Investigation of sintering temperature and concentration effects on sodium doped hydroxyapatite.
Abstract: Sodium doped hydroxyapatite (HA) (of different mol%) was calcined at four different temperatures for observing the sintering behavior and concentration effects. Doped and pure apatites were synthesized from waste egg shell as Ca precursor through precipitation method. The substitution limit was up to 8mol%. Atomic absorption (AAS) and UV spectrophotometric methods were followed to detect the presence of Na, Ca, P etc. Quantitative analysis was carried out by X-ray fluorescence (XRF) and electron diffraction spectroscopy (EDS). The influence of sintering temperature on phase composition was evaluated by using Fourier transform infrared spectrometer (FT-IR), X-ray diffraction (XRD) and Scanning electron microscopy SEM techniques. Crystallographic values were calculated and comparable discussion was presented. Observed data were also in excellent agreement with the standard JCPDS values for HA.
Article Type: Report
Subject: Sintering (Research)
Hydroxylapatite (Chemical properties)
Hydroxylapatite (Observations)
Authors: Kabir, Sumaya F.
Ahmed, Samina
Ahsan, Mainul
Mustafa, Ahmad I.
Pub Date: 04/01/2012
Publication: Name: Trends in Biomaterials and Artificial Organs Publisher: Society for Biomaterials and Artificial Organs Audience: Academic Format: Magazine/Journal Subject: Health Copyright: COPYRIGHT 2012 Society for Biomaterials and Artificial Organs ISSN: 0971-1198
Issue: Date: April, 2012 Source Volume: 26 Source Issue: 2
Topic: Event Code: 310 Science & research
Geographic: Geographic Scope: Bangladesh Geographic Code: 9BANG Bangladesh
Accession Number: 304842710
Full Text: Introdcution

Among all calcium phosphate minerals Hydroxyapatite (HA) has received significant attention due to its similarity with the inorganic component of in-vivo hard tissue (bone and tooth mineral) [1-3]. In addition HA also possess high biocompatibility and osteoconductivity [2,4]. However, bone minerals is considered to have a structure of pure HA with some impurities ([K.sup.+], [Mg.sup.2+], [Na.sup.+], [F.sup.-] etc) [5,6]. Thus, on mimicking natural apatite it's necessary to dope HA with some ionic substituents. Moreover, the structure of HA is flexible to accommodate a variety of cationic and anionic substituents with a modification in its lattice structure, crystallinity, morphology and thermal stability [1,7,8]. Besides, such alteration greatly enhances the mechanical, chemical and physiochemical properties of synthetic HA [8,9].

Sodium is one of the trace elements available in biological bone and tooth minerals. It plays an important role in cell adhesion, bone metabolism and resorption process [1]. Moreover, Sodium acts as an essential nutrient with important functions in regulating extracellular fluid volume and the active transport of molecules across cell membrane [10]. A certain portion of this element is the prerequisite for better nerve and muscle functioning. To regulate fluids and blood pressure, and to keep muscles and nerves running smoothly, adults are required to take 1,500 mg Na per day. In this regard, the sodium doped apatite would obviously act as a reservoir and carrier of sodium with better response in terms of the biological behavior and implantation. Hardly some researches have conducted on sodium doped hydroxyapatite (Na-HA). Thus our attempt was to synthesize doped HA of different sodium concentration at various sintering temperatures. Besides, to investigate these effect on phase properties of Na-HA.

The developed bioceramic material is expected to have a wide range of application. Mostly older people all over the world suffer from periodontal bone destruction which causes tooth loss, osteoporesis that leads to bone fracture and also from several bone defects [4,11,12]. Our effort is thus definitely a step forward in solving such problems. The synthesized Na-HA would be a promising biomaterial for bone repair, regeneration and also as bone and tooth filler.

Materials and Methods

Synthesis of Na-substituted apatite from egg shell as Calcium (Ca) precursor

In context of our country, all HA used are currently imported. So, our effort was to develop cost effective Na-HA. Besides, selection of raw material is an important factor for any methods to be developed. In this perspective, we choose eggshell as Calcium precursor.

Raw egg shell was thoroughly washed, calcined, powdered and finally characterized as previously mentioned [13]. Analar grade chemicals NaN[O.sub.3], N[H.sub.4]OH, HN[O.sub.3], [(N[H.sub.4]).sub.2]HP[O.sub.4] (99.99% pure) were used in this method, obtained from E. Merck. All the solutions were prepared using double distilled water.

