Synthesis and characterization of uniform radiopaque polystyrene

Synthesis and characterization of uniform radiopaque polystyrene microspheres for X-ray imaging by a single-step swelling process Anna Galperin,1 David Margel,2 Shlomo Margel1 1 Department of Chemistry, Bar-Ilan University, Ramat-Gan 52900, Israel 2 Institute of Urology, Rabin Medical Center, Beilinson Campus, Petah-Tikva, Israel Received 8 February 2006; revised 23 March 2006; accepted 3 April 2006 Published online 20 June 2006 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.a.30863 Abstract: Uniform radiopaque polystyrene microspheres of 2.3 6 0.2 lm were prepared by a single-step swelling of 2.3 6 0.2 lm polystyrene template microspheres, dispersed in an aqueous solution with methylene chloride emulsion droplets containing 2,3,5-triiodobenzoylethyl ester. After completing the swelling process, the methylene chloride was evaporated in order to lock the 2,3,5-triiodobenzoylethyl ester in the polystyrene microspheres. The influence of the weight ratio [2,3,5-triiodobenzoylethyl ester]/[polystyrene] on the % of entrapped 2,3,5-triiodobenzoylethyl ester was elucidated. Characterization of the radiopaque polystyrene microspheres was accomplished by light microscope, FTIR, TGA, SEM, XPS, and elemental analysis. The radiopacity of the microspheres was demonstrated

by an imaging technique based on X-ray absorption usually used in hospitals. This novel method of encapsulation of 2,3,5-triiodobenzoylethyl ester within polystyrene microspheres by a single-step swelling process may be used as a model for encapsulation of various hydrophobic radiopaque drugs within uniform polystyrene template particles of various diameters for different X-ray imaging needs, e.g., blood pool, body organs, embolization, dental composition, implants, protheses, and nanocomposites. Ó 2006 Wiley Periodicals, Inc. J Biomed Mater Res 79A: 544–551, 2006

INTRODUCTION

compounds containing a heavy atom substituent as physical mixtures with an appropriate polymer.8–10 Radiopaque polymer–salt complexes have been produced by incorporation of radiopaque heavy metal salts into an appropriate polymer ligand via chelation. Radiopaque polymers have also been formed by polymerization of methyl methacrylate with metal salts of vinyl monomers such a barium or zinc acrylates.16 Another approach to preparing radiopaque polymers is based on copolymerization of vinyl monomers containing covalently bound halogen atoms such as iodine with other vinyl monomers.4,11–21 Radiopaque polymers were also synthesized by grafting iodinecontaining molecules onto preformed high-molecular-weight polymers.5–7,15 The present article describes a novel method to prepare new radiopaque polymeric contrast agent particles, based on a single-step swelling of uniform template polystyrene (PS) microspheres dispersed in an aqueous solution with an hydrophobic swelling solvent such as methylene chloride containing 2,3,5triiodobenzoylethyl ester (TIBEE). After completing the swelling process, the methylene chloride is evaporated so as to lock the TIBEE in the PS microspheres.

Recent literature of novel biomaterials shows that there is an increasing interest in developing radiopaque polymers as contrast agents for X-ray imaging.1–21 The radiopaque polymeric agents may be used for various applications, e.g., imaging of blood pool1,2 or certain body organs3 in order to detect or diagnose various disease states, monitoring embolization process,4–10 evaluation of implants used in surgery to determine their exact location,11–16 and dental composition.16–18 There are different techniques to prepare radiopaque polymers of various types. For example, radiopaque polymer blends have been prepared by incorporating radio-opacifying agents such as heavy metal powders, inorganic salts of a heavy element, or organic

Correspondence to: S. Margel; e-mail: shlomo.margel@mail. biu.ac.il Contract grant sponsors: Minerva grant; Israeli Ministry of Science

Ó 2006 Wiley Periodicals, Inc.

