Evaluation of Hot-Pressed Hydroxyapatite/Poly-L-lactideComposite Biomaterial Characteristics

Nenad Ignjatovic,1 Edin Suljovrujic,2 Jaroslava Budinski-Simendic,3 Ivan Krakovsky,4 Dragan Uskokovic1

1 Institute of Technical Sciences of the Serbian Academy of Sciences and Arts, Knez Mihailova 35/IV, Belgrade,Serbia, and Montenegro

2 The Vinca Institute of Nuclear Sciences, P.O. Box 522, Belgrade, Serbia, and Montenegro

3 Faculty of Technology, University of Novi Sad, Bulevar Cara Lazara 1, Novi Sad, Serbia and Montenegro

4 Faculty of Mathematics and Physics, Charles University, V. Holesovickah 2, Prague, Czech Republic

Received 4 December 2003; revised 26 March 2004; accepted 26 March 2004Published online 14 June 2004 in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jbm.b.30093

Abstract: Hydroxyapatite/poly-L-lactide (HAp/PLLA) is a ceramic/polymer composite,whose application as a resorbable biomaterial for the substitution and repair of hard bonetissue is widely promising in orthopedic, oral, maxillofacial, and reconstructive surgery. Hotprocessing is a necessary step for obtaining HAp/PLLA composite blocks with mechanicalproperties similar to those of bones. In this article, the changes in structure and physicochem-ical properties of HAp/PLLA composite, hot pressed for different pressing times (5, 15, 30, 45,and 60 min), were studied. Because the morphology of HAp/PLLA composite biomaterial isvery sensitive to this procedure, its surface microstructure was analyzed by scanning electronmicroscopy (SEM) coupled with an energy-dispersive X-ray (EDX) detector system. Struc-tural changes occurring in the material, mostly changes in crystallinity of PLLA, were studiedby wide-angle X-ray structural analyses (WAXS) and infrared (FTIR) spectroscopy. Usingdifferential scanning calorimetry (DSC), thermogravimetric analysis (TGA), and gel perme-ation chromatography (GPC), the influence of hot pressing on the properties, crystallizationkinetics, and decomposition of HAp/PLLA composite biomaterial was analyzed. © 2004 WileyPeriodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 71B: 284–294, 2004

Keywords: composite biomaterials; hydroxyapatite; poly-L-lactide; hot pressing; crystallin-ity; mechanical properties

INTRODUCTION

Bone tissue repairs have been performed until now using anumber of different biomaterials.1 Various composite sys-tems, combining fibrous or particulate components for rein-forcement and organic polymers as matrix materials, aresome of them, and they have already been investigated. Thesecomposites can be classified into three groups (by combina-tions of reinforcing and matrix materials, which can be eitherbioresorbable or nonbioresorbable): both nonbioresorbablereinforcing and matrix components; bioactive particles forreinforcement with nonbioresorbable polymer as a matrix;bioactive but not totally resorbable reinforcing componentswith a bioresorbable matrix, in which complete resorption

still cannot be acquired.2 Substitution and repair of hard bonetissue are to be made by biomaterials which properties suchas biocompatibility, nontoxicity, and numerous biomechani-cal ones have to satisfy the requirements of a defect to berepaired. On the other hand, mechanical properties similar tothose of natural bones are also necessary.3 Good adhesion ofthe surrounding cells with the biomaterial surface to establisha hard connection in the implant–tissue interlayer is a pre-requisite for the successful repair.

Reinforcement of the ceramic phase by bioresorptive poly-mers results in the formation of bioresorptive composites,whose ceramic phase after application remains unchangedwhile the polymer resorbs with time, leaving space to new-formed body tissue.4 Chemically synthesized calcium hy-droxyapatite (HAp) is similar to the ceramic component ofhuman bone tissue.5 Bioresorptive polylactides (PLA) andtheir copolymers belong to the group of nontoxic polymers,because the final products of their degradation (CO2 andH2O) enter without difficulty three-carboxylic acids cycle,not disturbing the metabolism of the surrounding tissue in the

Correspondence to: D. Uskokovic (e-mail: uskok@itn.sanu.ac.yu)Contract grant sponsor: the Ministry of Science, Technology and Development of

the Republic of Serbia; contract grant number: No. 1431 (Molecular designing ofmonolith and composite materials)

© 2004 Wiley Periodicals, Inc.

