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Preparation and mechanical properties of nanocomposites of poly(d ,l -lactide) with Ca-deficient hydroxyapatite nanocrystals
Biomaterials 22 (2001) 2867}2873
Preparation and mechanical properties of nanocomposites of poly(D,L-lactide) with Ca-de"cient hydroxyapatite nanocrystals Xianmo Deng*, Jianyuan Hao, Changsheng Wang Chengdu Institute of Organic Chemistry, Academia Sinica, P.O. Box 415, Chengdu 610041, People+s Republic of China Received 24 May 2000; accepted 9 January 2001
Abstract Nanocomposites of high molecular poly(D,L-lactide) (PLA) with Ca-de"cient hydroxyapatite nanocrystals (d-HAP) were successfully prepared through solvent-cast technique. Such composites are of great importance to make bone-like substitutes as d-HAP nanocrystals have similar composition, morphology and crystal structure as natural apatite crystals. Of all the PLA solvents studied, N,N-dimethylformamide is the best one to disperse d-HAP nanocrystals. The resultant sol is a blue, stable dispersion that could preserve several days with only slight precipitation. The bright-"eld TEM micrograph shows that d-HAP nanocrystals form homogeneous dispersion in the PLA matrix at a microscopic level. The tensile modulus for PLA/d-HAP nanocomposites increases with d-HAP loading. Theoretical prediction of the modulus has been made by assuming the nanocomposites as short "ber "lled systems. The calculated values based on Halpin}Tsai equations show excellent agreement with the experimental results. The yield stress for the nanocomposites has not been undermined by the presence of the nanocrystals. This preservation of strength for PLA/d-HAP nanocomposites may be due to the homogeneous dispersion of d-HAP nanocrystals in the PLA matrix as well as the good interfacial adhesion. 2001 Published by Elsevier Science Ltd. Keywords: Nanocomposites; Poly(D,L-lactide); Ca-de"cient hydroxyapatite nanocrystals
1. Introduction Various shapes and sizes of bone defects caused by trauma, tumor and infections must be "lled with suitable substances to promote bone repair. The most common bone substitutes used in clinic are autografts, allografts, polymers such as polymethylmethacrylate (PMMA) and ceramics such as dense and porous hydroxyapatite (HAP), tricalcium phosphate ceramics and bioglass ceramic in the systemof Na O}CaO}SiO }P O [1}3]. But none of the materials presently available is entirely suitable for orthopedic applications, each su!ering speci"c disadvantages [4}6]. For examples, autografts are restricted by their donor bone supply, allografts might elicit an immunologic response in the human body, and PMMA would lead to local necrosis of surrounding tissues due to a great deal of heat released on the setting process.
* Corresponding author. E-mail address: [email protected] (X. Deng).
Clinically, bioactive ceramics are most widely used and investigated and the most prominent example is hydroxyapatite (HAP). Because this ceramics allows osteogenesis to occur, it provides a bony contact and even bonds with host bone [7]. However, whether used in block or granular forms, pure HAP could not degrade in the human body. In the case of block form, the high failure rate of HAP ceramics further impedes its clinical applications. Thus recently, much attention has been focused on HAP/polymer composites [8}10], especially those containing biodegradable polymers. The polymeric parts are metabolized and excreted, and the ceramic constituents are assimilated in the body. The brittleness of HAP ceramics is also improved by mixing with tough organic polymers. Recently a few reports have been presented concerning synthesis of nanocrystals of Ca-de"cient hydroxyapatite (d-HAP) by hydrothermal methods [11}12]. d-HAP is of greater biological interest than stoichiometric HAP (sHAP) since the Ca/P ratio in bone is lower than 1.67. When d-HAP or s-HAP is used as a synthetic material, there are di!erences in their behavior. It has been
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reported that d-HAP elicits an immediate precipitation of biologically equivalent apatite on its surface when immersed in a simulated physiological #uid, whereas precipitation on s-HAP requires some induction time [13]. In addition to the compositional similarity, the hydrothermally synthesized d-HAP crystals also have similar morphology and crystal structure as natural apatite crystals [11]. Poly(D,L-lactide) (PLA) is one of the main polymer groups used in biomaterials research. It is biodegradable, essentially non-toxic, elicits only a mild in#ammatory response and the lactic acid yielded after hydrolysis is a normal intermediate of carbohydrate metabolism without accumulation in vital organs. In view of biomimetics and nanomaterials, a combination of above two components is of great signi"cance and expected to result in a promising nanocomposite. This nanocomposite has possible prospects for application as bone implant in the human body. However, it is a challenging work to make such nanocomposites as d-HAP nanocrystals are synthesized in aqueous environment and especially tend to agglomerate when being dried. Liu [14] has previously reported mechanical and physicochemical characteristics of nano-HAP/Polyactive2+ 70 : 30 composites produced by co-precipitation method. The existence of HAP nanocrystals showed strong ability to promote the calci"cation of the composites compared with the pure polymer. In this paper, solvent-cast technique was employed to make PLA/d-HAP nanocomposites with special design to achieve good dispersion of nanocrystals in the polymer matrix. High molecular PLA was chosen as material matrix to endow the nanocomposites with good mechanical strength. Transmission electron microscopy (TEM) was used to evaluate the dispersion of nanocrystals in the dimethyl formamide (DMF) solvent and the polymer PLA matrix. The in#uence of d-HAP loading on mechanical properties of the materials were also determined by tensile tests.