Key words: radiopaque microspheres; polystyrene microspheres; single-step swelling; encapsulation; X-ray imaging

SYNTHESIS AND CHARACTERIZATION OF RADIOPAQUE PS MICROSPHERES

Ugelstad and coworkers invented a useful multi-step swelling method of uniform template particles with various acrylate monomers and initiators for the production of different uniform-sized particles of controlled, desired properties.22,23 This basic swelling process was then significantly elaborated by Cheng et al.24 and Hosoya and coworkers and Svec and coworkers.25–28 The first step of the multi-step swelling method is associated with the activation of template uniform particles (usually PS) formed by either emulsion or dispersion polymerization processes. The activation of the template particles is accomplished by swelling the particles dispersed in aqueous phase with emulsion droplets of a swelling solvent, e.g. dibutyl phthalate or 1-chlorodecane. The first swelling step stimulates the swelling of the template particles in the subsequent steps. When the activation-swelling step is completed, the second swelling step takes place by introducing the slightly enlarged template particles to monomers, initiator, and porogens. This can be done in one step, or through sequential addition of each component. The initiator can be added in the first or second swelling step. Polymerization of the monomers within the uniformly swollen particles can then be induced by increasing the temperature. An alternative swelling method was invented by Okubo et al., and was named: ‘‘the dynamic swelling method’’.29,30 According to this method, uniform PS template particles can be swollen enormously, while maintaining their uniformity, by slow, continuous, dropwise addition of water into an ethanol/water medium containing the template particles and hydrophobic monomer/s and initiator (e.g. styrene and benzoyl peroxide [BP]). Polymerization can then be performed, as previously described, by increasing the temperature. According to the dynamic swelling method, there is no need to use a swelling solvent, and the process can be performed in a one-step procedure. A new method for preparing particles of narrow size distribution and controlled properties, i.e. surface area, based on a single-step swelling process of template uniform microspheres was recently published by Margel and coworkers.31–33 According to this process, the swelling of the template particles with the initiator and monomer/s via a swelling solvent is accomplished in a single step, in contrast to the multi-swelling steps where the swelling with these reagents is accomplished in two or more steps. The present article is different from the previous ones in that the single-step swelling process was used for entrapping TIBEE rather than acrylate monomers within uniform PS template microspheres. Evaporation of the swelling solvent, after completing the swelling process, produces radiopaque PS microspheres of narrow size distribution. TIBEE, rather than 2,3,5-triiodobenzoic acid (a parent compound for synthesis of various contrast agent

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derivatives used in medicinal X-ray imaging), was chosen in this study to be used as a model X-ray contrast agent for the encapsulation process. The reason for this is the fact that 2,3,5-triiodobenzoic acid is not suitable for the encapsulation process described here, since this acid is slightly soluble in water and does not dissolve significantly in methylene chloride. On the other hand, the ethyl ester derivative of 2,3,5-triiodobenzoic acid, TIBEE, is relatively more hydrophobic, soluble in methylene chloride, and thereby suitable for the described encapsulation process.

EXPERIMENTAL Materials The following analytical grade chemicals were purchased from Aldrich, and were used without further purification: 2,3,5-triiodobenzoic acid, ethyl acetate, magnesium sulfate, concentrated sulfuric acid, BP (98%), sodium dodecyl sulfate (SDS), polyvinylpyrrolidone (PVP, m.w. 360,000), ethanol (HPLC), 2-methoxy ethanol (HPLC), and methylene chloride (HPLC). Styrene (Aldrich, 99%) was passed through activated alumina (ICN) to remove inhibitors before use. Water was purified by passing deionized water through Elgastat Spectrum reverse osmosis system (Elga, High Wycombe, UK).

Methods Synthesis of TIBEE (Scheme 1) Concentrated H2SO4 (4 mL) was added to a solution of 2,3,5-triiodobenzoic acid (20 g, 40 mmol) in 250 mL of ethanol. The reaction mixture was then refluxed overnight. The resulting mixture was evaporated, and the residue was then dissolved in 250 mL of ethyl acetate. The organic phase was washed with H2O, saturated NaHCO3, and brine. The organic layer was then dried over MgSO4, filtered, and evaporated to produce the desired orange solid product (yield 83%). 1 H NMR (CDCl3) d 8.28 (d, 1H, H6, J ¼ 2 Hz), 7.71 (d, 1H, H4, J ¼ 2 Hz), 4.38 (q, 2H, CH2, J ¼ 7.2 Hz), 1.39 (t, 3H, Me, J ¼ 7.2 Hz). 13C NMR (CDCl3) d 165.99 (CO), 148.57 (6),

Scheme 1.

Synthesis of TIBEE.

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141.69 (ipso), 139.79 (4), 133.20 (2), 106.49 (3), 93.66 (5), 62.50 (CH2), 14.05 (Me). MS (FABþ): m/z 529 (MHþ, 100), 483 (M-OCH2CH3, 46). Elemental analysis, calculated: C, 20.47; H, 1.33; O, 6.06; I, 72.12; experimental: C, 20.77; H, 1.35; O, 5.37; I, 72.51.