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process.6 Compared with pure hydroxyapatite, compositebiomaterials of HAp/PLLA type (or its copolymers) induce,on their surfaces, formation of a larger number of osteoblastsnecessary for repair of bone defects. The concentration ofalkaline phosphates, needed for osteoblast differentiation, ishigher near HAp/PLLA composite biomaterial (or its copol-ymers) than near pure PLLA.7 Chemically synthesized HApenables on its surface good adhesion, differentiation, andgene expression of osteoblasts, as well as absorption ofproteins responsible for the adhesion.8 Response of the or-ganism to the composite biomaterial implanted depends onnumerous factors; the most important among them are bio-compatibility, porosity (size and distribution of pores), sur-face microstructure, etc. In addition, the behavior of HAp/PLLA composite biomaterial depends on the characteristicsof HAp and PLLA also. Bioresorption time of PLLA dependson the ratio and distribution of amorphous/crystalline phasein the polymer, their molecular weights, polymer porosity,etc. Generally, a decrease in polymer crystallinity decreasesthe bioresorption time.9 HAp/PLLA composite biomaterialwas obtained by synthesis and subsequent polymerization orforging of a HAp and PLLA mixture.2,3 Blocks of HAp/PLLA composite material of required densities and mechan-ical characteristics were obtained by hot pressing at the PLLAmelting temperature.10–12 These blocks exhibited extremelyhigh mechanical characteristics.13 During hot pressing ofhighly porous HAp/PLLA composite biomaterial and itstransformation into a nonporous one, physicochemicalchanges occurred in the PLLA phase, while the HAp phaseremained stable. PLLA molecular weight decreased duringthermal and mechanical treatments.14

Nazhat et al.15 also obtained a hydroxyapatite/polylactidecomposite by hot pressing. The aim of this work was to studythe effects of the hot pressing on the structure and character-istics of HAp/PLLA composite biomaterial. The reason forusing scanning electron microscopy (SEM) coupled withenergy-dispersive X-ray analysis (EDX) was to determine thechanges in surface microstructure. Structural changes occur-ring in the material were studied by wide-angle X-ray struc-tural analyses (WAXS) and infrared (FTIR) spectroscopy.Also, the aim was to point out the changes in crystallinity andthermal properties of the polymer (PLLA) phase, being veryimportant for in vitro and in vivo examinations, as well as forthe clinical use of this biomaterial.

MATERIALS AND METHODS

Calcium-hydroxyapatite was obtained by precipitation of cal-cium nitrate and ammonium phosphate in an alkaline me-dium. The gel was dried at room temperature and calcined at1100°C for 6 h. HAp powder of a 100-nm average particlesize, obtained by grinding the HAP granules, was used as aconstituent in processing of the composite biomaterial. Com-mercial poly-L-lactide (PLLA, Fluka, Germany) with a mo-lecular weight of 100,000 g/mol (Mw) was used as a polymercomponent in the study. After complete dissolution of PLLA

in chloroform, HAp powder was added and thus obtainedmixture was vacuum evaporated, according to the previouslydescribed procedure.10

HAp to PLLA ratio in all samples was 80:20. PLLAfraction of 20 mass % represents a volume fraction of 41%.10

Natural bone tissue consists of 60 vol % of inorganic phase(mostly HAp) while remaining 40 vol % is connective tissue(of polymer nature). By HAp to PLLA weight ratio in thecomposite material, the concept of natural bone is preserved.

Highly porous HAp/PLLA composite in cylindrical molds(10 mm in diameter) was compacted by hot pressing(CARVER Press Inc., Auto C Model 3889) at a temperatureof 176 � 1°C and pressure of 98.1 MPa, for 5, 15, 30, 45, and60 min. Two grams of HAp/PLLA composite biomateral waspoured into a mold, previously heated to a predeterminedpressing temperature. Upon insertion of molds, the pressurewas reached for 30 s, and then the pressing was performedwith a pressure hold-up time of 5, 15, 30, 45, and 60 min.After pressing, the pressure was allowed to drop to theatmospheric for 20 s. Subsequently, the samples were pulledfrom the molds for 1 min. The cooling rate was of predom-inant influence on the formation of crystalline regions inPLLA. To minimize this influence, all samples were cooleddown in the same way. After being tipped out, all sampleswere dried in a drying oven at a temperature of 25°C andrelative humidity of 65%. Five hours later, the samples weretaken out from the oven and analyzed.