2.2. Synthesis of high molecular PLA High molecular PLA was synthesized in bulk using Al(i-Bu) /H PO /H O as the initiator [15]. The polym erization was carried out under nitrogen atmosphere at the temperature above the melting point of D,L-lactide. The resulting polymer was then dissolved in CHCl , and recovered by precipitation in excess diethyl ether. The puri"ed product was dried under vacuum at 403C for 48 h. The weight average molecular weight was 7.3;10, as determined by intrinsic viscosity measurement in tetrahydrofuran at 373C using the relation []" 1.04;10\ M . The molecular weight distribution was 2.1 determined by gel permeation chromatography (GPC). 2.3. Dispersion of d-HAP nanocrystals in DMF Approximately 2 g of d-HAP nanocrystals was dispersed in 200 ml DMF using ultrasonics for 30 min. The dispersion was then "ltered for removal of large aggregates and a blue sol "ltrate was obtained. The accurate concentration of the sol was determined by precipitation 10 ml of the sol into an excess of ethyl ether, followed by weighting the precipitate. 2.4. Preparation of PLA/d-HAP nanocomposites PLA/d-HAP nanocomposites were prepared through solvent-cast technique. Pre-weighted PLA was added into a #at-bottom #ask that contained the d-HAP sol. The #ask was sealed o! and stirred at room temperature until PLA totally dissolved into the sol. The resulting mixture was poured into an aluminum mould and then heated to 1403C for 1 h in a thermostat oven for removal of the solvent. The "nally obtained nanocomposite "lm was furthered dried under vacuum at 803C for 12 h. 2.5. Transmission electron microscopy (TEM)
2. Materials and methods 2.1. Preparation of d-HAP nanocrystals d-HAP nanocrystals were synthesized by a hydrothermal method described elsewhere [11]. A (NH ) HPO aqueous solution of 200 ml (11.4 wt%) was slowly dropped into a stirred, 400 ml Ca(NO ) aqueous solution (16.8 wt%). The pH for both solutions was 10}12, adjusted with ammonium hydroxide solution, and the reaction was carried out at room temperature. The resultant precipitates were put into an autoclave in a solid-solution ratio of 2 wt% and hydrothermally treated at 1403C and 0.3 MPa for 5 h, followed by centrifugal washing with deionized water. The hydrothermal slurry was "nally dried and a white power was obtained.
TEM was used to evaluate the dispersion of d-HAP nanocrystals in the DMF solvent and the PLA polymer matrix as well. All microscopic investigations were performed on a JEOL JEM-100CX TEM operating at 100 kV. The sol sample was visualized directly by dripping a drop into a gold TEM grid (hexagonal 400 mesh), which was dried completely on "lter paper at room temperature. The bulk nanocomposites were obtained from several pieces of composite "lms that were overlapped, and then treated under pressure at 1403C to melt into thick pellets. These pellets were cut to form a triangular block face (approximately 1 mm;0.5 mm) for microtoming. Ultrathin sections were microtomed from this face, at room temperature, using a ultramicrotome. A water-"lled boat
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was attached to the knife, so that after cutting, the ultrathin sections could be #oated onto water. The sections were collected on gold TEM grids and observed after drying in air. 2.6. Tensile tests The stress}strain curves of solution-cast "lms were obtained at 153C on an Instron 4302 machine. The "lms were 0.1 mm thick and tested at a stretching speed of 50 mm/min. At least "ve specimens were measured and the mechanical tensile data were determined from the curves on the average of three specimens. 2.7. Scanning electron microscopy (SEM) SEM was performed with an AMRAY 1000B equipment operated at 18 kV to examine the fracture surfaces of the nanocomposites. All samples were coated with a thin layer of gold by means of a polaron sputtering apparatus.
3. Results 3.1. Characterization of d-HAP nanocrystals The hydrothermally formed d-HAP nanocrystals are rod-like particles with the length between 40}80 nm and the width between 20}40 nm. The Ca/P ratio and the surface area of these nanocrystals were determined as 1.61 by an atomic absorption spectrometer for calcium combined with an ultraviolet spectrophotometer for phosphorus, and 80 m/g by BET method, respectively. 3.2. Dispersion of d-HAP nanocrystals in DMF and the PLA polymer matrix In order to prepare PLA/d-HAP nanocomposites through solvent-cast technique, the solvent selected to disperse HAP nanocrystals should be able to dissolve the PLA polymer. Several PLA solvents such as chloroform, dichloroethane, benzene and DMF were studied to evaluate their dispersive abilities for the nanocrystals. It was found that DMF is the best solvent to disperse the d-HAP nanocrystals. The acquired dispersion was a stable, blue sol that could preserve for several days with only slight precipitation. Fig. 1a is the TEM image of d-HAP nanocrystals dispersed in the DMF solvent. As is shown that most particles are detached from each other with only slight aggregation emergence. The obtained nanocomposites were semitransparent "lms with the thickness about 0.1 mm. Fig. 1b shows a typical bright-"eld TEM micrograph of the nanocomposite containing 1.8 v% d-HAP nanocrystals. It is
Fig. 1. TEM micrographs of d-HAP nanocrystals: (a) dispersed in the DMF solvent; (b) dispersed in the PLA polymer matrix (containing 1.8 v% d-HAP); (c) higher magni"cation of nanocrystals in (b).
shown that homogeneous distribution of the nanocrystals at a microscopic level is observed in the PLA matrix. So far this dispersion level is the best working result for polymer/HAP nanocrystals composites. Fig. 1c is a higher magni"cation photo of the nanocrystals in Fig. 1b. Close contact between the polymer and the d-HAP nanocrystals is observed with no cracks or voids observed in the interface of two phases. 3.3. Mechanical properties of PLA/d-HAP nanocomposites The stress}strain behavior of PLA/d-HAP nanocomposites were characteristic of quasi-brittle materials, all specimens breaking at a point shortly after the yield
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Fig. 2. Dependence of tensile modulus on d-HAP loading for PLA/dHAP nanocomposites; the solid line was calculated from Eq. (5).
