05Solubility enhancement of ibu

J Sol-Gel Sci Technol (2014) 72:627–636 DOI 10.1007/s10971-014-3485-3

ORIGINAL PAPER

Solubility enhancement of ibuprofen using tri-ureasil-PPO hybrid: structural, cytotoxic, and drug release investigation Beatriz B. Caravieri • Pollyanna F. de Oliveira • Ricardo A. Furtado • Denise C. Tavares • Eduardo J. Nassar • Katia J. Ciuffi • Eduardo F. Molina

Received: 3 July 2014 / Accepted: 20 August 2014 / Published online: 27 August 2014 Ó Springer Science+Business Media New York 2014

Abstract Herein, we used tri-ureasil organic–inorganic hybrid material (tU5000) in order to enhance the solubility of nonsteroidal anti-inflammatory drugs and fine tuning the drug delivery profile. For the first time, we used tU5000 as a film-forming agent in order to provide an alternative vehicle for transdermal drug delivery systems which the cell viability of practically 100 % for the highest and the lowest tested concentrations of pure tU5000 indicated that the material was not cytotoxic. The physicochemical properties of the tU5000 drug carrier and drug-loaded hybrids were systematically studied using powder X-ray diffraction, differential scanning calorimetry, small-angle X-ray scattering, and Fourier-transform infrared spectroscopy. The structural changes of tU5000 as well as the relationships between the drug content and in vitro drug release behaviors were investigated. The results showed that the ibu molecules were homogeneously distributed in the tU5000 xerogels contributing to fine-tuning the drug delivery profile. Considering the ability to incorporated high drug content, simple and mild preparation procedure by one-pot sol–gel route, high stability of the materials, sustained-release property, this class of hybrid based on polymers and inorganic compounds may have potential applications in the design of pharmaceutical formulation as ophthalmic (contact lenses), transdermal (patches) and implantable (soft tissue) drug delivery systems.

Electronic supplementary material The online version of this article (doi:10.1007/s10971-014-3485-3) contains supplementary material, which is available to authorized users. B. B. Caravieri  P. F. de Oliveira  R. A. Furtado  D. C. Tavares  E. J. Nassar  K. J. Ciuffi  E. F. Molina (&) Department of Chemistry, Universidade de Franca, Av. Dr. Armando Salles Oliveira 201, Franca, SP 14404-600, Brazil e-mail: [email protected]

Keywords Tri-ureasil hybrid  Drug delivery  Sustained release  Structural characterization  Ibuprofen

1 Introduction Over the past decades, the use of organic–inorganic (O–I) hybrid structures as drug carriers has become increasingly attractive. The properties of these materials do not simply correspond to the sum of the contributions of their individual components; in fact, they are the result of the strong synergy created in extensive hybrid interfaces [1–3]. O–I hybrid structures are simple to process and amenable to design on the molecular scale, enabling the creation of multifunctional materials [4–6]. For example, it is possible to incorporate inorganic clusters or nanoparticles with specific optical, electronic, or magnetic features into organic polymer matrices, which paves the way for developing advanced drug delivery systems [7, 8]. In the pharmaceutical domain, nonsteroidal antiinflammatory drugs (NSAIDs) display analgesic, antipyretic, and anti-inflammatory activities; [9, 10] they are well known for their efficient and unsurpassed therapeutic effects in numerous chronic and acute conditions [11, 12]. However, NSAIDs of interest such as ibuprofen (ibu) and naproxen have relatively short half-life in plasma and are poorly water-soluble, [13, 14] making it necessary to administer high systemic doses of these medications [15, 16]. Unfortunately, their repeated administration culminates in severe gastrointestinal side effects such as stomach ulceration, bleeding, and perforation, because in the body the drug reaches targeted and nontargeted sites [17]. Several research groups have investigated mesostructured silica materials as drug delivery systems for NSAIDs [18–21]. Although these systems exhibit sustained-release

123

628

J Sol-Gel Sci Technol (2014) 72:627–636

Scheme 1 Structure of the organic–inorganic tri-ureasil hybrid tU5000 (left) and the ibuprofen drug (right)

properties, their drug loading capacity is relatively low, which causes burst release and short-term release. Interestingly, modification of mesostructured silica leads to much stronger electrostatic interaction between the carboxylic acid and the amine group (modified MCM-41), which improves drug loading and slows down ibu release [22]. To achieve higher bioavailability and solubility of poorly water-soluble NSAIDs, researchers have added surface-active agents to these drugs, which affords watersoluble salts, and increases the drug particles wettability and micronization [23, 24]. However, this method is not always successful. In recent years, research groups have developed and used a number of macromolecules to enhance NSAIDs solubility and limit their side effects [25]. Drug adsorption and distribution in such systems rely on site specificity, resistance to degradation, and minimization of side effects, which depend on the modifications performed on the carrier macromolecule [26]. Senel and Koc¸ have already reported that a new class of polypropylene oxide-cored PAMAM dendrimers increased the solubility, drug loading, and sustained drug release of NSAIDs such as ibuprofen, ketoprofen, and diflunisal. [27] The concentration, core size, and generation of the polypropylene oxide PPO cored PAMAM determined improvement in these parameters. In a previous paper, our group described the synthesis of the tri-ureasil O–I hybrid (labeled tU5000) and demonstrated its unique potential as drug carrier, photochromic device, and dye adsorbent [28]. However, the question of existence or not of chemical interactions between tU5000 and drug is still open. Thus, the goal of this work was to study the interactions between ibu drug and tU5000 hybrid material, aiming to understand and to optimize the drug delivery behavior. The structural changes of tU5000 carriers as well as the relationships between the drug content and in vitro drug release behaviors were in-deep investigated. The physicochemical properties of the tU5000 drug carrier and drug-loaded hybrids were systematically studied using powder X-ray diffraction (XRD), differential