Synthesis of PS template microspheres PS template microspheres of narrow size distribution were prepared according to a procedure similar to that described in the literature.26–28 Briefly, these microspheres were synthesized in a three-neck round-bottom flask equipped with a condenser and immersed in a constant temperature silicone oil bath at a preset temperature. In a typical experiment, PS microspheres with an average diameter of 2.3 6 0.2 lm were formed by introducing into the reaction flask (1 L) a solution containing PVP (3.75 g, 1.5% w/v of total solution) dissolved in a mixture of ethanol (150 mL) and 2-methoxy ethanol (62.5 mL). The temperature of the mechanically stirred solution (200 rpm) was then preset to 738C. Nitrogen was bubbled through the solution for 15 min to exclude air, and then a blanket of nitrogen was maintained over the solution during the polymerization period. A deaerated solution containing the initiator BP (1.5 g, 0.6% w/v of total solution) and styrene (37.5 mL, 16% w/v of total solution) was then added to the reaction flask. The polymerization reaction continued for 24 h, and was then stopped by cooling to room temperature. The microspheres formed were washed by extensive centrifugation cycles with ethanol and then with water. The particles were then dried by lyophilization. Uniform PS microspheres of sizes ranging between 0.2 and 5.0 lm were prepared similarly by changing polymerization parameters, e.g., monomer or initiator concentration.

Preparation of radiopaque PS microspheres by a single-step swelling process In a typical experiment, PS template microspheres of 2.3 6 0.2 lm were swollen up to 4.2 6 0.2 lm by methylene chloride containing TIBEE by adding, to a 20 mL vial, 10 mL SDS aqueous solution [1.5% (w/v)] and 1.6 mL of the swelling solvent containing 0.125 g TIBEE. Emulsion droplets of the swelling solvent were then formed by sonication (Sonics and Materials, model VCX-750, Ti-horn 20 kHz) of the mixture at 48C for 60 s. An aqueous suspension (3.5 mL) of the PS template microspheres (7% w/v) was then added to the stirred methylene chloride emulsion. After the swelling of the PS particles was completed, and the mixture did not contain any small emulsion droplets of the swelling solvent, as verified by optical microscopy, evaporation of the entrapped methylene chloride was performed by shaking the open vial containing the aqueous mixture of swollen particles for 4 h at 408C. The resulting uniform radiopaque PS microspheres were then washed by several centrifugation cycles with water, and then lyophilized. In several cases, where, in addition to the radiopaque PS microspheres, TIBEE needle crystals were also observed by light microscope observations, removal of the needles was performed by filtration of the aqueous suspenJournal of Biomedical Materials Research Part A DOI 10.1002/jbm.a

GALPERIN, MARGEL, AND MARGEL

sion of particles through Whatman filter paper of 41 mesh. The TIBEE crystal-free radiopaque PS aqueous suspension was then washed by several centrifugation cycles with water, and lyophilized.

Characterization 1

H and 13C NMR spectra were obtained on Bruker DPX-300 spectrometer. For chloroform-d chemical shifts are expressed in ppm downfield from tetramethylsilane used as internal standard. Mass spectra were obtained on a Finnigan 4021 spectrometer (FAB, fast atom bombardment). Optical microscope pictures of the microspheres dispersed in the liquid phase were obtained with an Olympus microscope, model BX51. Particle average size and size distribution were determined by measuring the diameters of more than 100 particles on optical micrographs with the image analysis software AnalySIS Auto (Soft Imaging System GmbH, Germany). Fourier transform infrared (FTIR) analysis was performed with a Bomem FTIR spectrophotometer, model MB100 (Hartman & Braun). The analysis was performed with 13-mm KBr pellets that contained 2 mg of the detected material and 198 mg KBr. The pellets were scanned 200 times at 4 cm1 resolutions. Elemental analysis was performed using an elemental analysis instrument, model EA1110 (CE Instruments). Iodine elemental analysis was performed by the Microanalysis Lab, Institute of Chemistry, The Hebrew University of Jerusalem, Jerusalem. Surface elemental analysis was obtained by X-ray photoelectron spectroscopy (XPS), model AXIS-HS (Kratos Analytical, England), using Al Ka lines, at 109 Torr, with a take off angle of 908. The reported values of both the XPS and elemental analysis are an average of measurements performed on at least three samples of each of the tested particles, and have a maximum error of about 10 and 2%, respectively. Thermogravimetric analysis (TGA) was performed with a TC15 system equipped with TGA, model TG-50, Mettler Toledo. The analysis was performed with 6 mg of dried samples in a dynamic nitrogen atmosphere (200 mL/min) with a heating rate of 108C/min. Surface morphology was characterized with JEOL Scanning Electron Microscope (SEM), model JSM-840. The radiopacity of the microspheres was demonstrated by a CT scanner (MARCONI, HeliCAT II). Quantitation (in Hounsfield units) of the opacification of the microspheres was performed by the image-processing software CDP DiagNet v. 5.55. The measured and calculated percentage of TIBEE entrapped in the PS microspheres were calculated using the following equations: %TIBEEmeasured ¼