Fig. 4. Fracture surfaces of (a) pure polymer; (b) the nanocomposite containing 5.3 v% d-HAP.
Fig. 3. Dependence of yield stress on d-HAP loading for PLA/d-HAP nanocomposites.
maximum on the stress}strain curves but prior to neck formation. The relationship between the d-HAP loading and the tensile modulus for the nanocomposites is illustrated in Fig. 2. The presence of the "lled nanocrystals substantially increase the tensile modulus relative to the pristine polymer. For the nanocomposite containing 10.5 v% dHAP, the modulus even reaches to 2.47 GPa as compared to 1.66 GPa for pure PLA. Fig. 3 shows the dependence of yield stress on d-HAP loading. It is observed that the yield stress for the nanocomposites varies irregularly with d-HAP loading. In a "rst approximation, the yield stress is almost unaffected by d-HAP loading. 3.4. Fracture surface observation by SEM The fracture mode of the nanocomposites was quite similar to that of the pure polymer. Fracture usually initiated at the stress concentration at one end of the fracture surface and propagates perpendicularly against
the loading direction. Fig. 4a and b shows the fracture surfaces of the nanocomposite containing 5.3 v% d-HAP and the pure polymer, respectively. In the middle of the photos, a macroscopic shear ribbon perpendicular to the direction of crack growth is observed for both specimens. Several such shear ribbons could be observed along the whole fracture surface. The fracture surfaces between two shear ribbons indicate brittle fracture and appear much rougher for the nanocomposite than for the pure polymer.
4. Discussion 4.1. Praparation of d-HAP nanocrystals and PLA/d-HAP nanocomposites The dispersion degree of d-HAP slurry after hydrothermal treatment was greatly decided by synthesis conditions such as the initial solution concentrations, their pH values and so on. A relatively strong basicity for the reaction solutions was found to be bene"cial for obtaining less-aggregated hydrothermal slurry. Actually, only under well-controlled synthesis conditions, the
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"nally obtained nanocrystals could turn into a blue sol in the DMF solvent. The dried d-HAP nanocrystals were redispersed in organic solvent to prepare PLA/d-HAP nanocomposites through solvent-cast technique. Among the several PLA solvents studied, such as chloroform, dichloroethane, benzene and DMF, DMF is the best to disperse the nanocrystals. As shown in Fig. 1a, well-dispersed nanocrystals were observed in the TEM image. The dispersion was a blue sol and remained stable for several days with only slight precipitation. This feature o!ers possibility to prepare real PLA/d-HAP nanocomposites through solvent-cast technique. It seems that the solvents with a greater polarity are more e!ectively to disperse the HAP nanocrystals than those with a lower polarity. However in dispersing media such as water or methanol, the polarity of which is larger than that of DMF, the dispersion of the nanocrystals is much poorer than that in the DMF solvent. This fact suggests that the polarity of the solvents is not the only or the dominating factor that determines their dispersive abilities. The well-dispersed nanocrystals in the DMF solvent may be ascribed to its speci"c molecular structure, which is e!ective to stable the HAP nanocrystals. The TEM observation of the nanocomposite indicates good dispersion of d-HAP nanocrystals in the polymer matrix (Fig. 1a and b). Since the nanocomposites were prepared through solvent-cast technique, the gradual removal of the solvent from composite solutions may induce serious agglomeration of the nanocrystals. However, this case had not actually occurred. A possible reason for this may be due to the high viscosity of the solutions resulting from PLA component that e!ectively prevents d-HAP nanocrystals form agglomeration. 4.2. Advantages of PLA/d-HAP nanocomposites as bone implant materials An excellent bone implant material should be able to rapidly induce osteogenesis and form tightly bony bonds with host bone. In mature bone HAP nanocrystals are all irregular shaped thin plates of carbonate apatite with average lengths and widths of 50;25 nm and thicknesses of 2}3 nm [16}18]. Further, natural apatite is a Cade"cient hydroxyapatite with the Ca/P ratio lower than 1.67. These characteristics have little similarity to those for synthetic HAP commonly used in the form of polygonal sintered coarse particles with polycrystalline structure. From the view of biomemetics, better osteogenesis would be achieved if synthetic HAP has more similarity to bone mineral in composition, crystal structure, crystallinity and morphology [19}21]. In this paper, our hydrothermally synthesized HAP is Ca-de"cient rod-like nanocrystals with the length between 40}80 nm and the width between 20}40 nm. These nanocrystals has much more similarity to natural bone mineral in the mentioned
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compositional and morphological aspects and therefore better osteoconductivity for the nanocomposites is expected. Another advantage for the PLA/d-HAP nanocomposites as bone implant materials is that they could be totally assimilated in the human body. Thus, ideal bone implant materials may be produced from these nanocomposites that could rapidly form bony bonds with host bone and gradually be replaced by the production of new bone. 4.3. Mechanical properties of PLA/d-HAP nanocomposites The incorporating of d-HAP nanocrystals into the PLA polymer matrix could signi"cantly improve the rigidity of the materials (Fig. 2). This increase in modulus with "ller content could be theoretically predicted by assuming the nanocomposites as short "ber "lled systems. For a composite containing unidirectional arrayed "bers, the e!ect of "ller on moduli could be described using Halpin}Tsai equations [22]: 1!K* < E " E , * 1#2(l /d )K < * 1#2K2 < E2 " E , 1!K2 <
(1) (2)
with (E /E )!1 K* " , (3) (E /E )#2(l /d ) (E /E )!1 K " . (4) 2 (E /E )#2 Where E* and E2 are the composite moduli in the longitudinal and transverse direction of the "bers orientataion, respectively; E and E are the matrix and the "ller moduli, respectively; V is the volume fraction of the "ller; l is the average length of the "ller; d is the average thickness of the "ller. However for the cases of PLA/dHAP nanocomposites, the spatial orientation of the nanocrystals is not a one-dimensional alignment but a random array in three dimensions. For such a composite, the modulus could be estimated from below experimental equation on the basis of Eqs. (1) and (2) [23]: 3 5 E " E* # E2 . 8 8
(5)
The solid line in Fig. 2 is the prediction of the modulus for the nanocomposites obtained from Eq. (5). In the calculation, the moduli of the PLA matrix (E ) and the
d-HAP nanocrystals (E ) are taken as 114.0 and 1.66 GPa, respectively [24]; the value of l /d for the nanocrystals is taken as 3. It is seen that the experimental values fall on the solid line indicating the good agreement between prediction and experiment.