Microstructure characterization was made by a JSM 5300scanning electron microscope coupled with a QX2000 Ana-lyzer System (Oxford Instruments, UK) for the electron-dispersion analysis (EDX). For SEM analyses, the fracturesurfaces of samples were covered by a thin layer of gold,under vacuum evaporation. Porosity, that is, relative density,of HAp/PLLA composite biomaterial blocks was determinedgeometrically from the mass to volume sample ratio. Wide-angle X-ray structural (WAXS) analyses were made by aPhilips PW 1710 diffractometer with Ni-filtered CuK�1, �2

radiation. The data were collected in the 2� region from 5 to90°, with a step length of 0.03° and exposure time of 4 s perstep. Infrared reflection spectra were obtained at room tem-perature using a BOMEM DA8 Fourier-transform infraredspectrometer in the spectral range between 500–4000 cm�1

(KBr beamsplitter, Globar source, MCT detector). For DSCmeasurements, a Perkin-Elmer Model DSC-2 differentialscanning calorimeter fitted with a data acquisition system wasused. Samples of 10 � 0.5 mg were analyzed in a nitrogenflow by heating (20 K/min) from 310–490 K. Because thechanges in thermogram, in the measured temperature interval,are due to polymeric PLLA component, the HAp/PLLA com-posite thermogram was normalized to the polymer mass (2 �0.1 mg). The enthalpy of melting (�Hm) and the cold crys-tallization (�HC) of the polymer (PLLA) phase, in the HAp/PLLA composite, of each sample were determined (from thearea under endothermal peak for �Hm and exothermal peaksfor �HC), and the degree of crystallinity (�) was calculatedaccording to � � �H0/�Hf � (�Hm � �Hc)/�Hf, where �Hf

is the heat of fusion of a perfectly (100%) crystalline PLLA.16

285HOT-PRESSED HAp/PLLA COMPOSITE BIOMATERIAL CHARACTERISTICS

The presented results are average values for at least fiveidentically prepared samples. The error was calculated on thebasis of mean square deviation. Thermogravimetric (TGA)analyses were made in a nitrogen atmosphere by aPerkinElmer TGA-2 device, in the 20–500°C temperaturerange, and at 10°C/min heating rate.

Gel permeation chromatography (GPC) was used to de-termine the average molecular weights and polydispersityindex (PI) of PLLA. A GPC Device Labio (Czech Republic),equipped with a refractometric detector and Biospher GMB1000 column (size 10 microns), was used for measurements.After hot pressing of HAp-PLLA composites, pure PLLAwas extracted by being dissolved in chloroform and vacuumdried until obtaining a constant weight of the samples. Then,solutions of pure PLLA in tetrahydrofurane (THF) with 2%vol of toluene (as an internal standard) were prepared at 50°Cusing an electromagnetic stirrer (0.02 � 0.001 g of PLLA in1 cm3 of solvent). Calibration was conducted with polysty-rene standards with narrow molecular weight distributions,which covered a molecular weight range of 1000–10,000,000(218,000, 52,000, and 13,700). Tetrahydrofurane was used asa moving solvent. Measurements were preformed at roomtemperature (25°C). Flow rate of mobile phase was 1 mL/min.

Compressive strength and elasticity modulus of the HAp/PLLA composite biomaterial blocks after pressing were de-termined. Compressive strength of cylindrical blocks (h � 10mm and d � 10 mm) was measured on an INSTRON M 1185instrument. The load stress velocity was 1 mm/mm. Theobtained values are arithmetic means for five (n � 5) sam-ples, for each given pressing time.

