Key Engineering Materials Vols. 254-256 (2004) pp. 997-1000 online at http://www.scientific.net © (2004) Trans Tech Publications, Switzerland
Porous Hydroxyapatite and Glass Reinforced Hydroxyapatite for Controlled Release of Sodium Ampicillin A.C. Queiroz1,2, S. Teixeira1,3, J.D. Santos1,3, F.J. Monteiro1,3 1
INEB - Instituto de Engenharia Biomédica, Laboratório de Biomateriais, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal; email:
[email protected] 2 Escola Superior de Tecnologia e Gestão, 4901 Viana do Castelo Codex, Portugal 3 Faculdade de Engenharia da Universidade do Porto, Departamento de Engenharia Metalúrgica e Materiais, Rua Dr. Roberto Frias, 4200-465 Porto, Portugal
Keywords: Hydroxyapatite, glass reinforced hydroxyapatite, porous, drug delivery
Abstract. Porous hydroxyapatite (HA) and glass reinforced hydroxyapatite (GRHA) with adequate macro and microporous structure were developed, aiming at being used as drug delivery carrier of antibiotics, for the in situ treatment of periodontitis. Materials were characterised by XRD and FTIR, presenting no changes from similar dense materials. Mercury intrusion porosimetry revealed micropores of less than 1 mm, accounting for 15% of the total porosity. Compression tests have shown close values for HA and GRHA, with the former showing slightly higher values of strength. Ampicillin adsorption was more effective on porous than on dense HA, and was similar for HA and GRHA. Introduction Periodontitis is an oral disease that promotes, in its most severe form, maxillar alveolar bone loss [1, 2]. Current systemic approach leads to the use of long-lasting treatments with very high dosages of antibiotics and anti-inflammatory drugs. This may induce drug resistance of oral and medial pathogens to common antibiotics [3] and particularly when they are present locally in less than the minimum inhibitory concentration (MIC). A local delivery system should provide the locally required amount of antibiotic, and this should naturally correspond to a significant decrease, when compared to a systemic treatment. Also the period of treatment should decrease [3]. To maximise local antibiotic delivery, for a system where the drug is adsorbed, the surface area of the drug releasing agent must be maximised. For example, polymer porous scaffolds have been widely used for soft tissue replacement and drug delivery. By proposing the use of a porous bioactive ceramic scaffold, the aim is not only to locally deliver a specific drug, in this case sodium ampicillin, a wide spectrum antibiotic, but also, after delivery, to continue acting by inducing cell attachment, osseointegration and vascularisation, promoting rapid healing of the bone tissue after an efficient treatment of the periodontitis infection. In this work both dense and highly porous structures of hydroxyapatite (HA) and glass-reinforced hydroxyapatite (GRHA) were obtained. Hydroxyapatite (HA) is a well-known bioceramic extensively used in medical applications. Glass-reinforced hydroxyapatite (GRHA) [4] has been found to present higher mechanical strength than HA, both as coatings [5] and dense materials, and also to be able to degrade faster in contact with living tissues, due to the presence of controlled amounts of b-TCP, eventually leading to a faster response from the host bone.