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The yield stress of the nanocomposite varies little with d-HAP loading (Fig. 3). This is generally the case when there is good adhesion between the polymer and the "ller [25,26]. Thermal gravimetic analysis demonstrated that there was 6 wt% organic matter on the surfaces of the d-HAP nanocrystals that were separated from the nanocomposites. This organic layer, mainly composed of PLA molecules, are tightly bonded onto the surfaces of d-HAP nanocrsystals indicating good adhesion between the matrix and the "ller. Further, the preservation of strength for PLA/d-HAP nanocomposites has close relation with the nanocrystals' dispersion in the PLA matrix. If the nanocrystals aggregate heavily, the internal crack of the aggregate particles would make them no longer bear load, thus resulting in a decreased strength for the nanocomposites. Fortunately, this is not the case for the nanocomposites in which homogeneous disperison of the nanocrystals in a micro-scale is observed (Fig. 1c). 4.4. Fracture mechanism The presence of d-HAP nanocrtystals does not in#uence the fracture mode of the nanocomposites relative to the pristine polymer. Fracture usually initiates at the stress concentration at one end of the fracture surface and propagates with brittle fracture or ductile shear alternatively arranged along the fracture surface. The ductile shear deformation consumes most of the fracture energy and gives rise to shear ribbons perpendicular to the direction of crack growth. The fracture surface for the nanocomposite appears rougher than the pure polymer due to the presence of d-HAP nanocrystals.
5. Conclusions In this paper, preparation of PLA/d-HAP nanocomposites has been realized by solvent-cast technique. Such composites are of great importance to make bone-like substitutes as d-HAP nanocrystals have similar composition, morphology and crystal structure as those of natural apatite crystals. Of all the PLA solvents studied, N,N-dimethylformamide is the best to disperse d-HAP nanocrystals. The obtained dispersion is a stable, blue sol that could preserve for several days with only slight precipitation. This feature o!ers possibility to prepare real PLA/d-HAP nanocomposites through solvent-cast technique. The obtained nanocomposites are semitransparent "lms with the thickness about 0.1 mm. The SEM observation indicates close contact between the polymer matrix and the "lled nanocrystals. The dispersion of nanocrystals in the polymer matrix is homogeneous at a microscopic level. The tensile modulus for the nanocomposites increases with d-HAP loading. Theoretical prediction of the
modulus has been made by assuming the nanocomposites as short "ber "lled systems. The calculation values based on Halpin}Tsai equations show excellent agreement with the experimental results. The yield stress for the nanocomposites has not been undermined by the presence of the nanocrystals. This preservation of strength for PLA/d-HAP nanocomposites may be due to the homogeneous dispersion of d-HAP nanocrystals in the PLA matrix as well as the good interfacial adhesion.
References [1] Lin FH, Yao CH, Huang CW, Sun JS. The bonding behavior DP-bioactive glass and bone tissue. Mater Chem Phys 1996;46:36}42. [2] Kurashina K, Kurita H, Takeuchi H, Hirano M, Klein CPAT. Osteogenesis in muscle with composite graft of hydroxyapatite and autogenous calvarial periosteum: a preliminary report. Biomaterials 1995;16:119}23. [3] Denissen HW, de Groot K. Immediate dental root implants from synthetic dense calcium hydroxyapatite. J Prosthet Dent 1979; 42:551}6. [4] Lord CF, Gebhardt MC, Tomford WW, Mankin HJ. Infection in bone allografts. J Bone Jt Surg 1988;70A:369}76. [5] Berry BH, Lord CF, Gebhardt MC, Mankin HJ. Allograft fractures: frequency, treatment, and end results. J Bone Jt Surg 1989;72A:825}33. [6] Aebi M. In: Aebi M, Regazzoni B, editors. Bone transplantation, updating on osteochondral auto and allografting. Berlin: Springer, 1987. [7] Idem. Clin Orthop 1981;157:259. [8] Liu Q, de Wijn JR, Bakker D, Van Blitterswijk CA. Surface modi"cation of hydroxyapatite to introduce interfacial bonding with polyactive2+ 30/70 in a biodegradable composite. J Mater Sci Mater Med 1996;7:551}7. [9] Kikuchi M, Suetsugu Y, Tanaka J, Akao M. Preparation and mechanical properties of calcium phosphate/copoly-L-lactide composites. J Mater Sci Mater Med 1997;8:361}4. [10] Verheyen CCPM, Klein CPAT, de Blieckhogervorst JMA, Wolke JGC, van Blitterswijn CA. Evaluation of hydroxyapatite/poly(Llactide) composites: physico-chemical properties. J Mater Sci Mater Med 1993;4:58}65. [11] Li YB, de Wijn J, Klein CPAT, Van de Meer S. Preparation and characterization of nanograde osteoapatite-like rod crystals. J Mater Sci Mater Med 1994;5:252}5. [12] Li YB, de Groot K, de Wijn J, Klein CPAT, Van de Meer S. Morphology and composition of nanograde calcium phosphate needle-like crystals formed by simple hydrothermal treament. J Mater Sci Mater Med 1994;5:326}31. [13] Bett JAS, Christner LG, Hall WK. Studies of the hydrogen held by solids. XII. Hydroxyapatite catalysts. J Am Chem Soc 1967;89:5535. [14] Liu Q, de Wijn J, van Blitterswijk CA. Nano-apatite/polymer composites: mechanical and physicochemical characteristics. Biomaterials 1997;18:1263}70. [15] Deng XM, Xiong CD, Cheng LM, Huang HH, Xu R. Studies on the block copolymerization of D,L-lactide and poly(ethylene glycol) with aluminum complex catalyst. J Appl Polym Sci 1995;55:1193}6. [16] Lowenstam HA, Weiner S. On biomineralization. New York: Oxford University Press, 1989.