% Iodine 100 72

ð1Þ

%TIBEEcalculated ¼

WTIBEE 100 ð0:24 þ WTIBEE Þ

ð2Þ

where % iodine is obtained from the elemental iodine analysis, 72 is the percentage of iodine in TIBEE, WTIBEE is the weight (g) of the TIBEE that was used in the swelling


Figure 1.

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Synthesis of radiopaque PS microspheres by a single-step swelling process.

process, and 0.24 is the weight (g) of the PS template microspheres that were used in the swelling process.

RESULTS AND DISCUSSION Figure 1 describes the fundamental process of preparation of the radiopaque PS microspheres via a single-step swelling of the PS template microspheres with methylene chloride containing TIBEE, followed by evaporation of the methylene chloride after completion of the swelling process. The PS template microspheres have a size and size distribution of 2.3 6 0.2 lm. As a consequence of their swelling with 1.6 mL methylelne chloride containing 0.12 g TIBEE, their size distribution was maintained, while their diameter increased from 2.3 6 0.2 lm to 4.2 6 0.2 lm. Evaporation of the methylene chloride to form the final radiopaque PS microspheres leads to shrinking of the swollen particles, more or less, to the initial size and size distribution of the template microspheres, as shown in Figure 2. Figure 3 depicts FTIR spectra of the PS template microspheres (A), TIBEE (B), and the radiopaque PS microspheres prepared by the single-step swelling of

Figure 2. A light microscope picture of the radiopaque PS microspheres formed by swelling of the particles with methylene chloride containing TIBEE, followed by evaporation of the swelling solvent. The swelling and evaporation processes were accomplished according to the Experimental section, with 1.6 mL methylene chloride containing 0.12 g TIBEE.

the template microspheres (C). Figure 3(A) shows a typical IR spectrum of PS: the transition frequencies 1492 and 3000–3100 cm1 correspond to the aromatic CH stretching bands, 2849 and 2922 cm1 correspond to the CH2 stretching bands, and 700 cm1 corresponds to vibrational band of CC. Figure 3(B) shows an IR spectrum of TIBEE: the transition frequencies at 1300 cm1 correspond to the CO stretching band and 1732 cm1 correspond to carbonyl CO stretching band. Figure 3(C) shows the IR spectrum of the radiopaque PS microspheres. This spectrum is composed of peaks belonging both to PS (700, 1492, 3000 cm1, etc.) and to TIBEE (1300, 1732 cm1), and thus demonstrates the presence of the X-ray contrast agent in the PS microspheres. Figure 4 illustrates TGA thermograms of the PS template microspheres (A), TIBEE (B), and the radiopaque PS microspheres prepared by the single-step

Figure 3. FTIR spectra of the template PS microspheres (A), TIBEE (B), and the radiopaque PS microspheres (C). The PS template microspheres and the TIBEE were prepared according to the Experimental section. The radiopaque PS microspheres were synthesized according to the Experimental section, with 1.6 mL methylene chloride containing 0.12 g TIBEE. Journal of Biomedical Materials Research Part A DOI 10.1002/jbm.a

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Figure 4. TGA thermograms of the PS template microspheres (A), TIBEE (B) and the radiopaque PS microspheres (C). The PS template microspheres and the TIBEE were prepared according to the Experimental section. The radiopaque PS microspheres were synthesized according to the Experimental section, with 1.6 mL methylene chloride containing 0.12 g TIBEE.

swelling of the template microspheres (C). The TGA thermogram of the PS template microspheres [Fig. 4(A)] indicates almost 100% weight loss at temperatures between 350 and 4508C, due to the decomposition of the PS polymer. The TGA thermogram of the TIBEE [Fig. 4(B)] shows almost 100% weight loss at temperatures between 170 and 3008C, due to the decomposition of TIBEE. The TGA thermogram of the radiopaque PS microspheres [Fig. 4(C)] represents the thermal behavior of both the PS microspheres and the TIBEE. This figure indicates two slopes: the first one, started at 1708C, belongs to the decomposition of TIBEE, and the second one, started at 3508C, belongs to the decomposition of the PS microspheres.