123

scanning calorimetry (DSC), small-angle X-ray scattering (SAXS), Fourier-transform infrared (FTIR) spectroscopy, and we will assess the drug release kinetics by in situ UV– vis spectroscopy. Additionally, we use the XTT assay to examine the viability of human lung fibroblasts cells in the presence of pure tU5000 (See Scheme 1 for the structure of tU5000 hybrid matrix and ibu drug) and for the first time we used tU5000 as a film-forming agent in order to provide an alternative vehicle for transdermal drug delivery systems.

2 Materials and methods 2.1 Materials Glyceryl poly(oxypropylene) triamine with a molecular weight of 5000 g mol-1 (JeffamineÒ T-5000) was donated by Herminio Muchon Filho, Account Manager, Huntsman Performance Products, Brazil. (3-Isocyanatopropyl)triethoxysilane (ICPTES), ethanol (CH3CH2OH), tetrahydrofuran (THF), and ibuprofen sodium salt (ibu, a-methyl-4(isobutyl)phenylacetic acid, oral lethal dosage (LD50) of 636 mg kg-1 for rats (Material Safety Data Sheet Ibuprofen MSDS)) (ibu) were purchased from Sigma-Aldrich. All these chemicals were used as received. 2.2 Tri-ureasil hybrid synthesis The tri-ureasil hybrid, represent by the notation tU5000, were prepared according to the conventional method described in the previous work [28]. The ureasil cross-link agent was covalently bound to both ends of the macromer by reacting the NH2 groups of the Jeffamine T5000 with the –N=C=O group of ICPTES at a 1:3 molar ratio [28, 29]. These reagents were refluxed in THF for 24 h with magnetic stirring. Then, THF was eliminated by evaporation at 60 °C, to give a viscous precursor. In a second step, silanol moieties (–Si(OH)3) were generated. Subsequent condensation reactions furnished ureasil cross-linking siloxane

J Sol-Gel Sci Technol (2014) 72:627–636

nodes (see synthetic scheme of the reaction in ref. 28). All the samples were prepared under the same conditions. Addition of 4 mL of a water–ethanol mixture (0.04 v/v) containing 165 mg kg-1 HCl (oral toxicity limit 900 mg kg-1) to 1.5 g of the precursor induced –(SiOCH2CH3)3 hydrolysis, which gave the final product tU5000. To obtain the drug-loaded sample (1, 3, 5, 10, 15, 20, 30, and 50 wt% ibu-loaded tU5000), ibu powder was dissolved in water–ethanol hydrolysis solution and added to tU5000. Cylindrical monolithic tU5000 xerogels with diameter and height of approximately 20 and 3 mm, respectively, were obtained after drying under vacuum at 70 °C for 24 h. 2.3 Characterization 2.3.1 Powder X-ray diffraction (XRD) XRD patterns were recorded on a Rigaku MiniFlex II ˚) Desktop X-ray Diffractometer using Cu Ka (k = 1.54 A radiation at a scan speed of 0.04° s-1. 2.3.2 Differential scanning calorimetry (DSC) DSC measurements were carried out using a TA Instrument model Q100. Disk sections of approximately 20 mg were cut off from the hybrid matrix and sealed in a 40-mL hermetic aluminum pan. Each sample was heated from -80 to 350 °C at 10 °C min-1. The purge gas was highpurity nitrogen (N2) supplied at a flow rate of 75 cm3 min-1.

629

detector distance was 700 mm, and the time necessary for each data collection was 30 s. The beam center was calibrated using silver behenate with the primary reflection peak at 1.076 nm-1. 2.4.2 In vitro ibuprofen release The ibu release from the ibu-loaded tU5000 samples were monitored by UV–vis spectroscopy. To this end, about 0.1 g of monolithic ibu-loaded tU5000 was immersed into 100 mL of phosphate buffered saline (PBS, pH 7.4). The release assays were carried out thermostatically at 37 °C and stirred (70 rpm). The water volume considered here was about 20 times higher than that required for the solubilization of all the ibu loaded in the xerogel. All the experiments were conducted on three independent samples; the results are the average of the data. The amount of ibu in the medium was determined by UV spectroscopy, kmax = 221 nm. UV–vis absorption data were recorded within the wavelength range of 190–500 nm using an Agilent Technologies Cary 60 dual beam spectrophotometer fitted with a fiber optic coupler equipped with a solarization-resistant immersion probe. A 2-mm solution thickness can be analyzed using the immersion probe, which makes the technique sensitive to the ibu molecules delivered in aqueous solutions. The acquisition scan rate was 300 nm min-1; a full spectrum was recorded in 60 s. Standard stock ibu aqueous solutions with different concentrations were measured and used to quantitatively determine the cumulative drug release in phosphate buffer. 2.4.3 Toxicity/viability