X. Deng et al. / Biomaterials 22 (2001) 2867}2873 [17] Robinson RA. An electron microscope study of the crystalline inorganic component of bone and its relationship to the organic matrix. J Bone Jt Surg 1952;34A:389}434. [18] Weiner S, Price PA. Disaggregation of bone into crystals. Calcif Tissue Int 1986;39:365}75. [19] Posner AS. The mineral of bone. Clin Orthop Rel Res 1985;200:87}99. [20] Ellies LG, Garter JM, Natiella JR, Featherstone JDB, Nelson DGA. Quantitative analysis of early in vivo tissue response to synthetic apatitic implants. J Biomed Mater Res 1988;22:137}48. [21] Li Y, Klein GPAT, de Wijn J, van de Meer S, Groot K. Shape change and phase transition of needle-like non-
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stochiometric apatite crystals. J Mater Sci Mater Med 1994;5:263}8. Halpin JS, Tsai SM. E!ects of environmental factors on composite materials. AFML-TR 67-423, June 1969. Mallick PK. Fiber-reinforced composites: materials, manufacturing, and design. New York: Marcel Dekker Inc., 1988. p. 110}2. Gilmore RS, Katz JL. Elastic properties of apatite. J Mater Sci 1982;17:1131}41. Nicholais L, Nicodemo L. Strength of particlulate composite. Polym Eng Sci 1973;13:469. Sahu S, Broutman LJ. Mechanical properties of particulate composites. Polym Eng Sci 1972;12:91.
Preparation and mechanical properties of nanocomposites of poly(D,L-lactide) with Ca-de"cient hydroxyapatite nanocrystals Xianmo Deng*, Jianyuan Hao, Changsheng Wang Chengdu Institute of Organic Chemistry, Academia Sinica, P.O. Box 415, Chengdu 610041, People+s Republic of China Received 24 May 2000; accepted 9 January 2001
Abstract Nanocomposites of high molecular poly(D,L-lactide) (PLA) with Ca-de"cient hydroxyapatite nanocrystals (d-HAP) were successfully prepared through solvent-cast technique. Such composites are of great importance to make bone-like substitutes as d-HAP nanocrystals have similar composition, morphology and crystal structure as natural apatite crystals. Of all the PLA solvents studied, N,N-dimethylformamide is the best one to disperse d-HAP nanocrystals. The resultant sol is a blue, stable dispersion that could preserve several days with only slight precipitation. The bright-"eld TEM micrograph shows that d-HAP nanocrystals form homogeneous dispersion in the PLA matrix at a microscopic level. The tensile modulus for PLA/d-HAP nanocomposites increases with d-HAP loading. Theoretical prediction of the modulus has been made by assuming the nanocomposites as short "ber "lled systems. The calculated values based on Halpin}Tsai equations show excellent agreement with the experimental results. The yield stress for the nanocomposites has not been undermined by the presence of the nanocrystals. This preservation of strength for PLA/d-HAP nanocomposites may be due to the homogeneous dispersion of d-HAP nanocrystals in the PLA matrix as well as the good interfacial adhesion. 2001 Published by Elsevier Science Ltd. Keywords: Nanocomposites; Poly(D,L-lactide); Ca-de"cient hydroxyapatite nanocrystals
1. Introduction Various shapes and sizes of bone defects caused by trauma, tumor and infections must be "lled with suitable substances to promote bone repair. The most common bone substitutes used in clinic are autografts, allografts, polymers such as polymethylmethacrylate (PMMA) and ceramics such as dense and porous hydroxyapatite (HAP), tricalcium phosphate ceramics and bioglass ceramic in the systemof Na O}CaO}SiO }P O [1}3]. But none of the materials presently available is entirely suitable for orthopedic applications, each su!ering speci"c disadvantages [4}6]. For examples, autografts are restricted by their donor bone supply, allografts might elicit an immunologic response in the human body, and PMMA would lead to local necrosis of surrounding tissues due to a great deal of heat released on the setting process.
* Corresponding author. E-mail address: [email protected] (X. Deng).
