Investigation of HA Nano-Crystallite Size Crystallographic Characterizations in NHA, BHA and HA Pure Powders and
Their influence on Biodegradation of HA
Ebrahim Karamian
1,a, Amirsalar Khandan
3,b, Mustafa Eslami
2,c, Hassan Gheisari
4,dand Niosha Rafiaei
5,e1,2,5 Assistant professor and M. Sc. Students, Department of Materials Engineering,
Najafabad Branch, Islamic Azad University, P.O. Box 517, Isfahan, Iran
3 M. Sc. Student, Department of Mechanical Engineering, Khomeinishahr. Branch, Islamic Azad University, Isfahan, Iran
4 M. Sc. Student, Department of Materials Engineering, Shahreza Branch, Islamic Azad University, Isfahan, Iran
a [email protected], b,*[email protected],c [email protected],
d [email protected], e [email protected]
Keywords: NHA; lattice parameters; c/a ratio; HA; XRD; Nano-crystal; BHA; Biodegradation; SBF Abstract. The mineral or inorganic component of bone is a calcium phosphate idealized as a calcium hydroxyapatite, HA, Ca10(PO4)6(OH) 2. Changes in the composition of the apatite affect its lattice parameters, morphology, crystallinity (reflecting crystal size and/or perfection) and finally dissolution properties. In regard to the above points, lattice parameters are expected to have better dissolution properties. Estimation of HA nano crystallite size lattice parameters is one of the most important factors for dissolution properties of bone apatites. In present work, the composition of apatite induce complex structures at the unit-cell level and play a role in influencing the dissolution rate of apatites, which may favour osteointegration. The samples are consist of NHA, bovine bone heated at 850 C for 3 h; BHA, human bone heated at 900 ºC for 2 h and PHA, HA pure powder.
The results estimated by XRD data indicate that increasing in c/a ratio of HA, which is leading to increasing in crystallinity, induce decrease at HA dissolution or improving its chemical solubility in simulated body fluid (SBF). As, it was concluded that the biodegradation of HA decrease in PHA sample.
Introduction
Synthetic hydroxyapatite (HA) is the most helpful because of its similarity to natural bone in both crystalline structure and chemical composition. HA is a material that has significant research synthesis and characterisation of pure nano porous hydroxyapatite applications, especially in the biomedical field. HA, with the chemical formula Ca10 (PO4)6(OH)2, is the main component of bone and teeth. This bioceramic has been widely used in dental materials, constituent implants and bone substituent materials due to its excellent biocompatibility, bioactivity, osteoconductivity, non- toxicity and non-inflammatory nature. HA is manufactured in many forms and can be prepared as a dense ceramic, powder, ceramic coating or porous ceramic as required for the particular applications [1]. Tissue diseases and defects, particularly bone diseases are serious health condition that directly affects the quality of life of the sufferers [2]. Ceramics used for this purpose are termed bioceramics. Replacement of tissues has two alternatives: (1) transplantation and (2) implantation.
The significant advantages of bioceramics as implants over transplants are availability, reproducibility and reliability. Moreover, they do not pose the risk of any viral or bacterial infection to patients [3,4]. Apatites have good bioactivity and osteoconducivity, and are used as bulk ceramics, coatings and granules in clinical applications. Thus, HA has been widely used in dental implants, alveolar bridge augmentation, orthopedics, maxillofacial surgery, etc [5]. New knowledge on apatite lattice constants (LC) may shed light on not only the crystal structure behavior but also
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on understanding their physico-chemical properties to facilitate the design and fabrication of specific biomaterials. Basic apatite structure is hexagonal with space group P63/m, although a few monoclinic forms with space group P1121/b were also reported [6]. A research program is carried out by the author’s group to develop computing models to aid in the understanding and design of new biomaterials. The emphasis was placed on the relationship of the structure (or LCs) and atomic properties of these apatites. Advances in high performance computing techniques allow calculating LC based on first principles of quantum mechanism [7]. Hydroxyapatite has a hexagonal symmetry and unit cell lattice parameters a = 0.95 nm and c = 0.68 nm (fig.1). Taking into account the lattice parameters and its symmetry, its unit cell is considered to be arranged along the c-axis. Unlike the other calcium phosphates, hydroxyapatite does not break down under physiological conditions.