2.4 Fourier-Transform infrared (FTIR) spectroscopy FTIR spectra were acquired on a Perkin Elmer FTIR Spectrometer Frontier equipped with the ATR accessory, at room temperature. The spectra were collected over the 4,000–400 cm-1 range by averaging 15 scans at a spectral resolution of 4 cm-1. Prior to collecting the spectra, the xerogels were vacuum dried at 50 °C for about 24 h, to reduce the levels of adsorbed water and solvent. 2.4.1 Small-angle X-ray scattering (SAXS) The nanostructure of the materials was investigated by small-angle X-ray scattering (SAXS) experiments, performed at the D01A-SAXS1 beamline of the National Synchrotron Light Laboratory (LNLS, Campinas, Brazil). A vertical position-sensitive X-ray detector and a multichannel analyzer were used to record the SAXS intensity, I(q), as a function of the modulus of the scattering vector q = (4p/k) sin(h/2), q being the scattering angle and k the wavelength of the X-ray (k = 0.148 nm). The sample-to-

Considering that an objective of this study was to evaluate the safety of pure tU5000 xerogel for subsequent applications, it is preferred that normal human cells are used in cytotoxicity assays. In the present study, to evaluate cytotoxicity, normal human lung fibroblasts (GM07492A) were employed. The cell lines were maintained as monolayers in plastic culture flasks (25 cm2) in HAM-F10 ? DMEM 1:1 (Sigma-Aldrich) culture medium supplemented with 10 % fetal bovine serum (Nutricell), antibiotics (0.01 mg mL-1 streptomycin and 0.005 mg mL-1 penicillin; SigmaAldrich), and 2.38 mg mL-1 HEPES (Sigma-Aldrich), at 37 °C, with 5 % CO2. The sample preparation was carried out according to ISO 10993-12:2007. A tU5000 xerogel measuring 1 cm2 and with 0.5-mm thickness was cut into small pieces of 1 mm 9 5 mm. The sample was sterilized under ultraviolet radiation at the two surfaces for 30 min, and submitted to extraction into 900 lL of culture medium without serum at 37 °C for 24 h; the sample was sonicated for 30 min prior to the treatment of the cells. Then, the medium was

123

630

J Sol-Gel Sci Technol (2014) 72:627–636

Fig. 1 Photographs of tU5000 with different percentages (wt%) of ibuprofen

transferred to another flask and supplemented with 100 lL of fetal bovine serum completing the volume to 1,000 lL. This solution was taken as 100 %. The cytotoxic activity was measured using thein vitro Toxicology Colorimetric Assay Kit (XTT; Roche Diagnostics) according to the manufacturer’s instructions and ISO 10993-5:2009. The XTT-based system furnishes a reliable index to measure mitochondrial activity; also, it satisfactorily provides information on cell viability/function for more complex drug screening paradigms [30]. The XTT assay helps to evaluate the sensitivity of cell lines to chemicals; it is widely used to quantitatively determine cell proliferation and the cytotoxic effects of chemicals [31]. For these experiments, the cells (104 cells/well)were plated onto 96-well microplates, each well received 100 lL HAM F10/DMEM medium containing tU5000 immediately after the sonication.The concentrations tested ranging from 0.78 to 100 %. The negative (without treatment) and positive (dimethylsulfoxide–DMSO 25 %) controls were included. After incubation at 37 °C for 24 h, the medium was removed; cells were washed with 100 lL of PBS (phosphate buffered saline) and exposed to 100 lL of HAM-F10 medium without phenol red. Then, 25 lL of XTT were added to each well. The microplates were covered and incubated at 37 °C for 17 h. The absorbance of the sample was measured in a multi-plate reader (ELISA–Asys–UVM 340/Microwin 2000) at a test wavelength of 450 nm and a reference wavelength of 620 nm. Cell viability was expressed as a percentage of untreated cells, which served as the negative control group and was designated as 100 %. The experiments were performed in triplicate.

3 Results and discussion 3.1 Structural characterization Sample transparency attests that the inorganic phase (ICPTES, in the present case) is covalently bound to and homogeneously distributed within the network [32]. Therefore, we evaluated the transparency of ibu-loaded tU5000 with different ibu contents (Fig. 1). The tU5000 hybrid with the highest ibu content (50 wt%) was as

123

Fig. 2 XRD patterns of ibuprofen and of unloaded and ibuprofenloaded tU5000 with different percentages of ibuprofen

transparent as the samples with less ibu, evidencing the good solubility of the ibu molecules within the matrix. Figure 2 corresponds to the powder X-ray diffractograms of ibu, unloaded tU5000, and ibu-loaded tU5000. The tU5000 structure remained the same after incorporation of different ibu contents (1, 3, 5, 10, 15, 20, 30, and 50 wt%). The unloaded and ibu-loaded tU5000 were totally amorphous, as a consequence of the highly cross-linked network. Essentially, all the matrixes displayed one broad Gaussian-shaped peak centered at ca. 21.3–21.6°, assigned to ordering within the siliceous network [33]. Incorporation of 1–50 wt% ibu into tU5000 did not significantly affect aggregation of the polyether (PPO) network and afforded transparent materials. In general, poorly soluble drugs tend to crystallize upon encapsulation [7]. The absence of crystalline ibu in the loaded tU5000 with high drug content (C20 wt%) indicated that the ibu molecules were highly soluble in the PPO chains present in the matrix (Fig. 2). Previous studies have demonstrated polypropylene oxide cored PAMAM significantly enhance the solubility of NSAIDs and due to this core containing PPO a higher solubility enhancement abilities than normal PAMAM dendrimers was observed [27].