Table I demonstrates the influence of the weight ratio [TIBEE]/[PS] on the % TIBEE entrapped in the PS microspheres, as calculated from elemental iodine analysis. This table indicates that upon increasing the weight ratio [TIBEE]/[PS] from 0.12 to 0.16, 0.25, and 0.5, the weight % TIBEE entrapped in the PS microspheres increased from 11.3% to 14.9, 20.5, and 32%, respectively. On the other hand, there is no significant change in the weight % TIBEE entrapped in the PS particles (31%) when the weight ratio [TIBEE]/ [PS] increased from 0.5 to 1.0 or 2.0. Table I also demonstrates that when the weight ratio [TIBEE]/[PS] is between 0.12 and 0.5, the measured and the calculated weight % TIBEE entrapped in the PS microspheres are similar, while at weight ratios [TIBEE]/[PS] 1.0 and 2.0, the calculated weight % TIBEE entrapped in the PS microspheres (50.0 and 66.6, respectively) is significantly higher than that of the measured values (30.7 and 31.0, respectively). The calculated values of the weight % TIBEE entrapped in the PS microspheres are based on the assumption that all the TIBEE that was used for the swelling process was entrapped in the PS particles. Table I, therefore, shows that all the TIBEE used for the swelling process at weight ratios [TIBEE]/[PS] up to 0.5 was entrapped in the PS particles. On the other hand, there is no indication of additional permeation of TIBEE to the PS particles when the weight ratio [TIBEE]/ [PS] increased from 0.5 to 1.0 or 2.0. Table I therefore illustrates that under the experimental conditions, the maximum capacity of TIBEE in the PS particles is 32%. In addition to the results shown in Table I, light microscopy observations showed that at weight ratio [TIBEE]/[PS] below 0.5, only uniform microspheres exist in the aqueous suspension. On the other hand, when the weight ratio [TIBEE]/[PS] is above 0.5, in addition to the microspheres, needle crystals were also observed. Also, the concentration

TABLE I Influence of the Weight Ratio [TIBEE]/[PS] on the % of TIBEE Entrapped in the PS Microspheresa Weight % TIBEE in the PS Microspheres [TIBEE] (g)

[TIBEE]/[PS] (w/w)

Weight % Iodineb

Measuredc

Calculatedd

0.03 0.04 0.05 0.06 0.08 0.12 0.24 0.48

0.12 0.16 0.20 0.25 0.33 0.50 1.0 2.0

8.2 10.8 12.2 14.8 18.2 23.0 22.2 22.5

11.3 14.9 16.9 20.5 25.2 32.0 30.7 31.0

11.1 14.3 16.9 20.0 25.0 33.3 50.0 66.6

a The radiopaque PS microspheres were prepared according to the Experimental section with 1.6 mL methylene chloride containing different amounts of TIBEE. b Data from elemental analysis. c Calculated from iodine elemental analysis according to the Experimental section. d The calculations were performed according to the Experimental section, and are based on the assumption that all the TIBEE that was used in the swelling process was entrapped in the radiopaque PS microspheres.



of the needle crystals increased with increasing weight ratio [TIBEE]/[PS]. Figure 5, for example, illustrates a typical light microscopy picture of microspheres with the needle crystals obtained at weight ratio [TIBEE]/[PS] of 2.0. The removal of the needles from the suspensions of the radiopaque microspheres was easily accomplished by filtration through Whatman filter paper of 41 mesh. Elemental analysis, FTIR, MS, and NMR clearly demonstrated that these needle crystals are composed of pure TIBEE. It should be noted that TIBEE needle crystals were observed by light microscopy only after the evaporation of methylene chloride, but not during the swelling process. These results may indicate that the excess TIBEE, which does not penetrate to the PS microspheres during the swelling process, was adsorbed onto the surface of the microspheres. Evaporation of the methylene chloride, after completing the swelling process, probably detached the loosely surface adsorbed TIBEE into the aqueous phase where the TIBEE can then grow to needle shape. The bulk and surface composition of the radiopaque particles prepared at different [TIBEE]/[PS] weight ratios were measured by elemental and XPS iodine analysis, respectively. Table II demonstrates, as expected, that the % iodine content of the radiopaque microspheres increased as this ratio increased. For example, at weight ratios 0.12, 0.25, and 0.5, the iodine content of the radiopaque particles is 8.2, 14.8, and 23.0%, respectively. On the other hand, the % iodine at the surface of the radiopaque particles is significantly lower (1.6–1.8%), and does not depend on the weight ratio [TIBEE]/[PS]. These results, particularly those presented by the XPS, may indicate