Lattice parameter measurements are used in many situations to characterize biomaterials. For example, knowledge of the lattice parameters can provide information on the thermal properties of a material, an indirect method to determine the compositions in a solid solution, a measure of the strain state, or an analysis of the defect structure. Therefore, it is important to determine the lattice structure with the highest precision. Fortunately, X-ray diffraction can provide such information to an accuracy of several significant figures if care is taken during the experiment and subsequent analysis.
Fig.1. Structure, Unit Cell, of HA [8]
Materials and Methods
Bovine bones were boiled for 2 h to remove flesh and fat. The bones were heated at 110 C for 2 h to remove moisture. To prevent blackening with soot during heating, the bones are cut into small pieces of about 10 mm thick and heated at 500 C (bone ash) for 2 h in air to allow for evaporation of organic substances. The resulting black bone ash was heated for 3 h at 850 C. This synthesis is called thermal decomposition of bone resource to create natural HA, NHA. According to the above same way, a Scapular bone has been heated at 900 C for 2 h to create biological HA, BHA. And synthesized nano sized hydroxyapatite, PHA, was created by sol-gel method. Phase structure analyses were carried out by X-ray diffraction (XRD) analysis using a Philips X’Pert-MPD diffractometer with Cu Ka radiation (λ1 = 0.15418 nm) over the 2θ range of 20–80. The obtained experimental patterns were compared to the standards compiled by the Joint Committee on Powder Diffraction and Standards (JCDPS) which involved card # 09-432 for HA. The crystallite size of prepared powders was determined using XRD patterns and modified Scherrer equation. And c/a ratio has been calculated by XRD data.The SBF was prepared according to the procedure described by Kokubo [9]. The ion concentrations of the SBF are similar to those in human blood plasma. The powders were weighted in amount of 0.3 g and pressed in disc steel mould with 6 mm in diameter for 2 min under 9000 N force. Disc samples were soaked in the cell SBF solution (pH 7.4) at 37 C and physiological saline (pH 5.2) for 1, 6, 11 and 14 days at a solid/liquid ratio of 1 mg/ml without refreshing the soaking medium. After soaking for the predicted time periods, the disc samples were rinsed with deionized water, and dried in an oven at 110C for 1 h.
The lattice parameters, a and c, are calculated by using the following equation eq.1:
1/d2=
¾
(h2+hk+k2)/a2 +l2/c2 (1) Where d is distance between the planes with Miller indices (h, k and l).
Also, crystallinity (Xc) is calculated by using XRD data and following equations.
Xc = 1- (V112/300/I300) (2)
where I300 is the intensity of (300) diffraction peak and V112/300 is the intensity of the hollow between (112) and (300) diffraction peaks of the produced powders.
Results and Discussions
XRD Results. Figures 1 and 2 show the XRD pattern of pure HA, NHA and BHA samples, respectively. They illustrate that there is just HA phase, Hydroxyapatite, in all the samples. (All the peaks belong to HA)
(a) (b)
Ref. Code Score Compound Name Scale Factor Chemical Formula 01-074-0565 81 Hydroxylapatite 1.007 Ca10 (PO4)6 (OH)2
(a)
Ref. Code Score Compound Name Scale Factor Chemical Formula 00-024-0033 63 Hydroxylapatie 0.967 Ca5 (P O4)3 (OH)
(b)
Fig 1. a) XRD pattern of sample pure HA, PHA, and b) XRD pattern of sample BHA
(c)
Visible Ref. Code Score Compound Name
Displacement [°2Th.]