Clinically, bioactive ceramics are most widely used and investigated and the most prominent example is hydroxyapatite (HAP). Because this ceramics allows osteogenesis to occur, it provides a bony contact and even bonds with host bone [7]. However, whether used in block or granular forms, pure HAP could not degrade in the human body. In the case of block form, the high failure rate of HAP ceramics further impedes its clinical applications. Thus recently, much attention has been focused on HAP/polymer composites [8}10], especially those containing biodegradable polymers. The polymeric parts are metabolized and excreted, and the ceramic constituents are assimilated in the body. The brittleness of HAP ceramics is also improved by mixing with tough organic polymers. Recently a few reports have been presented concerning synthesis of nanocrystals of Ca-de"cient hydroxyapatite (d-HAP) by hydrothermal methods [11}12]. d-HAP is of greater biological interest than stoichiometric HAP (sHAP) since the Ca/P ratio in bone is lower than 1.67. When d-HAP or s-HAP is used as a synthetic material, there are di!erences in their behavior. It has been
0142-9612/01/$ - see front matter 2001 Published by Elsevier Science Ltd. PII: S 0 1 4 2 - 9 6 1 2 ( 0 1 ) 0 0 0 3 1 - X
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reported that d-HAP elicits an immediate precipitation of biologically equivalent apatite on its surface when immersed in a simulated physiological #uid, whereas precipitation on s-HAP requires some induction time [13]. In addition to the compositional similarity, the hydrothermally synthesized d-HAP crystals also have similar morphology and crystal structure as natural apatite crystals [11]. Poly(D,L-lactide) (PLA) is one of the main polymer groups used in biomaterials research. It is biodegradable, essentially non-toxic, elicits only a mild in#ammatory response and the lactic acid yielded after hydrolysis is a normal intermediate of carbohydrate metabolism without accumulation in vital organs. In view of biomimetics and nanomaterials, a combination of above two components is of great signi"cance and expected to result in a promising nanocomposite. This nanocomposite has possible prospects for application as bone implant in the human body. However, it is a challenging work to make such nanocomposites as d-HAP nanocrystals are synthesized in aqueous environment and especially tend to agglomerate when being dried. Liu [14] has previously reported mechanical and physicochemical characteristics of nano-HAP/Polyactive2+ 70 : 30 composites produced by co-precipitation method. The existence of HAP nanocrystals showed strong ability to promote the calci"cation of the composites compared with the pure polymer. In this paper, solvent-cast technique was employed to make PLA/d-HAP nanocomposites with special design to achieve good dispersion of nanocrystals in the polymer matrix. High molecular PLA was chosen as material matrix to endow the nanocomposites with good mechanical strength. Transmission electron microscopy (TEM) was used to evaluate the dispersion of nanocrystals in the dimethyl formamide (DMF) solvent and the polymer PLA matrix. The in#uence of d-HAP loading on mechanical properties of the materials were also determined by tensile tests.
2.2. Synthesis of high molecular PLA High molecular PLA was synthesized in bulk using Al(i-Bu) /H PO /H O as the initiator [15]. The polym erization was carried out under nitrogen atmosphere at the temperature above the melting point of D,L-lactide. The resulting polymer was then dissolved in CHCl , and recovered by precipitation in excess diethyl ether. The puri"ed product was dried under vacuum at 403C for 48 h. The weight average molecular weight was 7.3;10, as determined by intrinsic viscosity measurement in tetrahydrofuran at 373C using the relation []" 1.04;10\ M . The molecular weight distribution was 2.1 determined by gel permeation chromatography (GPC). 2.3. Dispersion of d-HAP nanocrystals in DMF Approximately 2 g of d-HAP nanocrystals was dispersed in 200 ml DMF using ultrasonics for 30 min. The dispersion was then "ltered for removal of large aggregates and a blue sol "ltrate was obtained. The accurate concentration of the sol was determined by precipitation 10 ml of the sol into an excess of ethyl ether, followed by weighting the precipitate. 2.4. Preparation of PLA/d-HAP nanocomposites PLA/d-HAP nanocomposites were prepared through solvent-cast technique. Pre-weighted PLA was added into a #at-bottom #ask that contained the d-HAP sol. The #ask was sealed o! and stirred at room temperature until PLA totally dissolved into the sol. The resulting mixture was poured into an aluminum mould and then heated to 1403C for 1 h in a thermostat oven for removal of the solvent. The "nally obtained nanocomposite "lm was furthered dried under vacuum at 803C for 12 h. 2.5. Transmission electron microscopy (TEM)
2. Materials and methods 2.1. Preparation of d-HAP nanocrystals d-HAP nanocrystals were synthesized by a hydrothermal method described elsewhere [11]. A (NH ) HPO aqueous solution of 200 ml (11.4 wt%) was slowly dropped into a stirred, 400 ml Ca(NO ) aqueous solution (16.8 wt%). The pH for both solutions was 10}12, adjusted with ammonium hydroxide solution, and the reaction was carried out at room temperature. The resultant precipitates were put into an autoclave in a solid-solution ratio of 2 wt% and hydrothermally treated at 1403C and 0.3 MPa for 5 h, followed by centrifugal washing with deionized water. The hydrothermal slurry was "nally dried and a white power was obtained.
TEM was used to evaluate the dispersion of d-HAP nanocrystals in the DMF solvent and the PLA polymer matrix as well. All microscopic investigations were performed on a JEOL JEM-100CX TEM operating at 100 kV. The sol sample was visualized directly by dripping a drop into a gold TEM grid (hexagonal 400 mesh), which was dried completely on "lter paper at room temperature. The bulk nanocomposites were obtained from several pieces of composite "lms that were overlapped, and then treated under pressure at 1403C to melt into thick pellets. These pellets were cut to form a triangular block face (approximately 1 mm;0.5 mm) for microtoming. Ultrathin sections were microtomed from this face, at room temperature, using a ultramicrotome. A water-"lled boat
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was attached to the knife, so that after cutting, the ultrathin sections could be #oated onto water. The sections were collected on gold TEM grids and observed after drying in air. 2.6. Tensile tests The stress}strain curves of solution-cast "lms were obtained at 153C on an Instron 4302 machine. The "lms were 0.1 mm thick and tested at a stretching speed of 50 mm/min. At least "ve specimens were measured and the mechanical tensile data were determined from the curves on the average of three specimens. 2.7. Scanning electron microscopy (SEM) SEM was performed with an AMRAY 1000B equipment operated at 18 kV to examine the fracture surfaces of the nanocomposites. All samples were coated with a thin layer of gold by means of a polaron sputtering apparatus.