RESULTS AND DISCUSSION

Scanning Electron Microscopy Analysis

To examine the influence of hot pressing time on the micro-structure and distribution of HAp and PLLA phases, fracturesurfaces of the HAp/PLLA composite biomaterial blockswere studied by SEM. The microstructure of a highly porousHAp/PLLA composite surface (before hot pressing) is shownin Figure 1(a). The surface has spherical and ellipsoidal-cylindrical pores, approximately the same in diameter (2 to 4�m), formed by removal of the solvent during vacuum evap-oration. The microstructure of fracture surfaces shown inFigure 1(b,c) was obtained after 15 and 60 min of hot press-ing of HAp/PLLA, respectively. After 60 min of hot pressing,nonporous HAp/PLLA composite blocks were obtained withHAp and PLLA in very close contact. Very fine distributionof HAp particles in the PLLA base can be seen in Figure 1(c).After hot pressing, PLLA is in intimate contact with HAp.SEM images prove the penetration of PLLA into the poroussurface of HAp granules.11

EDX spectra of HAp and PLLA are presented in Figure 2.Peaks originating from HAp can clearly be seen in the spectrumshown in Figure 2(a). A spectrum with peaks corresponding to

the PLLA phase is shown in Figure 2(b). The spectra presentedin Figure 2 define HAp and PLLA distribution in the fracturesurfaces illustrated by SEM images (Fig. 1).

Wide-Angle X-ray Structural Analysis

Highly porous HAp/PLLA was used as a starting and refer-ence material. In comparison with it, changes occurring in the

Figure 1. SEM images of HAp/PLLA composite biomaterial: (a) be-fore hot pressing; (b) after 15 min of hot pressing; (c) after 60 min ofhot pressing.

286 IGNJATOVIC ET AL.

composite during hot pressing were analyzed. The analysis ofHAp, PLLA, and HAp/PLLA composite biomaterial con-firmed the changes in the polymer phase during hot pressing,while HAp under given conditions remained unchanged.14

Effects of the hot pressing on the structural changes oc-curring in the material, mostly changes in HAp and in thecrystalline phase of PLLA component, were studied byWAXS. Diffraction spectra of PLLA (I crystalline, II amor-phous), HAp, and HAp/PLLA composite are shown in Fig-ures 3 and 4. WAXS confirmed the existence of HAp (also,HAp phase in HAp/PLLA composite) of high crystallinity.14

On the other hand, the most intense peaks of PLLA appear at2� � 15, 17, and 19°, which are comparable with the � form

of PLLA crystallized in a pseudo-orthorhombic unit cell ofdimensions: a � 1.07, b � 0.595, and c � 2.78 nm.17 Broaddiffusion bands observed in the spectra, at small angles (2�from 7 to 30°), were typical of amorphous polymeric (PLLA)component. In the case of HAp/PLLA composite (before hotpressing), the presence of two peaks for the PLLA componentis evident: (1) the most intense one at 2� � 17°; and (2) amuch smaller one at 2� � 19° (Fig. 4). They have a charac-teristic shape, and besides a wide peak corresponding to theamorphous phase of PLLA there is a HAp diffraction line,dominant in all spectra. Namely, all diffraction peaks of HApin the composite biomaterial appear at almost the same anglesand intensity to profile ratio values, before and after hot

Figure 2. EDX spectra: (a) HAp; (b) PLLA.

287HOT-PRESSED HAp/PLLA COMPOSITE BIOMATERIAL CHARACTERISTICS

pressing. It confirms that the applied pressure, temperature,and the presence of PLLA do not affect the HAp crystalstructure (single cell symmetry and crystallite size). There-fore, only changes in PLLA peak, which are not easy tofollow, indicate the changes in HAp/PLLA composite bio-material during compacting. The most intense peak (1) of thePLLA component is in a very narrow angle band togetherwith the diffraction line of HAp, and they completely overlap.Because in the case of HAp not a single change with hotpressing (at applied temperatures and pressures) at 2� � 17°was found, it is evident that the changes in width and inten-sity of peaks of the composite material result from thechanges in the crystal phase of PLLA induced by hot press-ing. Decrease in intensity and different shape of peaks at2� � 17° and 2� � 19° for various processing times (0, 15,

and 60 min) are evident by comparing the diffractograms[Fig. 4(b)]. The intensity and sharpness of the PLLA peak at2� � 17° decrease continuously with increasing processingtime (to 60 min). After 15 and 60 min of hot pressing, the areaof peak 1 (2� � 17°) decreases 1.5 and 1.85 times, respec-tively. The given results indicate a significant decrease inPLLA crystallinity during hot pressing. On the other hand,hot pressing of highly porous HAp/PLLA composite bioma-terial at the PLLA melting temperature for 60 min, yielded acomposite biomaterial of very high compressive strength andelasticity modulus.13

