Elsevier

Acta Biomaterialia

Volume 5, Issue 5, June 2009, Pages 1686-1696
Acta Biomaterialia

Mesoporous calcium silicate for controlled release of bovine serum albumin protein

https://doi.org/10.1016/j.actbio.2009.01.012Get rights and content

Abstract

The purpose of this study is to synthesize mesoporous calcium silicate (CS or wollastonite, CaSiO3) and evaluate its possible application in protein/drug delivery. First, calcium silicate was synthesized by wet chemical method and then mesoporosity was created by acid modification of the synthesized CS particle using hydrochloric acid at pH 7, 4.5, and 0.5. The results showed that a hydrated silica gel with abundant Si–OH functional group formed on the surface of calcium silicate due to acid modification. This surface layer had mesoporous structure, with pore diameter between 4 and 5 nm. BET specific average surface area increased to 221, 333, and 356 m2 g−1 due to acid modification at pH 7, 4.5, and 0.5, respectively, whereas the surface area for unmodified CS particles was 65 m2 g−1. Protein adsorption studies indicated that mesoporous CS has higher ability to adsorb bovine serum albumin and lysozyme compared to unmodified particles. The release kinetics showed that proteins on mesoporous CS released sequentially over one week, whereas the proteins on unmodified particle followed burst release kinetics within a few hours. Human osteoblast cell–material interaction study showed that these materials were biocompatible and promoted excellent bone cell proliferation. In summary, this work has demonstrated the potential to produce mesoporous CS as a carrier for protein/drug delivery for bone regeneration and other biomedical applications.

Introduction

Bone-forming growth factors, such as bone morphogenetic protein (BMP) and transforming growth factor (TGF-β), have been widely investigated in orthopedics and hard tissue engineering due to their ability to stimulate bone cell growth, differentiation, and accelerate bone tissue regeneration [1], [2]. Delivery of growth factors must be performed in a controlled manner to prevent deleterious side-effects in non-target tissues. Ideally, growth factors should be given sequentially in a manner that mimics the time profile of the healing process [2], [3], [4]. Some biomaterials have been developed for controlled protein delivery, including biopolymers [5], [6], [7], [8], inorganic ceramics [9], [10], [11], [12], [13], [14], [15], [16], [17], [18], and their composites [19], [20]. These carriers act as reservoirs for the controlled delivery of protein, so they should have high capability for protein adsorption and keep their biological activity. Carriers can also serve as a scaffold to support tissue formation, and thus they should be biocompatible with the bone tissue. Finally, they should be non-immunogenic and are often required to be biodegradable once tissue regeneration is complete [2], [3]. Most of the biopolymers are not bioactive, as they cannot chemically bond to living bone.

Inorganic deliveries, including calcium phosphate ceramics [10], [11], [12], [13], bioglass and bioglass-ceramics [14], silica gel [15], [16], and calcium phosphate cements [17], [18], have been extensively investigated due to their excellent bioactivity with bone tissue. However, protein loading on these inorganic carriers is carried out by surface physical adsorption. The adsorption capacity is limited, which mainly depends on the surface area of materials. Moreover, protein release from these materials shows burst release behavior due to weak bonds between protein and the carriers. Recently, mesoporous carbon [21], [22] and silicon [23], [24], [25] have been developed for protein delivery due to their high surface areas, which can host a high amount of protein into mesoporous structure. However, their bioactivity with bone tissue is limited [26], which has restricted their applications in bone regeneration.

Recently, calcium silicate or wollastonite (CS) has been regarded as a candidate for bone replacement biomaterial due to its good bioactivity. In vitro and in vivo studies showed that CS ceramic could induce a bone-like apatite layer formation in simulated body fluid (SBF) [27] and chemically integrate into the structure of living bone tissue [28]. To improve the mechanical properties, CS coating on titanium alloy was also developed by plasma spray [29], [30]. Recently, nano-sized calcium silicate particle was synthesized and its bioactivity was demonstrated by in vitro study [31], [32]. The analyses of these in vitro and in vivo studies reveal that the primary reason for bioactivity of calcium silicate is the formation of Si–OH on its surface when exposed to body fluid. CS is a chain-silicate mineral which consists of a network of covalently bonded silica that is interrupted and modified by Ca2+ cations [33], [34]. This weakly bonded, network-modifying Ca2+ is released to the solution as they exchange for hydrogen ions, resulting in the formation of Si–OH.

In this study, we have synthesized mesoporous calcium silicate as a vehicle for protein delivery with controlled release kinetics. In order to improve the bioactivity of CS and protein loading capacity, acid modification was applied. By acid modification, mesoporosity was created on the surface of CS, which provided very high surface area for protein adsorption. Acid treatment also produced abundant Si–OH functional group on the material surface, which helped to improve bioactivity and strengthened protein interactions with the material surface. A human osteoblast cell–material interaction study was conducted only to understand the cytotoxicity behavior of these porous Ca-silicates. The possibility of combining both bioactivity and controlled protein/drug release behavior can make these porous CS materials ideal for use in bone implant application.

Section snippets

Sample synthesis and characterization

Calcium silicate powder was synthesized by reacting aqueous solutions of 0.4 mol Ca(NO3)2·4H2O and 0.4 mol Na2SiO3·9H2O to form a white precipitate [31]. The white precipitate was then stirred for 4 h and filtered, followed by washing with deionized (DI) water. The precipitate was finally washed two times with 100% ethanol. The powder was dried at 80 °C for 24 h.

Mesoporous structure of CS was produced by acid treatment using hydrochloric acid (HCl) solution. The above dry calcium silicate powder was

Materials characterization

XRD spectra and the SEM-EDS analysis of calcium silicate before and after acid modification and silicic acid are shown in Fig. 1a and Table 1, respectively. After acid modification, an amorphous hump appeared in between 20° and 30° regions, which is attributed to hydrated silica. This hump was more obvious when wollastonite was treated at lower pH value. The spectrum of CS-0.5 was similar to that of silicic acid. From the EDS analysis, it was noted that Ca content decreased significantly after

Discussion

The purpose of this study is to prepare mesoporous wollastonite or calcium silicate by acid modification and evaluate its possible application in protein/drug delivery for bone regeneration. Calcium silicate has good bioactivity, and has been regarded as a candidate for bone replacement biomaterial. This study showed that a hydrated silica gel layer with mesoporous structures can be created by surface modification. Mesoporous structures have high surface area, which allow proteins to be hosted

Conclusion

Mesoporous calcium silicate (CS) or wollastonite was prepared by acid modification where hydrated silica gel with Si–OH functional group was formed on the surface of CS. This surface layer had a mesoporous structure, with pore diameter between 4 and 5 nm. Mesoporous CS particles also showed very high surface area. A maximum of 356 m2 g−1 of BET specific average surface area was achieved for the particle modified at pH 0.5. Protein adsorption studies showed that mesoporous wollastonite had higher

Acknowledgments

The authors gratefully acknowledge financial support from the National Science Foundation (Grant No. 0134476) and the Office of Naval Research (Grant Nos. N00014-1-04-0644 and N00014-1-05-0583).

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