled out as the same batch of MNPs with uniform particle size distribution was used for synthesis of different MCP compositions. The magnetic moments could however be quenched due to the formation of magnetic dead layer at the domain wall of MNPs. This could hinder the domain wall motion during application of the magnetic field, which might be responsible for the reduction in the saturation magnetization in MCPs. In this regard, pectin might form a magnetic dead layer at the surface of MNPs during the formation of the proposed MNP–pectin interface. The reduction in the magnetization for MCP-0.6 as compared to those of MCP-0.4 was attributed to formation of thicker dead layer of pectin at the domain boundary wall of encapsulated MNPs. It may therefore be inferred that the synthesis of hybrid nanostructures of MCPs resulted in pectin–MNP interface with formation of calcium pectinate at the periphery. This system as a whole exhibited superparamagnetic behavior with high magnetization at applied field of 2.5 T but appeared to be influenced by precursor pectin concentration.

16

M (emu/g)

4. Conclusion

14

12

10

8 0

50

100

150

200

250

Temperature (K) Fig. 10. ZFC and FC curve of MCP-0.4 recorded at 200 Oe.

300

A facile and an in-expensive method has been developed to synthesize a novel hybrid nanostructured material (MCP) by completely encapsulating stable superparamagnetic magnetite nanoparticles (MNPs) with pectin followed by cross linking with Ca2 + ions. The superparamagnetic nature of the as synthesized MCP-0.4 was confirmed by measuring ZFC–FC profile at an applied field of 200 Oe and the blocking temperature was found to be 93.3 K. The MCPs showed considerable magnetism, although their saturation magnetization value was lower than that of the bare MNPs and decreased with increasing precursor pectin concentration. The magnetite phase of the MNPs was confirmed from XRD and its encapsulation by calcium pectinate to form MCPs was probed by FT-IR, TGA and XPS studies. The MCPs were mostly spherically shaped with sizes ranging typically between 100 and

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150 nm in dry condition as evidenced by TEM and SEM. However, in aqueous medium at pH 4, these nanostructures showed swelling effect with an average size of  400 nm as determined by DLS measurement, and evidenced unimodal size distribution. This indicated that the method used in the synthesis of MCP nanostructures offered a good control over their sizes. Furthermore, the MCPs were reasonably stable in aqueous medium as revealed from their zeta potential measurements and they were found to be impermeable in acidic condition indicating minimal loss of magnetic material in simulated gastric condition. Overall, it may be considered that the novel magnetically responsive hybrid nanostructures comprising superparamagnetic MNPs and pectin could potentially find a wide range of biomedical applications, e.g., targeted delivery of drugs or other biomolecules, imaging by MRI or magnetic fluid hyperthermia.

Acknowledgements One of the authors (S.S.) would like to thank MHRD, India for awarding fellowship. Authors thank Institute Instrumentation Centre and Centre of Nanotechnology, IIT Roorkee for instrumental facilities. Authors also thank Dr. T. Shripathi, and Prof. Ajay Gupta of UGC DAE, CSR Indore Centre, India for extending their XPS experimental facility. References [1] [2] [3] [4] [5] [6] [7] [8] [9]

L. Liu, M.L. Fishman, J. Kost, K.B. Hicks, Biomaterials 24 (2003) 3333–3343. H.N. Englyst, S. Hay, G.T. MacFarlane, FEMS Microbiol. Ecol. 95 (1987) 163–171. S.A. Sande, Expert Opin. Drug Deliv. 2 (2005) 441–450. S.A. Sande, S. Sungthongjeen, T. Pitaksuteepong, A. Somsiri, P. Sriamornsak, Drug Dev. Ind. Pharm. 25 (1999) 1271–1276. S. Das, K.Y. Ng, J. Pharm. Sci. 99 (2009) 840–860. K. Cheng, L.Y. Lim, Drug Dev. Ind. Pharm. 30 (2004) 359–367. C.Y. Yu, H. Cao, X.C. Zhang, F.Z. Zhou, S.X. Cheng, X.Z. Zhang, R.X. Zhuo, Langmuir 25 (2009) 11720–11726. P. Opanasopit, A. Apirakaramwong, T. Ngawhirunpat, T. Rojanarata, U. Ruktanonchai, AAPS Pharm. Sci. Technol. 9 (2008) 67–74. C.C. Huang, T.Y. Liu, C.H. Su, Y.W. Lo, J.H. Chen, C.S. Yeh, Chem. Mater. 20 (2008) 3840–3848.