Differential Scanning Calorimetry Analysis

To get a more complete picture of the changes registered byWAXS, DSC measurements of HAp/PLLA biocompositebefore and after hot pressing were performed. The results areillustrated in Figures 5, 6, and 7. DSC curves of HAp/PLLAsamples before and after hot pressing, similar to the onesobtained for pure PLLA by other authors,16,18,19 are given inFigure 5. All changes recorded in the biocomposite originatefrom PLLA, because the HAp phase in the temperature rangestudied is stable.14 The crystallization behavior of pure PLLAand morphology of PLLA crystals were well summarized byMiyata.20,21 The curves (Fig. 5) indicate the presence of a fewcharacteristic transitions typical of pure semicrystalline polyL-lactide samples: (1) glass transition, (2) cold crystallization,and (3) melting. Except for certain similarities, DSC curveshave significantly different shapes, positions, and peak areas,which are the consequences of structural changes in amor-phous/crystalline fraction and changes in these fractions, in-duced by hot pressing. A decrease in temperature of glasstransition Tg, cold crystallization (TC1 and TC2) and meltingpoint Tm of the PLLA component with increasing hot press-

Figure 3. Diffractograms of PLLA [crystalline (I) and amorphous (II)],HAp and HAp/PLLA composite (before hot pressing and after 60 minof hot pressing).

Figure 4. (a) Diffractograms of PLLA [crystalline (I) and amorphous (II)], HAp and HAp/PLLA com-posite (b) diffractograms of HAp/PLLA composite before hot pressing and after 15 and 60 min of hotpressing.

288 IGNJATOVIC ET AL.

Figure 5. DSC curves of the HAp/PLLA composite biomaterial beforeand after 5, 15, 30, and 60 min of hot pressing.

Figure 6. (a) Melting (Tm), cold crystallizations (Tc1 and Tc2) and glass transition temperatures ofHAp/PLLA samples as functions of hot pressing time. (b) The enthalpy of melting (�Hm), coldcrystallization enthalpy (�HC), and real enthalpy (�H0) as functions of hot pressing time.

Figure 7. Crystallinity of the polymer (PLLA) component (of HAp/PLLA samples) as a function of absorbed dose calculated on polymer(PLLA) component weight part (I) and on HAp/PLLA composite weight(II). Calculation was made for two different values of heat of fusion ofa perfectly (100%) crystalline PLLA (�Hf1 � 93.7 Jg�1 is given byShikinami2 (�) and �Hf2 � 142 Jg�1 given by Loomis15 (■).

289HOT-PRESSED HAp/PLLA COMPOSITE BIOMATERIAL CHARACTERISTICS

ing time is evident [Fig. 6(a)]. The enthalpy of melting(�Hm), cold crystallization (�HC), and the real enthalpy(�H0) were determined [Fig. 6(b)], and the degree of crys-tallinity (�) was calculated according to � � � �H0/�Hf �(�Hm � �Hc)/�Hf, where �Hf1 � 93.7 Jg�1 is the heat offusion of a perfectly (100%) crystalline PLLA given byShikinami2 and �Hf2 � 142 Jg�1 given by Loomis.16 Figure7 shows the results of the changes in crystallinity of PLLA,calculated for the polymer weight part (I) and the weight ofthe whole HAp/PLLA sample (II). Rearrangement and crys-tallization of PLLA on the HAp particle surfaces duringcomposite preparation change the melting temperature ofPLLA, its enthalpy, and crystallinity. After hot pressing, amore compact HAp/PLLA composite biomaterial was ob-tained. Decrease in porosity and improvement in mechanicalproperties of the biomaterial are the result of hot pressing,due to easier flow of PLLA around HAp particles and highpressure applied.11,12 Due to thermomechanical factors,breaking of chains and, therefore, a decrease in PLLA mo-lecular weight took place during hot pressing.14 After hotpressing, PLLA of lower molecular weight was obtained,whose crystallization proceeds according to the mechanismdifferent from that for untreated PLLA. Different mecha-nisms of crystallization of PLLA in HAp/PLLA (from themelt and solution) lead to different appearance, patterns, andsizes of the crystals. This reflects on the crystallinity degreeand given composite melting temperature. The most signifi-cant effects caused by hot pressing are the changes in prop-erties of PLLA in the HAp/PLLA composite biomaterial. Theobtained melting enthalpy and crystallinity degree are con-siderably lower than those of untreated HAp/PLLA compos-ite biomaterial. These results are in accordance with the givenWAXS results. After 30 min of hot pressing, blocks with aporosity of 1.4 � 0.2% were obtained. After 60 min of hotpressing, the produced blocks have a density close to thetheoretical density and porosity of 0.4 � 0.2%. It is supposedthat porosity besides previously specified factors affects thecrystallization mechanism, melting enthalpy, and crystallinitydegree as well. Under applied pressure, it comes to the strainin PLLA globular molecules, their unfolding, and the gener-ation of linear forms. A decrease in crystallinity of the PLLAphase surrounding HAp particles under high pressure wasreported earlier by Shikinami.2 Lower crystallinity of PLLAin the HAp/PLLA composite compared with pure PLLA andthe formation of larger amorphous regions were found tooccur due to separation and decrease in PLLA crystal regionscaused by HAp particles under high pressure. HAp particlesplay a significant role in these changes. Due to temperaturegradient, in hot processed HAp/PLLA composite, strainedand unfolded PLLA globular molecules exist around the HApparticles (different heat capacities). It is noticeable that crys-tallinity decreases with increasing hot pressing time.