J Sol-Gel Sci Technol (2014) 72:627–636

631

Fig. 3 DSC curve of ibuprofen and of unloaded and ibu-loaded tU5000 prepared with different percentages (wt%) of ibuprofen

Fig. 4 SAXS patterns of unloaded and ibuprofen-loaded tU5000 with different percentages (wt%) of ibuprofen

A crucial thermal property concerning chain mobility in tU5000 hybrids is the glass-transition temperature (Tg). Figure 3 depicts the heat flow curves from the DSC analysis of unloaded and ibu-loaded tU5000. Only a single Tg (endothermic event) occurred at both low and high ibu content; this temperature shift as a function of the drug content, going from -64.0 °C for the unloaded hybrid to -55.6 °C for ibu-loaded tU5000 (50 wt%) clearly showing that the rigidity of the polymer is strongly increased by the incorporation of ibu molecules. More specifically, ibu slight increase Tg at small loadings (1 \ wt% \ 10); however, high drug content (C15 wt%) considerably increase Tg as compared with unloaded tU5000. Such a remarkable increasing effect (Fig. 3) on the Tg point upon drug incorporation into the tU5000 matrix could be explain by formation of transient inter/intra-PPO chain cross-linking by the ibu which this phenomena lead to a decrease of the local chain mobility. The fact that the single Tg is linearly related to the concentration of ibu indicates that the drug-loaded hybrid matrixes is characterized by a single relaxation process, which strongly suggests that the incorporation has produced a homogeneous glass solution where the two chemical species (drug-tU5000) are mixed at the molecular level [34, 35]. Taking this even further, the Tg of pure ibu [36] is -45 °C where this suggests the existence of a continuum of amorphous or glassy states between pure tU5000 and ibu and indicates that they mix. We conducted SAXS to analyze the nanostructural homogeneity of unloaded and ibu-loaded tU5000 and the effect of the matrix ibu content. The SAXS curves of unloaded and ibu-loaded tU5000 with different drug contents presented a single broad peak with maximum located at qmax values of *1.31 nm-1 (Fig. 4). This peak is

characteristic of ureasil dense cross-link nodes, which act as scattering centers bound at the PPO chain extremities [37]. Incorporation of low ibu content (1 \ wt% \ 10) into tU5000 did not affect the average correlation distance between two adjacent nodes (fd = 4.79 nm, as determined using the equation fd = 2p/qmax). However, tU5000 with high ibu content (C15 wt%) displayed slightly decreased fd: 4.79 in unloaded tU5000 as compared with 4.59 and 4.42 in ibu-loaded tU5000 20 and 50 wt%, respectively. Ibu addition into tU5000 widened the peak characteristic of spatially correlated siloxane-rich particles embedded in the polymer-rich phase. These effects generally stem from the presence of drug solubilized into the matrixes, increasing the electronic density of the polymeric phase and drastically reducing the contrast between the siloxane domains and the polymer. We recorded FTIR spectra to investigate how the drug and the hybrid matrix interacted. To this end, we followed how the characteristic vibration bands of the ibu molecules and tU5000 changed after ibu incorporation into tU5000. Figure S1 displays the FTIR spectrum recorded for the ibu salt as well as for unloaded tU5000 and ibu-loaded tU5000 matrixes between 600 and 1,900 cm-1. Figure S1 revealed that only the regions of the spectrum localized between 1,500 and 1,800 cm-1 do not present superimposition of bands characteristic of unloaded tU5000 and ibu. For this reason the detailed analysis of the vibration bands was only performed in these spectral regions. Analysis of the spectral signatures of the polyether backbone (800–1,400 cm-1) and of the urea linkage can provide valuable information about ibu solvation by different functional groups present in the tU5000 hybrid network. For this reason, we focused on these two spectral

123

632

J Sol-Gel Sci Technol (2014) 72:627–636

Fig. 5 FTIR of ibuprofen and of unloaded and ibuprofen-loaded tU5000 in the regions characteristic of the main vibrations of the hybrid matrix and ibuprofen: a 1,500–1,900 cm-1, b 800–1,400 cm-1 and c amide I and amide II region 1,530–1,700 cm-1

regions. The vibrational bands around 1,712 and 1,507 cm-1 in the spectra of ibu-loaded tU5000, characteristic of ibu carbonyl-stretching vibration (mC=OOH) and ring, respectively, confirmed ibu incorporation into tU5000 [38, 39]. These bands intensified as the ibu increased concentration in tU5000. For lower ibu content (B5 wt%) the band at 1,507 cm-1 is not visible, because the intensity of the ibu band is simply not be strong enough; so instead of shifting, the band appears at 1,513 cm-1 once enough ibu is present (C10 wt%) to have a visible intensity. The band at 1,712 cm-1 developed a more prominent double peak structure in the loaded hybrids, with two new intense broad bands at 1,707 and 1,735 cm-1 (Fig. 5a). These bands at 1,513, 1,707 and 1,735 cm-1 present in the loaded-tU5000 (with higher ibu content C10 wt%) probably indicates a changed environment between crystalline ibuprofen and the ibuprofen in the polymer matrix, to elicit these spectral changes. The 800–1,400 cm-1 region corresponds to the CH2 rocking and to the C–O and C–C stretching modes; in this range, the bands are very sensitive to conformational changes in the polymer backbone [40]. We assigned the bands in the FTIR spectra of unloaded and ibu-loaded tU5000 on the basis of FTIR assignments reported for triureasils [29, 33] and analogous urea-derived O–I hybrids (the so-called di-ureasils) [41–43]. The bands with maxima at 1,254 and 1,374 cm-1, related to the CH2 twisting and wagging modes, and the most intense band at 1,105 cm-1, assigned to the C–O stretching mode, did not shift upon ibu loading; in fact, the latter band simply broadening as a function of the ibu concentration (Fig. 5b), suggesting the chemical environment of the PPO chains changed upon ibu incorporation into tU5000. Lopes et al. [44] employed Raman spectroscopy to investigate how sodium diclofenac (SDCF) interacted with di-ureasil hybrids (PEO500 and