The different mol% Na-HA (Na4HA containing 4 mol% sodium and Na8HA containing 8mol% sodium) was prepared by wet precipitation method. A requisite amount of egg shell powder was dissolved in conc. HN[O.sub.3] and 50 mL of distilled water was added to this acidic egg shell solution and filtered to get clear solution. Final volume was made up to 100 mL with distilled water maintaining the pH of the solution at ~10.0 with aqueous ammonia. The doping solution (NaN[O.sub.3]) was mixed with the egg shell solution prior to the addition of phosphate precursor solution. [(N[H.sub.4]).sub.2]HP[O.sub.4] in ammonia (pH ~ 10.0) was added drop wise to this solution with continuous stirring condition. Yellowish gelatinous precipitates of doped HA was formed which was stirred for overnight in the mother solution for ripening. The precipitate was then filtered through a Buchner funnel and thoroughly washed with plenty of distilled water. At this stage, the filtered precipitate was dried at 110[degrees]C to remove any trace of water. The synthesized sample was then calcined at 900[degrees]C maintaining a fixed calcination time of 30 minutes. After calcination the sample was crushed to achieve fine powder [13]. Pure HA was also prepared by following the same procedure maintaining identical condition for comparison and was sintered only at 900[degrees]C. Synthesized doped apatites were crushed to fine powder and then subjected to calcination at different four temperatures.

Chemical analysis

Various analytical procedures were adopted for characterization of synthesized apatites. Presence of Ca, P and Na was analyzed by Atomic absorption spectroscopy (AAS) and ultra violet spectrophotometric methods (UV). EDS and XRF analysis were applied for quantitative determination of their composition.

FT-IR analysis

The functional groups were determined by Fourier transform infrared spectroscopy (FT-IR, Model no. FTIR -8900, SHIMADZU). Experimental spectra were obtained by using KBr disks with a 1:100 "samples-to-KBr" ratio and the samples were scanned in the wavenumber range of 4000 [cm.sup.-1] - 400 [cm.sup.-1] with an average of 30 scans. The resolution of the spectrometer was 4 [cm.sup.-1].

X-ray powder diffraction analysis

Effects of sintering temperatures on phase properties were observed by X-ray diffraction analyses of both pure and doped apaties (calcined at various temperatures). This investigation was performed by using PANalytical (X'Pert PRO XRD PW 3040). The intensity data were collected in 0.02[degrees] steps following the scanning range of 20 = 20[degrees] - 80[degrees] using CuK[alpha] ([lambda] = 1.54178[degrees]A) radiation. The observed phases were compared and confirmed using standard JCPDS files.

Results and Discussion

Chemical analysis

From the comparative study in Table 1, it can be summarized that though we maintain the Ca:P ratio1:66 in the prepared solution, but addition of Na changes this ratio in synthesized doped apatites. This observation supports that Na-HA is non-stoichiometry and Na incorporates in the lattice structure.

FT-IR analysis

The FT-IR spectra of both pure and Na-HA (4 and 8 mol%) calcined at three different temperatures 110[degrees]C, 300[degrees]C, 600[degrees]C representing the characteristic peaks for the phosphate (PO43) and O[H.sup.-] groups. The bands for doped apatites calcined at 110[degrees]C - 600[degrees]C appeared as broad spectra, which ultimately supported the formation of apatite but in poor crystalline form (Fig.1). However, increase in calcination temperature causes a sharp change in peak intensities which is dictating from the spectra of Na-HA apatite sintered at 900[degrees]C (Fig. 2). Band positions at 3569[cm.sup.-1] due to stretching mode of O[H.sup.-] group for pure HA is of high intensity whereas upon calcination this mode disappeared for both Na4HA and Na8HA apatites. (Fig. 2 and 3) [1,7]. Besides, the characteristic peak position for adsorbed water at 3430.34 cm-1 was appeared for all apatites (Na-HA and pure HA) sintered even at 900[degrees]C but in broad fashion. In addition, for Na doped apatite the gap between the band positions of P[O.sub.4.sup.3-] group at 603.4 cm-1 and 962.5 [cm.sup.-1] is wide rather than pure HA (where, P[O.sub.4.sup.3-] group at 602.4 [cm.sup.-1], 629.1 [cm.sup.-1] and 962.7 [cm.sup.-1]) recommended the formation of lattice structure of low crystallinity. XRD observation also supports this assumption. For doped apatites, vibrational modes of C[O.sun.3.sup.2-] group appears at 1421 [cm.sup.-1] even upon calcination at 900[degrees]C. Presence of this peak could be assigned to the presence of Na [1], because FT-IR spectra of pure HA show complete absence of this group (Fig. 3). Moreover, it can be infer that, on calcination above 1000[degrees]C, peak of C[O.sub.3.sup.2-] group for Na-HA would be disappear due to volatilization. Some extra broad peaks of P[O.sub.4.sup.3-] group are observed at 1016-1093 [cm.sup.-1] for all calcined apatites which supports the appearance of a-TCP at 900[degrees]C [7,8]. FT-IR band positions and their corresponding assignments of pure HA, Na8HA and Na4HA at various temperatures are tabulated in Table 2 and Table 3.