Infrared Spectroscopic Analysis

A decrease in PLLA molecular weight occurs during hotpressing.14 Dependence of the stability of HAp/PLLA com-

posite biomaterial blocks on the hot pressing time (15 and 60min) was studied by IR spectroscopy. IR spectra of HAp/PLLA composite biomaterial before and after 15 and 60 minof hot pressing are presented in Figure 8.

A spectrum of HAp/PLLA composite biomaterial, beforehot pressing, consists of spectral lines originating from itsconstituents HAp and PLLA as explained earlier.10,22–24

PLLA is characterized by an absorption band at 1760 cm�1

arising from the stretching vibrations of (CAO) group, aswell as bends ascribed to the bending vibrations of (CH3)group at 1386 and 1453 cm�1 and stretching vibrations of(COH) group at 2986 and 2933 cm�1. HAp is characterizedby a triplet of absorption bands at 626, 600, and 566 cm�1

arising from (OH) groups, doublet at 1093 and 1040 cm�1

from (PO4) groups, and an absorption band at 3573 cm�1

arising from the (OH) group. The spectra of HAp/PLLAcomposite biomaterial, obtained after 15 and 60 min of hotpressing, are slightly different from those registered beforehot pressing. Positions and intensities of absorption bands ofall three composite biomaterials are also slightly different.According to IR spectra, after 60 min of hot pressing, neitherformation of new phases nor disappearance of PLLA wererecorded. The changes occurring in PLLA during 30 and 60min are those already noted, that is, decrease in molecularweight14 and changes in temperature, melting enthalpy andcrystallinity degree. Nonappearance of new absorption bandsin the range of wave numbers from 400 to 4000 cm�1 in thespectra of HAp/PLLA composite biomaterial samples after15 and 60 min of hot pressing, confirms insignificant influ-ence of hot pressing on the stability of the material.

Thermogravimetric Analysis

Effects of the hot pressing time on the HAp/PLLA compositebiomaterial thermal stability were determined by thermo-gravimetric (TG) analyses. Compacting of HAp/PLLA com-posite biomaterial at the PLLA melting temperature, affectsthe PLLA thermal stability only, because the HAp phaseremains stable.14 TGA curves for HAp/PLLA composite bi-omaterial were compared with TGA curves for pure HAp and

Figure 8. FTIR reflection spectra of HAp/PLLA composite biomaterial(a) before and (b) after 15 and (c) 60 min of hot pressing.