123

PEO1900). As a consequence of drug–matrix interactions, the authors observed similar broadening of the bands typical of SDCF. They suggested that the amine group bound to the SDCF dichlorophenyl ring interacted with the urea groups or the ether-type oxygen atoms present in the hybrid matrices, to modify the chemical environment of the drug ring. Here, increased ibu loading into tU5000 affected the vibration bands, which may have changed the polymer backbone conformation and/or disrupted the PPO spatial packing. Finally, we closely inspected the region between 1,500 and 1,700 cm-1, related to the so-called ‘‘amide I’’ and ‘‘amide II’’ vibration modes mainly stemming from the C=O, C–N, and C–C stretching modes and the in-plane bending mode of the N–H group stretching vibration of the urea site [NHC(=O)NH] (Fig. 5c) [29]. The bands located at 1,565 (amide II) and 1,641 cm-1 (amide I) did not shift but broadened significantly upon incorporation of high ibu concentrations into tU5000 (Fig. 5c). These observations agreed with broadening of the bands ascribed to the ibu C=OOH group and ring, as a result of alterations in the chemical environment of this drug. The FTIR data corroborated the results from the DSC and SAXS analyses, which had evidenced that the PPO chains changed in the presence of ibu molecules. 3.2 Ibuprofen release kinetics Before used ibuprofen as model drug incorporated into tU5000, we tested sodium diclofenac (SDCF). The increasing opacity of the tU5000 hybrid observed by loading it above 3 wt% reveals the demixion of the SDCF crystals from the hybrid matrix (Figure S2). Thus, we selected ibu for the in vitro drug release assays, since this drug is poorly soluble in water and demonstrated

J Sol-Gel Sci Technol (2014) 72:627–636

633

Fig. 6 Cumulative percentage of the initial ibuprofen loading released into phosphate buffer, at 37 °C, from tU5000 containing a 1, 3, 5, and 10 wt% and b 15, 20, 30, 50 wt% of ibuprofen. All the

experiments were conducted on three independent samples; the results are the average of the data, being the standard deviation less than the symbol size

preferencial dissolution at the hydrophobic PPO chains existing in tU5000. We evaluated how the ibu content affected drug release from tU5000 by measuring the absorbance at 221 nm. Figure 6, parts a and b, brings the ibu release profiles from tU5000 in phosphate buffer (pH 7.4) at 37 °C. Regardless of the ibu loading (wt%), the ibu concentration increased with time, which demonstrated sustained profiles within eight hours. The cumulative drug release from ibu-loaded tU5000 with lower ibu content (1, 3, 5, and 10 wt%) showed that about 10, 25, 60, and 80 % of the loaded drug had been released after eight hours, respectively (Fig. 6a). In this case, hydration did not affect the hydrophobic nature of the PPO chains in tU5000. Consequently, ibu diffusion through the PPO network was much faster than the drug dissolution rate, allowing for a linear release profile [28]. The release rate obtained for larger drug loadings in tU5000 (C15 wt%) remains nearly constant around 80 % cumulative release. We had expected larger ibu release in the former case, but actually we only achieved 80 % release within the same eight hours (Fig. 6b). Probably, the good dissolution of the ibu molecules within the hydrophobic PPO chains (as demonstrated by XRD, DSC, SAXS, and FTIR analyses) hindered ibu diffusion. The water uptake is an important phenomenon in this class of hybrids formed by PEO/PPO polyether [43]. Molina et al. pointed out the contribution of the osmotic flow on the water uptake in ureasil-PEO1900 hybrid and the influence of this osmotic process on release of cisplatin molecules loaded into hybrid matrix. In this case, due to the hydrophilic character of PEO chains, when the matrix is immersed into water, the solute near the surface of the monolithic piece begins to dissolve. Thereafter, the dissolved solute is extracted from the

surface toward the solution, giving rise to an inverse water flux into the matrix, which in turn, dissolves the cisplatin loaded deep inside the matrix [43]. As a consequence of hydrophobic nature of PPO chains present in tU5000 hybrid the water uptake is hindered and only the solute (ibu molecules) present on the surface of the xerogel is dissolved thus presenting a sustained release profile. The sustained drug release results implied that it is possible to tune ibu release from tU5000 by changing the drug content in the tU5000 matrix; also, we succeeded in incorporating high ibu levels into tU5000. Generally, several mechanisms, such as pure diffusion, erosion control, and a combination of these mechanisms, may account for the overall drug release from a polymeric matrix [45]. Several factors such as polymer composition, molecular weight, crystallinity, hydrophilicity, degradation rate, porosity, and surface character can affect these release mechanisms [46]. Lippold and Peppas proposed a semiempirical power law equation to describe the different drug release mechanisms by diffusive transport through a monolithic polymeric matrix [47, 48]. We applied Eqs. (1) and (2) to the experimental ibu release profiles from tU5000 in phosphate buffer as shown in Fig. 7: Mt =M1 ¼ ktn   Mt log ¼ n log t þ log k M1