XRD analysis

The phase properties of all apatites were determined from XRD patterns. Our effort was to study the change in phase composition of both doped and pure apatites with respect to various calcined temperatures concentration variation. Fig. 4 (a, b, c, d) has represented the XRD patterns for Na8HA. XRD spectra (Fig. 4a) for oven dried (at 110[degrees]C) Na8HA is reasonably broad that represents the poor crystalline nature. This observation is similar for apatite calcined upto 600[degrees]C (Fig. 4b and 4c) which is also relevant with FT-IR data. However, a sharp change occurs in XRD pattern for this apatite sintered at 900[degrees]C (Fig. 4d). Thus it can be infer that increase in sintering temperature increases the crystallinity degree results in several distinct and sharp peaks. This observation is further satisfied by plotting the peak height (at 20 positions ~ 31.8228) as a function of various calcination temperatures. It is obvious from the curve (Fig. 5) that peak height at 900[degrees]C is sufficiently high which ultimately support the well crystalline behavior of the doped sample at this specific temperature. However, the observed intensity and d-spacing values of dope apatites are in excellent agreement with the JCPDS standard data (ref. code: 09-0432) for HA. The change of Crystal size and crystallinity with sintering temperature presented in Table 4 also strengthen this statement.






Lattice parameter changes

Lattice parameter of pure HA is slightly higher than doped apatites (Na4HA and Na8HA).Thus Na incorporation affects lattice parameter to a certain extent. Cell parameter a decreases up to 8 mol% Na substitution for calcined apatites (Fig. 6). On the contrary, lattice parameter c is consistent, not significantly affected by Na content but slightly increase with temperature effects (Table 4). This finding contradicts with previous report [1] where both the lattice parameters increase with sodium content. This may due to different nature of raw material and preparation procedure. The calculated Cell volume from these parameters is highest for HA while gradual decrease is observed with increased Na content (Fig. 7).

Effects of Na substitution on crystallographic properties

Crystallographic properties changes with Na incorporation. However, as lattice parameter varies with Na content indicates that crystallographic properties would also alter. But such alteration does not follow the mode of cell parameters. Crystal size and crystallinity increase with increased Na mol% but fairly lower than pure HA (Table 5). Moreover, Crystal size follows a linear relationship with sintering temperature (Fig. 8). As well Crystalline apatite only occur upon calcination at 900[degrees]C, so crystallinity was measured only for apatites sintered at this specific temperature.

Phase determination with calcination

It is well established that HA prepared by means of precipitation route decompose on calcination near 800[degrees]C to 900[degrees]C [1,7,8]. In XRD spectra appearance of a 2nd phases with HA for both calcined pure and substituted apatites supports this notion.

Diffraction angles and d-spacing values of (0210) plane of this phase for pure and doped apatites were presented in Table 6 and compared with the JCPDS value (JCPDS#09-0169). Both this values for pure HA are very similar to JCPDS data for [beta]-TCP. On the other hand, for Na-HA these values are slightly shifted from JCPDS data indicates that for Na-HA the second phase would be a combined phase of both sodium and calcium phosphate rather than pure [beta]-TCP.

Besides, Table 7 summarized the crystal size and volume fraction of 2nd phase for all apatites. From tabulated data it is obvious that crystal size of the 2nd phase for pure HA is smaller than that of doped HA. Moreover, the volume fraction (%v) of [beta]-TCP for pure HA is lowers than the new phase of Na-HA. Thus it can be concluded that sodium substitution destabilize HA phase.

SEM analysis

As previously mentioned crystallography properties of apatite firmly depends on the sintering temperature. Well defined crystalline structure of both pure and doped apatites appear at sintering temperature of 900[degrees]C, so the crystal morphology and micro structural features of different mol% Na-HA sintered at this temperature were further examined by capturing their SEM micrographs. SEM images (Fig. 9a, 9b, 9c) of pure and doped apatites appeared as a combination of different regular but agglomerated shapes, such as hexagonal, spherical, etc. However, no significant morphological changes was observed for doping effects, but SEM micrograph for 0 mol% Na content is evident of comparably more distinct hexagonal phases. Such morphological analyses support our previous report on basis of FT-IR and XRD analysis.