3. Results 3.1. Characterization of d-HAP nanocrystals The hydrothermally formed d-HAP nanocrystals are rod-like particles with the length between 40}80 nm and the width between 20}40 nm. The Ca/P ratio and the surface area of these nanocrystals were determined as 1.61 by an atomic absorption spectrometer for calcium combined with an ultraviolet spectrophotometer for phosphorus, and 80 m/g by BET method, respectively. 3.2. Dispersion of d-HAP nanocrystals in DMF and the PLA polymer matrix In order to prepare PLA/d-HAP nanocomposites through solvent-cast technique, the solvent selected to disperse HAP nanocrystals should be able to dissolve the PLA polymer. Several PLA solvents such as chloroform, dichloroethane, benzene and DMF were studied to evaluate their dispersive abilities for the nanocrystals. It was found that DMF is the best solvent to disperse the d-HAP nanocrystals. The acquired dispersion was a stable, blue sol that could preserve for several days with only slight precipitation. Fig. 1a is the TEM image of d-HAP nanocrystals dispersed in the DMF solvent. As is shown that most particles are detached from each other with only slight aggregation emergence. The obtained nanocomposites were semitransparent "lms with the thickness about 0.1 mm. Fig. 1b shows a typical bright-"eld TEM micrograph of the nanocomposite containing 1.8 v% d-HAP nanocrystals. It is
Fig. 1. TEM micrographs of d-HAP nanocrystals: (a) dispersed in the DMF solvent; (b) dispersed in the PLA polymer matrix (containing 1.8 v% d-HAP); (c) higher magni"cation of nanocrystals in (b).
shown that homogeneous distribution of the nanocrystals at a microscopic level is observed in the PLA matrix. So far this dispersion level is the best working result for polymer/HAP nanocrystals composites. Fig. 1c is a higher magni"cation photo of the nanocrystals in Fig. 1b. Close contact between the polymer and the d-HAP nanocrystals is observed with no cracks or voids observed in the interface of two phases. 3.3. Mechanical properties of PLA/d-HAP nanocomposites The stress}strain behavior of PLA/d-HAP nanocomposites were characteristic of quasi-brittle materials, all specimens breaking at a point shortly after the yield
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Fig. 2. Dependence of tensile modulus on d-HAP loading for PLA/dHAP nanocomposites; the solid line was calculated from Eq. (5).
Fig. 4. Fracture surfaces of (a) pure polymer; (b) the nanocomposite containing 5.3 v% d-HAP.
Fig. 3. Dependence of yield stress on d-HAP loading for PLA/d-HAP nanocomposites.
maximum on the stress}strain curves but prior to neck formation. The relationship between the d-HAP loading and the tensile modulus for the nanocomposites is illustrated in Fig. 2. The presence of the "lled nanocrystals substantially increase the tensile modulus relative to the pristine polymer. For the nanocomposite containing 10.5 v% dHAP, the modulus even reaches to 2.47 GPa as compared to 1.66 GPa for pure PLA. Fig. 3 shows the dependence of yield stress on d-HAP loading. It is observed that the yield stress for the nanocomposites varies irregularly with d-HAP loading. In a "rst approximation, the yield stress is almost unaffected by d-HAP loading. 3.4. Fracture surface observation by SEM The fracture mode of the nanocomposites was quite similar to that of the pure polymer. Fracture usually initiated at the stress concentration at one end of the fracture surface and propagates perpendicularly against
the loading direction. Fig. 4a and b shows the fracture surfaces of the nanocomposite containing 5.3 v% d-HAP and the pure polymer, respectively. In the middle of the photos, a macroscopic shear ribbon perpendicular to the direction of crack growth is observed for both specimens. Several such shear ribbons could be observed along the whole fracture surface. The fracture surfaces between two shear ribbons indicate brittle fracture and appear much rougher for the nanocomposite than for the pure polymer.
4. Discussion 4.1. Praparation of d-HAP nanocrystals and PLA/d-HAP nanocomposites The dispersion degree of d-HAP slurry after hydrothermal treatment was greatly decided by synthesis conditions such as the initial solution concentrations, their pH values and so on. A relatively strong basicity for the reaction solutions was found to be bene"cial for obtaining less-aggregated hydrothermal slurry. Actually, only under well-controlled synthesis conditions, the
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"nally obtained nanocrystals could turn into a blue sol in the DMF solvent. The dried d-HAP nanocrystals were redispersed in organic solvent to prepare PLA/d-HAP nanocomposites through solvent-cast technique. Among the several PLA solvents studied, such as chloroform, dichloroethane, benzene and DMF, DMF is the best to disperse the nanocrystals. As shown in Fig. 1a, well-dispersed nanocrystals were observed in the TEM image. The dispersion was a blue sol and remained stable for several days with only slight precipitation. This feature o!ers possibility to prepare real PLA/d-HAP nanocomposites through solvent-cast technique. It seems that the solvents with a greater polarity are more e!ectively to disperse the HAP nanocrystals than those with a lower polarity. However in dispersing media such as water or methanol, the polarity of which is larger than that of DMF, the dispersion of the nanocrystals is much poorer than that in the DMF solvent. This fact suggests that the polarity of the solvents is not the only or the dominating factor that determines their dispersive abilities. The well-dispersed nanocrystals in the DMF solvent may be ascribed to its speci"c molecular structure, which is e!ective to stable the HAP nanocrystals. The TEM observation of the nanocomposite indicates good dispersion of d-HAP nanocrystals in the polymer matrix (Fig. 1a and b). Since the nanocomposites were prepared through solvent-cast technique, the gradual removal of the solvent from composite solutions may induce serious agglomeration of the nanocrystals. However, this case had not actually occurred. A possible reason for this may be due to the high viscosity of the solutions resulting from PLA component that e!ectively prevents d-HAP nanocrystals form agglomeration. 