ð1Þ ð2Þ

where Mt and M? correspond to the cumulative mass of drug released at time t and at infinite time, respectively; k denotes the release constant incorporating structural and geometrical characteristics; and n is a release exponent characterizing the diffusional mechanism. The n values

123

634

Fig. 7 Plots of log (Mt/M?) against log t for ibuprofen release from tU5000 containing a 1, 3, 5, and 10 wt% and b 15, 20, 30 and 50 wt% of ibuprofen. All the experiments were conducted on three

provide direct information on the release kinetics: n ranging from 0.43 to 0.5 characterize a Fickian diffusion mechanism; n between 0.85 and 1 suggests a case II transport mechanism in which the zero-order rate controls drug release; and other n values indicate the superposition of both phenomena (anomalous transport kinetics) [49, 50]. Our group [28] and Yang et al. [51] recently employed this kinetic methodology in release studies of hydrophilic or hydrophobic drugs. Figure 7 contains the plot of log (Mt/ M?) versus log t for ibu-loaded tU5000 (the single solid line corresponding to fitting of Eq (1)). Initial burst was absent, so the ibu release profile corresponded to a linear curve. The release curves presented slopes lying between 0.45 and 0.54 ± 0.03, denoting a mechanism governed by pure ibu diffusion from the PPO hydrophobic chain (Fig. 7a). In the case of 1 wt% ibu-loaded tU5000, the slope of 0.36 ± 0.03 indicated anomalous transport kinetics. An n value of *0.55 ± 0.03 for the samples with high ibu content (15, 20 and 30 wt%) suggested that tU5000 released the drug molecules mainly by diffusion (Fig. 7b). However, the samples with 50 wt% of ibu content showed an n value of 0.77 in which suggests anomalous transport kinetics of ibu by tU5000 matrix. 3.3 Toxicity/Viability of tU5000 hybrid To ascertain tU5000 cytotoxicity, we accomplished XTT assays after treating GM07492A cells with tU5000 concentrations of 0.78–100 % for 24 h. Figure 8 shows the cell viability (%) in the GM07492A cultures. Treatment with tU5000 did not significantly reduce cell viability at any of the tested concentrations, demonstrating that tU5000 was biocompatible.

123

J Sol-Gel Sci Technol (2014) 72:627–636

independent samples; the results are the average of the data, being the standard deviation less than the symbol size

Fig. 8 Cell viability (%) of the GM07492A cell line after treatment with tU5000. Control (no treatment); DMSO (positive control-25 %). The X-axis correspond to tested concentrations (%) of pure tU5000 used in the essays. *Statistically different from the control

In order to provide an alternative vehicle for transdermal drug delivery systems (TDDS), we tested tU5000 hybrid matrix as a film-forming agent. The tU5000 film was prepared via in situ sol–gel process using the same conditions of the experimental section (see Tri-ureasil hybrid synthesis). As seen from Figure S3, the tU5000 organic– inorganic hybrid film was colorless, transparent and present good flexibility, which is easy to be coated on the skin surface and in situ formed a very thin and comfortable film with an aesthetical appearance. Moreover, the viscid gel turned entirely into a solid film after about 8 min. To the best of our knowledge, tri-ureasil hybrid films used in TDDS have not been reported. Thus, we will subject these materials to more intense studies, to elucidate the role

J Sol-Gel Sci Technol (2014) 72:627–636

played by the molecular weight of the polyether chains, experimental conditions for control the time to forming film and evaluate the skin irritation and adherence studies in vivo on pig ear skin, which could be valuable for the further development of novel dosage form for TDDS. In sum, tri-ureasil organic–inorganic hybrid film-forming can offer an opportunity to combine the film-forming properties of a polymer and the stability of an inorganic compound to fabricate highly functional materials which they can offer better cosmetic appearance, more dosage flexibility, ease of use, and higher simplicity of manufacture.

4 Conclusions Combination of the hybrid material technology with the sol–gel process affords a tri-ureasil matrix that constitutes a promising material for biomedical applications, especially in the area of implantable and patches drug delivery devices. The synthesized tU5000 hybrid significantly enhances NSAIDs solubility. XRD, DSC, SAXS, and FTIR analyses support the good solubility of ibuprofen within tU5000. It is possible to tune the release profile of the NSAID ibuprofen through the tU5000 matrix, which in turn exhibits excellent biocompatibility. This class of hybrid material based on polymers and inorganic compounds may have potential applications in the design of ophthalmic, transdermal and implantable drug delivery systems or sustained release of poorly water-soluble drugs, such as doxorubicin, cisplatin, docetaxel, or amphotericin, in addition to ibuprofen already tested. Acknowledgments The authors would like to thank the financial support received from the Brazilian agencies CAPES, CNPq, and FAPESP (project number 2013/02613-0). We thank the Brazilian Synchrotron Light Laboratory (LNLS) for SAXS measurements and the D01A-SAXS1 beamline staff for all assistance with the X-ray scattering experiments. We would also like to thank Leila A. Chiavacci for allowing us to use their laboratory premises (UNESP/Araraquara) during the UV–vis release experiments.