Our aim was to characterize and investigate the morphological properties of successfully prepared Na doped apatite (4 and 8 mol% Na-HA) from waste egg shell (as Ca precursor) as a function of temperature and concentration effects. Cell parameter, cell volume, crystal size and crystallinity changes with Na incorporation which ultimately confirms Na substitution on HA lattice. Furthermore, all these values was inferior to those of pure HA, means addition of Na deteriorate the crystalline nature of the mentioned apatite. Our next attempt would be to study its biocompatibility for investigate its suitability as dental implant, bone filler for treatment of various bone diseases.


The authors gratefully acknowledge the financial support from IGCRT, BCSIR and the assistance of Dr. Mohammad Mizanur Rahman, Assistant Professor of ACCE, DU for FTIR. Thank is also due to the Ministry of Science and Information & Communication Technology, Government of Bangladesh for granting NSICT fellowship to SFK.


[1.] Kannan S., Ventura J.M.G., Lemos A.F., Barba A. and. Ferreira J.M.F, Effect of sodium addition on the preparation of hydroxyapatites and biphasic ceramics. Ceramics International 2008; 34:7-13.

[2.] Wang J., Nonami T., Yubata K., Syntheses, structures and photophysical properties of iron containing prepared by a modified pseudo-body solution. J. Mater Med 2008;19:2663-2667.

[3.] Kalita S. J., Bhatt. H. A. Nanocrystalline hydroxyapatite doped with magnesium and Zinc: Synthesis and Characterization, Materials Science and Engineering C 2007;27:837-848.

[4.] Li Y., Nam C.T. and Ooi C. P., iron(iii) and manganese(ii) substituted hydroxyapatite nanoparticles; Characterization and cytotoxicity analysis, Journal of Physics, 2009; Conference Series 187,012024

[5.] Webster T. J., Massa-Schlueter E. A., Smith J. L. and Slamovich E. B., Osteoblast response to hydroxyapatite doped with divalent and trivalent cations. Biomaterials 2004;25: 2111-2121

[6.] Ren F., Xin R., Ge X. and Leng Y., Characterization and structural analysis of zinc-substituted hydroxyapatites. Acta Biomaterials 2009;5:3141-3149

[7.] Cacciotti I., Bianco A., Lombardi M. and Montanaro L., Mg-substituted hydroxyapatite nanopowders: Synthesis, thermal stability and sintering behavior, Journal of the European Ceramic Society 2009;29:2969-2978

[8.] Miyaji F., Kono Y. and Suyama Y. Formation and structure of zinc-substituted calcium hydroxyapatite. Materials Research Bulletin 2004;40:2 09-220

[9.] Tang Y., Chappell H. F, Martin T. D., Richard J. R. and Young J. L. Zinc incorporation into hydroxyapatite. Biomaterials 2009;30:2864-2872

[10.] Doyle M. E., Glass K. A. Sodium Reduction and Its Effect on Food Safety, Food Quality, and Human Health. Food Science and Technology 2009;9;44-56.

[11.] Hockin H. K. X.,Takagi S., Sun L., Hussain L., Chow L. C., William F., Yen J. H. Development of a nonrigid, durable calcium phosphate cement for use in periodontal bone repair, JADA 2006;37:1131-1138.

[12.] Choi J.W., Cho H. M., Kwak E. K., Kwon T. G., Ryoo H. M., Jeong Y. K., Oh K.S. and Shin H. I. Effect of Ag-Doped Hydroxyapaptite as a bone fille for inflamed Bone Defects. Key Engineering materials 2004;vols254-256:47-50.

[13.] Kabir S. F., Ahmed S., Ahsan M., Mustafa A. I. Dope hydroxyapatite from waste calcium source: Part 1-Na doped apatite. Material Science 2011;7(1);42-48.

[14.] Ahmed S. and Ahsan M. Synthesis of Ca-hydroxyapatite Bioceramic from Egg Shell and its Characterization. Bangladesh J. Sci. Ind. Res. 2008;43(4):501-512

Sumaya F. Kabir (a), Samina Ahmed * (b), Mainul Ahsanb, Ahmad I. Mustafa * (a)

(a) Department of Applied Chemistry and Chemical Engineering, University of Dhaka, Dhaka-1000, Bangladesh, bInstitute of Glass and Ceramic Research and Testing (IGCRT), Bangladesh Council of Scientific and Industrial Research (BCSIR), Dhaka-1205, Bangladesh

* Corresponding author:

Received 27 December 2011; Accepted 20 February 2012; Available online 6 May 2012
Table 1: Molar percentage concentration of precursors in HA