4.2. Advantages of PLA/d-HAP nanocomposites as bone implant materials An excellent bone implant material should be able to rapidly induce osteogenesis and form tightly bony bonds with host bone. In mature bone HAP nanocrystals are all irregular shaped thin plates of carbonate apatite with average lengths and widths of 50;25 nm and thicknesses of 2}3 nm [16}18]. Further, natural apatite is a Cade"cient hydroxyapatite with the Ca/P ratio lower than 1.67. These characteristics have little similarity to those for synthetic HAP commonly used in the form of polygonal sintered coarse particles with polycrystalline structure. From the view of biomemetics, better osteogenesis would be achieved if synthetic HAP has more similarity to bone mineral in composition, crystal structure, crystallinity and morphology [19}21]. In this paper, our hydrothermally synthesized HAP is Ca-de"cient rod-like nanocrystals with the length between 40}80 nm and the width between 20}40 nm. These nanocrystals has much more similarity to natural bone mineral in the mentioned
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compositional and morphological aspects and therefore better osteoconductivity for the nanocomposites is expected. Another advantage for the PLA/d-HAP nanocomposites as bone implant materials is that they could be totally assimilated in the human body. Thus, ideal bone implant materials may be produced from these nanocomposites that could rapidly form bony bonds with host bone and gradually be replaced by the production of new bone. 4.3. Mechanical properties of PLA/d-HAP nanocomposites The incorporating of d-HAP nanocrystals into the PLA polymer matrix could signi"cantly improve the rigidity of the materials (Fig. 2). This increase in modulus with "ller content could be theoretically predicted by assuming the nanocomposites as short "ber "lled systems. For a composite containing unidirectional arrayed "bers, the e!ect of "ller on moduli could be described using Halpin}Tsai equations [22]: 1!K* < E " E , * 1#2(l /d )K < * 1#2K2 < E2 " E , 1!K2 <
(1) (2)
with (E /E )!1 K* " , (3) (E /E )#2(l /d ) (E /E )!1 K " . (4) 2 (E /E )#2 Where E* and E2 are the composite moduli in the longitudinal and transverse direction of the "bers orientataion, respectively; E and E are the matrix and the "ller moduli, respectively; V is the volume fraction of the "ller; l is the average length of the "ller; d is the average thickness of the "ller. However for the cases of PLA/dHAP nanocomposites, the spatial orientation of the nanocrystals is not a one-dimensional alignment but a random array in three dimensions. For such a composite, the modulus could be estimated from below experimental equation on the basis of Eqs. (1) and (2) [23]: 3 5 E " E* # E2 . 8 8
(5)
The solid line in Fig. 2 is the prediction of the modulus for the nanocomposites obtained from Eq. (5). In the calculation, the moduli of the PLA matrix (E ) and the
d-HAP nanocrystals (E ) are taken as 114.0 and 1.66 GPa, respectively [24]; the value of l /d for the nanocrystals is taken as 3. It is seen that the experimental values fall on the solid line indicating the good agreement between prediction and experiment.
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The yield stress of the nanocomposite varies little with d-HAP loading (Fig. 3). This is generally the case when there is good adhesion between the polymer and the "ller [25,26]. Thermal gravimetic analysis demonstrated that there was 6 wt% organic matter on the surfaces of the d-HAP nanocrystals that were separated from the nanocomposites. This organic layer, mainly composed of PLA molecules, are tightly bonded onto the surfaces of d-HAP nanocrsystals indicating good adhesion between the matrix and the "ller. Further, the preservation of strength for PLA/d-HAP nanocomposites has close relation with the nanocrystals' dispersion in the PLA matrix. If the nanocrystals aggregate heavily, the internal crack of the aggregate particles would make them no longer bear load, thus resulting in a decreased strength for the nanocomposites. Fortunately, this is not the case for the nanocomposites in which homogeneous disperison of the nanocrystals in a micro-scale is observed (Fig. 1c). 4.4. Fracture mechanism The presence of d-HAP nanocrtystals does not in#uence the fracture mode of the nanocomposites relative to the pristine polymer. Fracture usually initiates at the stress concentration at one end of the fracture surface and propagates with brittle fracture or ductile shear alternatively arranged along the fracture surface. The ductile shear deformation consumes most of the fracture energy and gives rise to shear ribbons perpendicular to the direction of crack growth. The fracture surface for the nanocomposite appears rougher than the pure polymer due to the presence of d-HAP nanocrystals.
5. Conclusions In this paper, preparation of PLA/d-HAP nanocomposites has been realized by solvent-cast technique. Such composites are of great importance to make bone-like substitutes as d-HAP nanocrystals have similar composition, morphology and crystal structure as those of natural apatite crystals. Of all the PLA solvents studied, N,N-dimethylformamide is the best to disperse d-HAP nanocrystals. The obtained dispersion is a stable, blue sol that could preserve for several days with only slight precipitation. This feature o!ers possibility to prepare real PLA/d-HAP nanocomposites through solvent-cast technique. The obtained nanocomposites are semitransparent "lms with the thickness about 0.1 mm. The SEM observation indicates close contact between the polymer matrix and the "lled nanocrystals. The dispersion of nanocrystals in the polymer matrix is homogeneous at a microscopic level. The tensile modulus for the nanocomposites increases with d-HAP loading. Theoretical prediction of the
modulus has been made by assuming the nanocomposites as short "ber "lled systems. The calculation values based on Halpin}Tsai equations show excellent agreement with the experimental results. The yield stress for the nanocomposites has not been undermined by the presence of the nanocrystals. This preservation of strength for PLA/d-HAP nanocomposites may be due to the homogeneous dispersion of d-HAP nanocrystals in the PLA matrix as well as the good interfacial adhesion.
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