987

[10] S. Maenosono, T. Suzuki, S. Saita, J. Magn. Magn. Mater. 320 (2008) L79–L83. [11] G. Cheng, D. Romero, G.T. Fraser, A.R.H. Walker, Langmuir 21 (2005) 12055–12059. [12] J.W.M. Bulte, D.L. Kraitchman, NMR Biomed. 17 (2004) 484–499. [13] P. Dames, B. Gleich, A. Flemmer, K. Hajek, N. Seidl, F. Wiekhorst, D. Eberbeck, I. Bittmann, C. Bergemann, T. Weyh, L. Trahms, J. Rosenecker, C. Rudolph, Nat. Nanotechnol. 2 (2007) 495–499. [14] D.B. Shieh, F.Y. Cheng, C.H. Su, C.S. Yeh, M.T. Wu, Y.N. Wu, C.Y. Tsai, C.L. Wu, D.H. Chen, C.H. Chou, Biomaterials 26 (2005) 7183–7191. [15] F.X. Hu, K.G. Neoh, E.T. Kang, Biomaterials 27 (2006) 5725–5733. [16] C.H. Hou, S.M. Hou, Y.S. Hsueh, J. Lin, H.C. Wu, F.H. Lin, Biomaterials 30 (2009) 4700–4707. [17] C. Sestier, Z.G.M. Lacava, L.M. Lacava, M.F. Da Silva, R.B. Azevedo, N. Buske, C. Gansau, P.C. Morais, O. Silva, F. Pelegrini, D. Sabolovicm, J. Magn. Magn. Mater. 252 (2002) 403–405. [18] A.L.D. Silva, T. Trindade, B.J. Goodfellow, B.F.O. Costa, R.N. Correia, A.M. Gil, Biomacromolecules 8 (2007) 2350–2357. [19] G. Gnanaprakash, S. Mahadevan, T. Jayakumar, P. Kalyanasundaram, J. Philip, B. Raj, Mater. Chem. Phys. 103 (2007) 168–175. [20] N. Washington, C. Washington, C.G. Wilson, Physiological Pharmaceutics: Barriers to Drug Absorption, second ed., Taylor and Francis Inc., London, 2001. [21] V.R. Sinha, R. Kumria, Int. J. Pharm. 249 (2002) 23–31. [22] P.W. Schindler, Pure Appl. Chem. 63 (1991) 1697–1704. [23] A. Kumar, B. Sahoo, A. Montpetit, S. Behera, R.F. Lockey, S.S. Mohapatra, Nanomed. Nanotechnol. Biol. Med. 3 (2007) 132–137. [24] L.B. Asperilla, M. Darder, P. Aranda, L. Vazquez, M. Vazquez, E.R. Hitzky, J. Mater. Chem. 17 (2007) 4233–4238. [25] T. Xu, N. Zhang, H.L. Nichols, D. Shi, X. Wen, Mater. Sci. Eng. C 27 (2007) 579–594. [26] M. Mikhaylova, D.K. Kim, N. Bobrysheva, M. Osmolowsky, V. Semenov, T. Tsakalakos, M. Muhammed, Langmuir 20 (2004) 2472–2477. [27] P. Sriamornsak, Eur. J. Pharm. Sci. 8 (1999) 221–227. [28] G.D. Christian, Analytical Chemistry, sixth ed., John Wiley & Sons, Singapore, 2003. [29] G.T. Grant, E.R. Morris, D.A. Rees, P.J.C. Smith, D. Thom, FEBS Lett. 32 (1973) 195–198. [30] C. Rolin, B.U. Nielsen, P.E. Glaahn, in: S. Dumitriu (Ed.), Polysaccharides, Marcel Dekker, New York, 1998, pp. 377–432. [31] I.H.M. Van Oorschot, M.J. Dekkers, Geophys. J. Int. 145 (2001) 740–748. [32] I. Mika, H. Tatsuko, H. Hyoe, Thermochim. Acta 431 (2005) 68–72. [33] S. Si, A. Kotal, T.K. Mandal, S. Giri, H. Nakamura, T. Kohara, Chem. Mater. 16 (2004) 3489–3496. [34] T.J. Daou, G. Pourroy, S. Begin-Colin, J.M. Greneche, C. Ulhaq-Bouillet, P. Legare, P. Bernhardt, C. Leuvrey, G. Rogez, Chem. Mater. 18 (2006) 4399–4404. [35] A. Cesaro, F. Delben, S. Paoletti, Int. J. Biol. Macromol. 12 (1990) 170–176. [36] K.V.P.M. Shafi, A. Ulman, A. Dyal, X. Yan, N.L. Yang, C. Estournes, L. Fournes, A. Wattiaux, H. White, M. Rafailovich, Chem. Mater. 14 (2002) 1778–1787. [37] M.P. Morales, S.V. Verdaguer, M.I. Montero, C.J. Sterna, A. Roing, L. Casas, B. Martinez, F. Sandiumenge, Chem. Mater. 11 (1999) 3058–3064.