Precursors            Prepared solution        Synthesized samples

                    HA     Na8HA    Na4HA      HA     Na8HA    Na4HA

Ca(mol%)           62.43   62.43    62.43    62.43    57.98    59.98
P (mol%)           37.56   37.56    37.56    37.56    36.69    36.80
Na(mol%)            --      5.23     2.68      --       5.23     2.68
Ca/P               1.66     1.66     1.66     1.66     1.58     1.63
Na/(Na+Ca)(mol%)    --       --       --       --      8.27     4.27

Table 2: FT-IR band positions and their corresponding
assignments of Na8HA and Pure HA

               Observed band positions ([cm.sup.-1])

              Na8HA           Na8HA           Na8HA
Pure HA   110[degrees]C   300[degrees]C   600[degrees]C

562.3         561.2           560.3           561.8

602.4         602.3           602.9           603.5

629.1          --              --              --

962.7         961.9           960.1           963.4

--           1418.1          1418.6          1420.5

3430.5       3430.3          3432.6          3431.2

3572.6       3567.4          3568.2          3568.5

              Na8HA       Corresponding assignments
Pure HA   900[degrees]C

562.3         562.4       P[O.sub.4.sup.3-] bending

602.4         603.4       P[O.sub.4.sup.3-] bending

629.1          --         P[O.sub.4.sup.3-]
                          asymmetric stretching (v1)

962.7         962.5       P[O.sub.4.sup.3-]
                          symmetric stretching (v3)

--           1421.7       C[O.sub.3.sup.2-] group

3430.5       3431.7       [H.sub.2]O adsorbed (v2)

3572.6          ?         Structural O[H.sup.-]

Table 3: FT-IR band positions and their corresponding
assignments of Na4HA

             Observed band positions ([cm.sup.-1])

Na4HA 110    Na4HA 300    Na4HA 600    Na4HA 900    Corresponding
[degrees]C   [degrees]C   [degrees]C   [degrees]C   assignments

560.2          561.2        561.9        561.5      P[O.sub.4.sup.3-]
                                                    bending (V4)

601.6          602.4        602.3        602.8      P[O.sub.4.sup.3-]
                                                    bending (v4)

--               --           --           --       P[O.sub.4.sup.3-]
                                                    stretching (v1)

960.3          961.3        963.4        962.3      P[O.sub.4.sup.3-]
                                                    stretching (v3)

1417.3         1418.1       1418.4       1419.3     C[O.sub.3.sup.2-]
                                                    group (V3)

3231.2         3231.2       3231.3       3231.5     [H.sub.2]O
                                                    adsorbed (v2)

3567.2         3567.8       3568.2         --       Structural

Table 4: Cell parameters as of all apatites

Sample apatites

                                Lattice parameter         Cell volume
                        a (A[degrees])   c (A[degrees])   (A[degrees])

Pure HA 900[degrees]C        9.42             6.88          1580.60
Na8HA110[degrees]C           9.31             6.86          1539.41
Na8HA 300[degrees]C          9.36             6.86          1562.65
Na8HA 600[degrees]C          9.38             6.86          1565.98
Na8HA 900[degrees]C          9.39             6.87          1568.60
Na4HA 110[degrees]C          9.32             6.86          1542.72
Na4HA 300[degrees]C          9.38             6.86          1565.89
Na4HA 600[degrees]C          9.39             6.87          1568.60
Na4HA 900[degrees]C          9.40             6.87          1571.60

Table 5: Crystallographic information of apatites

Sample apatites          Crystal size   Crystallinity

Pure HA 900[degrees]C       690.90           5.01
Na8HA 110[degrees]C         340.23            --
Na8HA 300[degrees]C         515.23            --
Na8HA 600[degrees]C         521.12           0.39
Na8HA 900[degrees]C         659.63           3.32
Na4HA 110[degrees]C         321.58            --
Na4HA 300[degrees]C         412.54            --
Na4HA 600[degrees]C         415.43           0.62
Na4HA 900[degrees]C         451.20           2.34

Table 6: Diffraction angles and d-spacing values for
2nd phase appeared on calcined at 900[degrees]C

                                     (0210 plane)

                              d-spacing     2([degrees]
Sample apatites Pure HA      (A[degrees])     Theta)

                                2.8794        31.0180
Na8HA                           2.8528        31.2421
Na4HA                           2.8489        31.2327
JCPDS#09-0169 ([beta]-TCP)      2.8800        31.0260

Table 7: Crystal size and volume fraction
of 2nd phase at 900[degrees]C

                   Crystal        Volume
Sample apatites   size (nm)    fraction (%v)

Pure HA              46.5          18.1
Na8HA                56.7          28.6
Na4HA                54.2          26.4
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