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TABLE II Weight % Iodine on the Surface and the Bulk of the Radiopaque PS Microspheres Formed at Different Weight Ratio of [TIBEE]/[PS]a Weight % Iodine in the PS Microspheres [TIBEE]/[PS] (w/w)

Elemental Analysis

XPS

0.12 0.16 0.20 0.25 0.33 0.50

8.2 10.8 12.2 14.8 18.2 23.0

1.6 1.8 1.8 1.8 1.6 1.6

a

The radiopaque PS microspheres were prepared according to the Experimental section with 1.6 mL methylene chloride containing different amounts of TIBEE.

low adsorption of TIBEE on the surface of the radiopaque PS particles, independent of the weight ratio [TIBEE]/[PS]. Figure 6, indeed, demonstrates by SEM pictures, the smooth surface of the PS template microspheres [Fig. 6(A)] and the bumpy surface, due to the surface adsorbed TIBEE, of the radiopaque PS microspheres formed at weight ratio [TIBEE]/[PS] of 0.5 [Fig. 6(B)]. Figure 7 represents the X-ray visibility of the PS template microspheres (A) and the radiopaque PS microspheres containing different amounts of TIBEE: 11.3 (B), 20.5 (C), and 32.0% (D). The PS template microspheres (A) are transparent to the X-ray irradiation, while the PS microspheres containing TIBEE [Fig. 7(B– D)] show excellent radiopaque nature. Figure 7(B–D), as expected, clearly demonstrates that the X-ray visibility increases as the % TIBEE encapsulated in the PS microspheres increases. Table III quantitates (in Hounsfield Units) the opacification of the PS template microspheres and the PS microspheres containing different amounts of TIBEE. Table III illustrates that the intensity of the CT signal of the PS template microspheres is 700 6 50 HU, similar to the CT number of air, 1000 HU.34 This CT number of the PS template microspheres indicates that these microspheres are transparent to X-ray irradiation, as is air. Table III shows the significant increase in the intensity of CT signals as the weight % TIBEE in the PS microspheres increases. For example, the intensity of the CT signals of the radiopaque PS microspheres containing 11.3, 20.5, and 32.0% TIBEE are 556 6 66, 1789 6 264, and 3049 6 137 HU, respectively.

CONCLUSIONS Figure 5. A light microscope picture illustrating the radiopaque PS microspheres and the needle crystals prepared at weight ratio [TIBEE]/[PS] of 2.0. PS radiopaque microspheres were prepared according to the Experimental section, with 1.6 mL methylene chloride containing 0.48 g TIBEE.

The present article describes a novel method to prepare model radiopaque uniform microspheres composed of TIBEE encapsulated in PS template microspheres of 2.3 6 0.2 lm diameter. This method Journal of Biomedical Materials Research Part A DOI 10.1002/jbm.a

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TABLE III Opacification of the PS Microspheres Containing Different Amounts of TIBEEa Weight % TIBEE in the PS Microspheres 0 (PS template microspheres only) 11.3 14.9 20.5 25.2 32.0

Opacification (HU) 700 556 1244 1789 2588 3049

6 6 6 6 6 6

50 66 232 264 141 137

a

The radiopaque microspheres were prepared according to the Experimental section with 1.6 mL methylene chloride containing different amounts of TIBEE. Quantitation (in Hounsfield Units) of the opacification of the microspheres was performed by the image-processing software CDP DiagNet v. 5.55.