References 1. Sanchez C, Belleville P, Popall M, Nicole L (2011) Applications of advanced hybrid organic–inorganic nanomaterials: from laboratory to market. Chem Soc Rev 40:696–753 2. Sanchez C, Julian B, Belleville P, Popall M (2005) Applications of hybrid organic–inorganic nanocomposites. J Mater Chem 15:3559–3592 3. Sanchez C, Soler-Illia GJAA, Ribot F, Lalot T, Mayer CR, Cabuil V (2001) Designed hybrid organic-inorganic nanocomposites from functional nanobuilding blocks. Chem Mater 13:3061–3083 4. Kickelbick G (2006) Hybrid materials. Wiley-VCH, Weinheim 5. Soler-Illia GJAA, Azzaroni O (2011) Multifunctional hybrids by combining ordered mesoporous materials and macromolecular building blocks. Chem Soc Rev 40:1107–1150

635 6. Mehdi A, Reye C, Corriu RJP (2011) From molecular chemistry to hybrid nanomaterials. Design and functionalization. Chem Soc Rev 40:563–574 7. Harrisson S, Nicolas J, Maksimenko A, Bui DT, Mougin J, Couvreur P (2013) Nanoparticles with in vivo anticancer activity from polymer prodrug amphiphiles prepared by living radical polymerization. Angew Chem Int Ed 52:1678–1682 8. Ge Z, Liu S (2013) Functional block copolymer assemblies responsive to tumor and intracellular microenvironments for sitespecific drug delivery and enhanced imaging performance. Chem Soc Rev 42:7289–7325 9. Laine L, Smith R, Min K, Chen C, Dubois RW (2006) Systematic review: the lower gastrointestinal adverse effects of non-steroidal antiinflammatory drugs. Aliment Pharmacol Ther 24:751–767 10. Narayanan D, Geena MG, Lakshmi H, Koyakutty M, Nair S, Menon D (2013) Poly-(ethylene glycol) modified gelatin nanoparticles for sustained delivery of the anti-inflammatory drug ibuprofen-sodium: an in vitro and in vivo analysis. Nanomedicine 9:818–828 11. Crielaard BJ, Lammers T, Schiffelers RM, Storm GJ (2012) Drug targeting systems for inflammatory disease: one for all, all for one. J Control Release 161:225–234 12. Rosario-Mele´ndez R, Yu W, Uhrich KE (2013) Biodegradable polyesters containing ibuprofen and naproxen as pendant groups. Biomacromolecules 14:3542–3548 13. Gao Q, Xu Y, Wu D, Shen W, Deng F (2010) Synthesis. Characterization, and in Vitro pH-controllable drug release from mesoporous silica spheres with switchable gates langmuir 26:17133–17138 14. Yu D, Williams GR, Yang J, Wang X, Yang J, Li X (2011) Solid lipid nanoparticles self-assembled from electrosprayed polymerbased microparticles. J Mater Chem 21:15957–15961 15. Lamprecht A, Saumet J, Roux J, Benoit J (2004) Lipid nanocarriers as drug delivery system for ibuprofen in pain treatment. Int J Pharm 278:407–414 16. Jia H, Kerr LL (2013) Sustained ibuprofen release using composite poly(lactic-co-glycolic acid)/titanium dioxide nanotubes from Ti implant surface. J Pharm Sci 102:2341–2348 17. Mirzaagha B (2008) Design, synthesis and in vitro evaluation of vinyl ether type polymeric prodrugs of ibuprofen, ketoprofen and naproxen. Int J Pharm 356:167–173 18. Yang P, Gai S, Lin J (2012) Functionalized mesoporous silica materials for controlled drug delivery. Chem Soc Rev 41:3679–3698 19. Vallet-Regi M, Colilla M, Gonzalez B (2001) Medical applications of organic–inorganic hybrid materials within the field of silica-based bioceramics. Chem Soc Rev 40:596–607 20. Zhu Y, Shi J, Shen W, Dong X, Feng J, Ruan M, Li Y (2005) Stimuli-responsive controlled drug release from a hollow mesoporous silica sphere/polyelectrolyte multilayer core-shell structure. Angew Chem Int Ed 44:5083–5087 21. Balas F, Manzano M, Horcajada P, Vallet-Regi M (2006) Confinement and controlled release of bisphosphonates on ordered mesoporous silica-based materials. J Am Chem Soc 128: 8116–8117 22. Vallet-Regi M (2006) Ordered mesoporous materials in the context of drug delivery systems and bone tissue engineering. Chem Eur J 12:5934–5943 23. Makiko F, Naohide H, Kumi S (2000) Effect of fatty acid Esters on permeation of ketoprofen through hairless rat skin. Int J Pharm 205:117–125 24. Vergote GJ, Vervaet C, Driessche IV (2001) An oral controlled release matrix pellet formulation containing nanocrystalline ketoprofen. Int J Pharm 219:81–87 25. Perioli L, Ambrogi V, Bernardini C (2004) Potential prodrugs of non-steroidal anti-inflammatory agents for targeted drug delivery to the CNS. Eur J Med Chem 39:715–727