290 IGNJATOVIC ET AL.

PLLA [Fig. 9(a)]. In the given temperature interval, the HApphase is constant and only mass loss of PLLA was regis-tered.14 PLLA from the HAp/PLLA composite biomaterial,compared with pure PLLA, shows greater stability at tem-peratures up to 200°C. The reason for such a behavior aremost probably HAp particles, which in the composite act asbarriers preventing heat transfer, as well as cavities whichdetermine the system porosity, due to differences in heattransfer coefficients.25 Because of nonstationary heat conduc-tivity in the given temperature range, HAp particles andporosity cause seemingly higher thermal stability of PLLA.Above 470 K and at 10°C/min heating rate, the stationaryheat transfer through the sample and thermal degradation tocomplete mass loss of PLLA occur. Thermal stability ofPLLA from the composite is different and it decreases withincreasing hot pressing time [Fig. 9(b)]. PLLA is a thermallyvery sensitive polymer. During thermal degradation of poly-mers, it comes to the breaking of bonds in the basic chain,which results in the formation of oligomeric fragments thatfurther on degrade.26 As a consequence, molecular weights ofpolymers decrease while in the subsequent degradationstages, monomers and gas-products are formed. Our previousstudy confirmed a decrease in PLLA molecular weight afterhot pressing of 60 min, but a total degradation was notregistered.14 Owing to the planar conformation of PLLA andits basic structure defined as (OOOCH(CH3)OCOO)n onecan predict where the breaking of bonds in the basic chainwill occur.27 Thermal degradation of PLLA is a one-stepprocess with the first-order reaction kinetics.28 According tothe model given by McNeil and Lepier,29 cyclic oligomers,lactide, acetaldehyde, and carbon monoxide are the finalproducts of PLLA thermal degradation. Thermal degradationis dictated by the final OH groups that “attack” the basicchain, which bends under the influence of thermal energy.

Such a reaction can start at temperatures higher than 500 K.30

In each of these reactions, the OH group regenerates and theprocess continues. Depending on the basic chain spot “at-tacked” by the OH group, different products are obtained.Consolidation by hot pressing is characterized not only byhigh temperature but by high pressure as a function of time aswell.13 Due to mentioned reasons, the hot pressing time hasan important influence on the PLLA molecular weight and,therefore, on the stability of the obtained blocks. The de-crease in PLLA molecular weight is induced by hot press-ing.14 With decreasing molecular weight, PLLA chains be-come more flexible and mobile and, therefore, thermally lessstable.30 According to TGA curves given in Figure 9, HAp/PLLA composite biomaterial blocks get energy during hotpressing from outside (pressure, heat), which causes a de-crease in size of PLLA chains and, therefore, their stability.The obtained blocks will become less stable if the time of hotpressing is longer.

Gel Permeation Chromatography Measurements

Normalized fraction distributions of number-average molec-ular weight Mn for PLLA extracted from HAp-PLLA com-posites after different time of hot pressing are summarized inFigure 10. The curves presented in Figure 10 were normal-ized recalculated, which means that the obtained numericalfraction was transformed to the mass fraction of PLLA.

It can be seen that during hot pressing, despite a degrada-tion of PLLA, changes in average molecular weight areevident. The changes of normalized distribution functions fordifferent samples (from the highest to lower molecularweights and decrease in fractions of high molecular weightswith increasing hot pressing time) indicate that chain scissionof PLLA, gives rise to ratio of small molecular weight oli-gomers.

Figure 9. (a) TGA curves of PLLA (before hot pressing), HAp, and HAp/PLLA composite biomaterial;(b) TGA curves of HAp/PLLA before and after 5, 30, and 60 min of hot pressing.

291HOT-PRESSED HAp/PLLA COMPOSITE BIOMATERIAL CHARACTERISTICS

It is evident from the results presented in Figure 9 thatlonger hot pressing time decreases the thermal stability ofPLLA, according to the previously discussed thermome-chanical degradation mechanisms. Final products are unsta-ble polymers with shorter chains, as confirmed by the resultspresented in Figure 10. Curve 1 (Fig. 10) for the polymerbefore hot pressing shows the existence of a wider gamut ofmolecular weights of PLLA than that of curve 5 (60 min ofhot pressing). In this way, a polymer was obtained withnarrow distribution of molecular weights (curve 5) close to amaximum of 58,000 g/mol. The presence (Wg5 � 5.75 �10�6) of a greater number of PLLA of lower molecularweights (Mn5 � 58,000) after 60 min of hot pressing com-pared to PLLA before hot pressing (Wg5 � 4.65 � 10�6 andMn5 � 80,000) is in accordance with lower thermal stabilityof PLLA, shown in Figure 9.

Mechanical Properties

Mechanical strength, elasticity modulus, as well as porositycontent and distribution play a significant role in implantationand exploitation of the biomaterial. To achieve an optimumdistribution of stress occurring during strain and load of theimplant it is necessary that the used HAp/PLLA compositebiomaterial exhibits mechanical properties similar to the bonetissue it substitutes.