Figure 6. SEM pictures of the template PS microspheres (A) and the radiopaque PS microspheres (B). The radiopaque PS microspheres were prepared according to the Experimental section, with 1.6 mL of methylene chloride containing 0.12 g TIBEE.

phobic particles dispersed in an aqueous continuous phase. For future studies we plan to extend the present work concerning radiopaque particles to uniform PS template particles of different sizes (0.1–10 lm) as well as particles others than PS, e.g., biodegradable particles from poly(methyl methacrylate) and polylactic acid. These radiopaque particles of different nature and diameter will then be tested for different X-ray imaging needs, e.g., blood pool, body organs, embolization, dental composition, implants, protheses, and nanocomposites. References

is based on a single-step swelling process of the PS microspheres dispersed in an aqueous continuous phase with methylene chloride (as a swelling solvent) containing TIBEE. After completion of the swelling process, the methylene chloride is evaporated in order to lock the TIBEE in the PS particles. This new encapsulation model may be used as a general concept for encapsulation of other hydrophobic compounds such as drugs, fertilizers, contrast agents, etc. within hydro-

Figure 7. X-ray visibility of the template PS microspheres (A) and the radiopaque PS microspheres containing different amounts of TIBEE: 11.3 (B), 20.5 (C) and 32.0% (D). The radiopaque microspheres were prepared according to the Experimental part, with 1.6 mL methylene chloride containing different amounts of TIBEE. The X-ray images were obtained with a CT scanner. Journal of Biomedical Materials Research Part A DOI 10.1002/jbm.a

1. Torchilin VP. PEG-based micelles as carriers of contrast agents for different imaging modalities. Adv Drug Deliv Rev 2002;54:235–252. 2. Torchilin VP. Polychelating amphiphilic polymers to optimize the concentration of contrast agents used in medical diagnostic imaging. Chemtech 1999;29:27–34. 3. Torchilin VP. Polymeric contrast agents for medical imaging. Curr Pharm Biotechnol 2000;1:183–215. 4. Masahiko O, Takeshi Y, Tomohiro A, Hiroki U, Tadashi K, Seiichi H, Tatsuo K. Synthesis and properties of radiopaque polymer hydrogels. II. Copolymers of 2,4,6-triiodophenyl- or N-(3-carboxy-2,4,6-triiodophenyl)-acrylamide and p-styrene sulfonate. J Mol Struct 2002;602/603:17–28. 5. Horak D, Metalova M, Svec F, Drobnik J, Katal J, Borovicka M, Adamyan AA, Voronkova OS, Gumargalieva KZ. Hydrogels in endovascular embolization. III. Radiopaque spherical particles, their preparation and properties. Biomaterials 1987;8:142–145. 6. Jayakrishnan A, Thanoo BC, Rathinam K, Mohanty M. Preparation and evaluation of radiopaque hydrogel microspheres based on PHEMA/iothalamic acid and PHEMA/iopanoic acid as particulate emboli. J Biomed Mater Res 1990;24:993–1004. 7. Mottu F, Rufenacht DA, Laurent A, Doelker E. Iodine-containing cellulose mixed esters as radiopaque polymers for direct embolization of cerebral aneurysms and arteriovenous malformations. Biomaterials 2002;23:121–131. 8. Chithambara TB, Jayakrishnan A. Barium sulphate-loaded p(HEMA) microspheres as artificial emboli: Preparation and properties. Biomaterials 1990;11:477–481. 9. Chithambara TB, Jayakrishnan A. Tantalum-loaded silicone microspheres as particulate emboli. J Microencapsul 1991;8:95– 101.