123

636 26. Chie K, Kenji K, Kazuo M (2009) Synthesis of polyamidoamine dendrimers having poly(ethylene glycol) grafts and their ability to encapsulate anticancer drugs. Bioconjug Chem 11:910–917 27. Koc¸ FE, Senel M (2013) Solubility enhancement of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) using polypolypropylene oxide core PAMAM dendrimers. Int J Pharm 451:18–22 28. Molina EF, Marc¸al L, Carvalho HWP, Nassar EJ, Ciuffi KJ (2013) Tri-ureasil gel as a multifunctional organic–inorganic hybrid matrix. Polym Chem 4:1575–1582 29. Nolasco MM, Vaz PM, Freitas VT, Lima PP, Andre PS, Ferreira RAS, Vaz PD, Ribeiro-Claro P, Carlos LD (2013) Engineering highly efficient Eu(III)-based tri-ureasil hybrids toward luminescent solar concentrators. J Mater Chem A 1:7339–7350 30. Berridge MV, Herst PM, Tan AS (2005) Tetrazolium dyes as tools in cell biology: new insights into their cellular reduction. Biotechnol Annu Rev 11:127–152 31. Bernas T, Dobrucki J (1999) Reduction of a tetrazolium salt, CTC, by intact HepG2 human hepatoma cells: subcellular localisation of reducing systems. Biochim Biophys Acta 1451:73–81 32. Chujo Y, Saegusa T (1992) Adv Polym Sci 100:11–13 33. Freitas VT, Lima PP, Bermudez VZ, Ferreira RAS, Carlos LD (2012) Boosting the emission quantum yield of urea cross-linked tripodal poly(oxypropylene)/siloxane hybrids through the variation of catalyst concentration. Eur J Inorg Chem 32:5390–5395 34. Newman A, Engers D, Bates S, Ivanisevic I, Kelly RC, Zografi G (2008) Characterization of amorphous API: polymer mixtures using X-ray powder diffraction. J Pharm Sci 97:4840–4856 35. Mahieu A, Willart J, Dudognon E, Dane`de F, Descamps MA (2013) New protocol to determine the solubility of drugs into polymer matrixes. Mol Pharm 10:560–566 36. Kianfar F, Antonijevic MD, Chowdhry BZ, Boateng JS (2011) Formulation development of a carrageenan based delivery system for buccal drug delivery using ibuprofen as a model drug. J Biomater Nanobiotech 2:582–595 37. Dahmouche K, Santilli CV, Pulcinelli SH, Craievich AF (1999) Small-angle X-ray scattering study of sol–gel-derived siloxane– PEG and siloxane–PPG hybrid materials. J Phys Chem B 103:4937–4942 38. Freer AA, Bunyan JM, Shankland N, Sheen DB (1993) Structure of (S)-(?)-ibuprofen. Acta Crystallogr Sect C 49:1378–1380 39. Levin CS, Kundu J, Janesko BG, Scuseria GE, Raphael RM, Halas NJ (2008) Interactions of ibuprofen with hybrid lipid bilayers probed by complementary surface-enhanced vibrational spectroscopies. J Phys Chem B 112:14168–14175

123

J Sol-Gel Sci Technol (2014) 72:627–636 40. Yoon S, Ichikawa K, MacKnight WJ, Hsu SL (1995) Spectroscopic analysis of chain conformation of poly(propylene oxide)based polymer electrolytes. Macromolecules 28:4278–4283 41. Zea Bermudez V, Carlos LD, Alcacer L (1999) Sol–gel derived urea cross-linked organically modified silicates. 1. Room temperature mid-infrared spectra. Chem Mater 11:569–580 42. Molina EF, Pulcinelli SH, Santilli CV, Briois V (2012) Ligand exchange inducing efficient incorporation of CisPt derivatives into Ureasil–PPO hybrid and their interactions with the multifunctional hybrid network. J Phys Chem B 116:7931–7939 43. Molina EF, Pulcinelli SH, Santilli CV, Blanchandin S, Briois V (2010) Controlled cisplatin delivery from Ureasil-PEO1900 hybrid matrix. J Phys Chem B 114:3461–3466 44. Lopes L, Molina EF, Chiavacci LA, Santilli CV, Briois V, Pulcinelli SH (2012) Drug–matrix interaction of sodium diclofenac incorporated into ureasil-poly(ethylene oxide) hybrid materials. RSC Advances 2:5629–5636 45. Lemaire V (2003) Structural modeling of drug release from biodegradable porous matrices based on a combined diffusion/ erosion process. Int J Pharm 258:95–107 46. Kim S, Ha J, Lee Y (2000) Poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide)/poly(e-caprolactone) (PCL) amphiphilic block copolymeric nanospheres: II. Thermo-responsive drug release behaviors. J Control Release 65:345–358 47. Zuleger S, Lippold BC (2001) Polymer particle erosion controlling drug release. I. Factors influencing drug release and characterization of the release mechanism. Int J Pharm 217:139–152 48. Siepmann J, Peppas NA (2012) Modeling of drug release from delivery systems based on hydroxypropyl methylcellulose (HPMC). Adv Drug Deliver Rev 64:163–174 49. Ritger PL, Peppas NA (1987) A simple equation for description of solute release II. Fickian and anomalous release from swellable devices. J Control Release 5:37–42 50. Siepmann J, Peppas NA (2001) Modeling of drug release from delivery systems based on hydroxypropyl methylcellulose. Adv Drug Deliver Rev 48:139–157 51. Yang YQ, Guo XD, Lin WJ, Zhang LJ, Zhang CY, Qian Y (2012) Amphiphilic copolymer brush with random pH-sensitive/hydrophobic structure: synthesis and self-assembled micelles for sustained drug delivery. Soft Matter 8:454–464