Scale Factor Chemical Formula
* 01-024-0033 69 Hydroxylapatite 0.000 0.994 Ca10 (P O4 )6 (OH)2
Fig 2. XRD pattern of NHA sample
Position [°2Theta]
20 30 40 50 60 70 80
Counts
0 1000 2000
4.sd
Position [°2Theta]
20 30 40 50 60 70 80
Counts
0 100 400
HHA9002.sd
The amounts of HA biodegradation properties and weight loss of the samples is shown in table 1.
Values crystallographic parameters of HA phase at all the samples is shown in table 2.
The gradual increase of crystallinity suggests increasing in lattice parameters (a) which is leading to decreasing weight loss. In fact, HA dissolution was decreased by increasing the amount of crystallinity.
Table 1. The HA biodegradation properties of all the samples in biological solution
Table 2. Values crystallographic parameters of HA phase at all the samples Sample Lattice Par.
a (Aº)
Crystalinity, Xc (%)
Crystallite Size ( nm)
PHA 9.417 97.1 53.4
BHA 9.425 90.2 44.3
NHA 9.443 84.6 25.5
Conclusions
1) The results estimated by using the XRD data indicate that decreasing in lattice parameters induce increasing in crystallinity and decreasing lattice parameter which are improving of HA dissolution in the biological solutions.
2) So, we reached to the conclusion that the HA biodegradation improves in the PHA samples. In fact, there were at least weight losses and Ca release in these samples.
3) In general, values of weight loss of samples and Ca release in the solutions were occurred in physiological saline more than SBF in the all samples. It seems that presence of high level of chloride ion (Cl-) in the physiological saline, 153 ppm, rather than the SBF, 147 ppm, induce increasing HA dissolution.
4) In fact, it can conclude that presence of chloride ion and Na+ result in degradation of HA crystals in the samples.
Acknowledgement
The authors would like to extend their gratitude for the supporting provided by Isfahan Azad University of Khominishahr & Najafabad & Shahreza, Iran.
References
[1] Rozita Ahmad Ramli1, Rohana Adnan1, Mohamad Abu Bakar1 and Sam’an Malik Masudi,Synthesis and Characterisation of Pure Nanoporous Hydroxyapatite, Journal of Physical Science, Vol. 22(1), 25–37, 2011.
[2] Hench LL. Bioceramics: From concept to clinic. J Am Ceram Soc 1991, 74: 1487-510.
[3] Hench LL, Wilson J. Introduction to bioceramics. Singapore: World Scientific, 1993.
[4] Ranter BD, Hoffman AS, Schoen FJ, Lemons JE. Biomaterials science. An introduction tomaterials in medicine. San Diego: Academic Press, 1996; p. 484.
[5] Amjad Z. Calcium phosphates in biological and industrial systems. Boston: Kluwer Academic Publishers, 1998; pp. 67-83.
Sample % W.L of samples in SBF After 1
Weeks
% W.L of samples in Saline After 2
Weeks
Ca Release in SBF After 1 Weeks [Ca+2]
ppm
Ca Release in Saline After 2 Weeks [Ca+2]
ppm
PHA 0.39 1.06 48 61
BHA 0.62 1.67 55 68
NHA 3.31 4.02 62 115
[6] Elliott JC. Structure and chemistry of the apatites and other calcium orthophosphates.
Amsterdam: Elsevier; 1994.
[7] Langford JI, Louer D. Powder diffraction. Rep Prog Phys 1996;59:131–234.
[8] Figure 1. The reference used is from Optimisation of Plasma Sprayed Hydroxyapatite Coatings Tanya. J. Levingstone, BEng.
[9] T. Kokubo, H. Takadama, How useful is SBF in predicting in vivo bone bioactivity, Biomaterials 27 (2006) 2907–2915.
Investigation of HA Nanocrystallite Size Crystallographic Characterizations in NHA, BHA and HA Pure Powders and their Influence on Biodegradation of HA
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