10. Chithambara TB, Sunny MC, Jayakrishnan A. Tantalum-loaded polyurethane microspheres for particulate embolization: Preparation and properties. Biomaterials 1991;12:525–528. 11. Horak D, Metalova M, Rypacek F. New radiopaque polyHEMA-based hydrogel particles. J Biomed Mater Res 1997;34: 183–188. 12. Benzina A, Kruft MAB, van der Veen FH, Bar FH, Blezer R, Lindhout T, Koole LH. A versatile three-iodine molecular building block leading to new radiopaque polymeric biomaterials. J Biomed Mater Res 1996;32:459–466. 13. Jayakrishnan A, Chithambara TB. Synthesis and polymerization of some iodine-containing monomers for biomedical application. J Appl Polym Sci 1992;44:743–748. 14. Kruft MAB, Benzina A, Bar FH, van der Veen FH, Bastiaansen CWM, Blezer R. Studies of two new radiopaque polymeric biomaterials. J Biomed Mater Res 1994;28:1259– 1266. 15. Nirmala RJ, Juby P, Jayakrishnan A. Polyurethanes with radiopaque properties. Biomaterials 2006;27:160–166. 16. Moszner N, Saiz U, Klester AM, Rheinberger V. Synthesis and polymerization of hydrophobic iodine-containing methacrylates. Die Angew Makromol Chem 1995;224:115–123. 17. Davy KWM, Anseau MR. X-ray opaque methacrylate polymers for biomedical application. Polym Int 1997;43: 143–154. 18. Davy KWM, Anseau MR, Berry C. Iodinated methacrylate copolymers as X-ray opaque denture base acrylics. J Dent 1997; 25:499–505. 19. Saralidze KS, Aldenhoff BJ, Knetsch LW, Koole LH. Injectable polymeric microspheres with X-ray visibility. Preparation, properties, and potential utility as new traceable bulking agents. Biomacromolecules 2003;4:793–798. 20. Vasquez B, Ginebra MP, Gil FG, Planell JA, Bravo AL, Roman JS. Radiopaque acrylic cements prepared with a new acrylic derivative of iodo-quinoline. Biomaterials 1999;20:2047– 2053. 21. Emans PJ, Saralidze K, Knetsch LW, Gijbels JJ, Kuijer R, Koole LH. Development of new injectable bulking agents: Biocompatibity of radiopaque polymeric microspheres studies in a mouse model. J Biomed Mater Res A 2005;73:430–436. 22. Ugelstad J. Swelling capacity of aqueous dispersions of oligomer and polymer substances and mixtures thereof. Makromol Chem 1978;179:815–817. 23. Ugelstad J, Mork PC, Nordhuus I, Mfutakamba H, Soleimany E. Thermodynamics of swelling. Preparation and application of some composite, monosized polymer particles. Macromol Chem 1985;10/11 (Suppl):215–234.

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24. Cheng CM, Micale FJ, Vanderhoff JW, El-Aasser MS. Synthesis and characterization of monodisperse porous polymer particles. J Polym Sci Part A: Polym Chem 1992;30:235–244. 25. Hosoya K, Frechet JMJ. Influence of the seed polymer on the chromatographic properties of size monodisperse polymeric separation media prepared by a multi-step swelling and polymerization method. J Polym Sci Part A: Polym Chem 1993; 31:2129–2141. 26. Smigol V, Svec F, Hosoya K, Wang Q, Frechet JMJ. Monodisperse polymer beads as packing material for high-performance liquid chromatography. Synthesis and properties of monodisperse polystyrene and methacrylate latex seeds. Angew Makromol Chem 1992;195:151–164. 27. Smigol V, Svec F. Synthesis and properties of uniform beads based on macroporous copolymer glycidyl methylacrylate ethylene dimethylacrylate: A way to improve separation media for HPLC. J Appl Polym Sci 1992;46:1439–1448. 28. Liang YC, Svec F, Frechet JMJ. Preparation and functionalization of reactive monodisperse macroporous poly(chloromethylstyrene-co-styrene-co-divinylbenzene) beads by a staged templated suspension polymerization. J Polym Sci Part A: Polym Chem 1997;35:2631–2643. 29. Okubo M, Ise E, Yamashita T. Synthesis of greater than 10-lmsized, monodispersed polymer particles by one-step seeded polymerization for highly monomer-swollen polymer particles prepared utilizing the dynamic swelling method. J Appl Polym Sci 1999;74:278–285. 30. Okubo M, Shiozaki M. Production of micron-size monodisperse polymer particles by seeded polymerization utilizing dynamic swelling method with cooling process. Polym Int 1993;30:469–474. 31. Kedem M, Margel S. Synthesis and characterization of micrometer-sized particles of narrow size distribution with chloromethyl functionality on the basis of single-step swelling of uniform polystyrene template microspheres. J Polym Sci Part A: Polym Chem 2002;40:1342–1352. 32. Boguslavsky L, Margel S. Synthesis and characterization of micrometer-sized homo and composite polyacrylonitrile particles of narrow size distribution on the basis of a single-step swelling of uniform polystyrene template microspheres. J Polym Sci Part A: Polym Chem 2004;42:4847–4861. 33. Akiva U, Margel S. Surface-modified hemispherical polystyrene/ polybutyl methacrylate composite particles. J Colloid Interface Sci 2005;288:61–70. 34. Wolf GL. Targeted delivery of imaging agents: an Overview. In: Torchilin VP, editor. Handbook of Targeted Delivery of Imaging Agents. Boca Raton: CRC; 1995, p 11–12.