Some authors point out that mechanical properties of thiskind of composites depend on the weight content of HAp andPLLA.32–35 These properties are important for various com-posite applications as well as in vitro and in vivo research.34

A change in HAp weight content causes changes in thecrystallinity degree that have a significant impact on themechanical properties.15 However, this study did not consider

a possible increase in mechanical properties with the samecontent of HAp and PLLA, as we did. By varying the basicparameters of hot pressing, a wide spectrum of HAp/PLLAbiocomposite blocks of different properties was obtained.The change in temperature, pressure, and time of hot pressingdirectly affects the compressive strength and elasticity mod-ulus of the blocks. Figure 11 shows the values of compressivemodules and compressive strengths achieved for different hotpressing times.

Pressing at the melting temperature of the polymer pro-vides efficient compacting followed by a continual decreasein porosity. Compressive strength and modulus could not bedetermined before hot pressing because the composite wasnot compacted. However, already after 15 min of hot pressing

Figure 10. Normalized fraction distributions of number-average molecular weights Mn for PLLAextracted from HAp–PLLA composites before and after hot pressing for different times.

Figure 11. Dependence of the compressive modules and compres-sive strength of the HAp/PLLA composite biomaterial blocks on thehot pressing times.

292 IGNJATOVIC ET AL.

cylindrical blocks exhibit an average compressive strength of90 MPa and modulus of 4.0 GPa. After 60 min, compressivestrength is higher and amounts to 123 MPa and modulus to9.7 GPa.

The maximum modulus of 9.7 GPa was registered with theblocks exhibiting the maximum compressive strength after 60min of compacting. As evident, the compressive strength ofHAp/PLLA composites increases with prolonged hot press-ing time. Natural bone elasticity modulus is in the interval of0.09–18.6 GPa and compressive strength in the range of1.9–167 MPa.3

Using PLLA of 430,000 g/mol molecular weight (Mw),compressive strength of 140 MPa and modulus of 10.0 GPawere obtained.13

The blocks with modulus of 9.7 GPa and compressivestrength of 123.0 MPa are very similar to the natural bonetissue.

By prolonging the hot pressing time, the composite poros-ity decreases, while the compressive strength increases asexpected.13 The results obtained by WAXS, DSC, and TGanalysis indicate that, besides macrochanges in compositeporosity, structural changes in PLLA phase occur, resulting inchanges in compressive strength and modulus. Under theinfluence of increased pressure and temperature during hotpressing, globular molecules of PLLA unzip into more linearforms with changed chain orientation and density. PLLAchains orientation and packing density have a significantimpact on the final mechanical properties.

With prolonged hot pressing time, over 60 min, an in-crease in compressive strength and modulus can be expecteduntil significant decrease in molecular weight of PLLA takesplace.

The connection between HAp and PLLA in the interfacesurface, before the hot pressing, is of weak ion nature and ismost probably established between the Ca2� ion and oxygenfrom PLLA.35 A possible improvement in this connectionduring hot pressing is not excluded.

CONCLUSION

Designing of HAp/PLLA composite biomaterial block prop-erties was achieved by combining high pressure, temperature,and time. Hot pressing causes a decrease in crystallinity ofPLLA in the HAp/PLLA composite biomaterial as proved byWAXS and DSC analysis. The WAXS results indicate asignificant decrease in PLLA crystallinity during hot press-ing. DSC analyses confirmed these changes in crystallinitydegree but also indicated changes in glass transition, coldcrystallization, and melting temperature of the PLLA poly-mer. In the given time interval of hot pressing from 0 to 60min, insignificant qualitative changes in the HAp and thePLLA phase were recorded by IR spectroscopy. This com-posite has higher degree of crystallinity than hot pressedblocks, and therefore their bioresorption is potentially longer.Because of that, it would be necessary to examine, in in vivoconditions, the properties the HAp/PLLA composite bioma-

terial blocks obtained after different times of hot pressing,which will be the subject of our further work.

During hot pressing, despite PLLA degradation, smallchanges in average molecular weights are evident. Thechanges of normalized distribution functions for differentsamples indicate that chain scission of PLLA gives rise toratio of smaller molecular mass oligomers. HAp/PLLA com-posite biomaterial blocks with a compressive strength of 123MPa and E-modulus of 9.7 GPa, the value close to that of thenatural bone, were obtained by hot pressing of 60 min.

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