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Materials and Methods Dense materials were obtained in disc shapes by uniaxially pressing at 180 MPa, followed by sintering at 1200 ºC, and then milled and sieved until granules of 250-850 mm were obtained. Porous HA and GRHA samples with approximately 5x5x5 mm were obtained by immersing polyurethane (PU) foams in the slurries respectively containing HA and GRHA, at 50 ºC. The slurries were obtained by mixing, commercially pure HA (P-120 Plasma Biotal) or GRHA powders, water and a tensoactive agent (LM3 commercial detergent) at pre-established proportions (HA (g): water (ml):tensoactive agent (ml)). After immersion, the sponges were squeezed, for excess removal, and dried overnight. PU was removed by burn-out method as previously described [6]. The best results were obtained with the 6:6:0.2 ratio for both materials. Materials were characterised by SEM, FTIR with split pea accessory and XRD. Microporosity was characterised by mercury intrusion porosimetry, and compressive strength tests were performed in a Lloyd LR 30 K tensile testing equipment, with a load cell of 5 KN. Samples were then tested for drug adsorption and releasing capability. Both dense granules (D) and porous (P66) samples were put in contact with a 10 mg/mL sodium ampicillin solution for 24 hours, at 37ºC, with continuous agitation of 250 rpm. The amount of sodium ampicillin adsorbed was measured from the supernatant, using UV spectroscopy at 230 nm. Adsorption in similar sets of porous samples (VP66, VP64) were carried out in vacuum for 15 min. at 37 ºC, and following a sequence similar to the previously described one, but being agitated at 110 rpm. Results and Discussion SEM observations of the ceramic sponges revealed an interconnective macroporous structure where very well organised macro and micro porosity was observed (Fig.1). These features were found for both HA and GRHA, with this last presenting some less organised aspects, namely more irregular distribution of micropores, as result of the glass liquid sintering and reaction with HA, occuring during sintering cycle. This kind of structure is considered to be adequate, as the macroporosity induces osteconductivity and allows for cells to enter the structure, and nutrients to reach all the cells by vascularisation, while microporosity allows improved cell adhesion [7]. The results of mercury intrusion porosimetry have confirmed that besides macroporosity, there is microporosity, as detected by SEM, with the majority of micropores occuring at diameters less than 1 mm. Micropores are responsible for approximately 15 % of the total porosity. Previous work had established macroporosity as being around 73% and corresponding to pore size between 150-400 mm [8]. Pores distribution for HA and GRHA were similar. Both FTIR and XRD analyses for porous HA and GRHA presented similar features to those previously found [8] with the same dense materials, indicating that the presence of the tensoactive agent and PU sponge did not affect the phase composition. Compression tests were carried out on porous HA (with two different (HA:W:TA) ratios) and GRHA ( Fig.2). For similar contents of ceramic powders, HA was more resistant than GRHA, in particular withstanding higher stress for the highest values of strain achieved in both cases.
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Fig 1. Porous 6:4:0.2 (HA:W:TA) structures of HA a) b) and c) and GRHA d) e) and f). (SEM images).
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The adsorption studies, including for the porous samples an initial stage of adsorption in vacuum, have shown that both granular and porous material could adsorb sodium ampicillin, but adsorption was much more effective, in the case of HA sponge, due to the significantly increased of surface area put in contact with the ampicillin containing solution (Fig. 3). Adsorption onto GRHA followed the same trend as in the case of HA samples.
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Fig. 3. Sodium ampicillin adsorption onto dense HA granules (D); porous 6:6:0.2 (P66) and vacuum adsorption onto porous 6:6:0.2 (VP66) and 6:4:0.2 (VP64).
Conclusions Porous structures of both HA and GRHA, being composed of interconnected macropores, associated to a well distributed range of micropores seem adequate for drug releasing of this antibiotic. Their morphologies seem to adapt well to be used as scaffolds for bone tissue to invade and proliferate, if adequate conditions are created. Porous HA and GRHA showed to be able to adsorb larger amounts of antibiotic than granules, becoming a better candidate material to be used as a scaffold for drug delivery in the case of periodontitis. HA and GRHA did not present very relevant differences. Acknowledgements The authors would like to acknowledge FCT Project “TEXMED”, ref. POCTI/FCB/41402/2001 for financial support. References [1] Krejci CB, Bissada NF, Farah C, Greenwell H.:J Periodontol 1987; 58: 521-528. [2] RA Seymour, Heasman PA, MacGregor IDM. The pathogenesis of periodontal disease. In: MacGregor IDM, editor. Drugs, Diseases, and the Periodontium. Oxford: Oxford University Press, 1992. p. 1-10. [3] Steinberg D, Friedman M, Soskolne A, Sela MN: J Periodontol 1990; 61: 393-398. [4] Santos JD, Silva PL, Knowles JC, Talal S, Monteiro FJ: J Mater Sci: Mater Med 1996; 7: 187-189. [5] Ferraz MP, Monteiro FJ, Santos JD: J Biomed Mat Res 1999; 45: 376-383. [6] Queiroz A.C.,Teixeira S.,Santos J.D.,Monteiro F.J.: Key Engineering Materials, submitted [7] Nishihara K: Clin Materials 1993; 12: 159-167. [8] Queiroz AC, Santos JD, Monteiro FJ, Gibson IR, Knowles JC: Biomaterials 2001; 22: 1393-1400.
