Effects of magnesium doping on the properties of hydroxyapatite/sodium alginate biocomposite

Abstract

A novel hydroxyapatite/sodium alginate biocomposite doped with magnesium was developed to enhance the physical, mechanical and bioactivity properties in bone implant applications. Specifically, magnesium was doped in hydroxyapatite (HA) (Ca₁₀(PO₄)₆(OH)₂/sodium alginate (SA) (NaC₆H₇O) by using precipitation method. This research also explored the effects of magnesium doping on HA/SA samples. The prepared powder was uniaxially pressed and sintered at 1300°C. The characterisation of Mg-doped HA/SA at various concentration ranging from 0.5 wt-% to 1.5 wt-% were performed through Field Emission Scanning Electron Microscopy (FESEM) analysis. The maximum relative density and hardness of Mg-doped HA/SA were fixed at 92% and 4.11 GPa respectively and at 1.0 wt-% for samples of magnesium doping. Based on the microstructure analysis by FESEM it is evident that the elements were distributed evenly in Mg-doped hydroxyapatite/sodium alginate (HA/SA). These results proved that the Mg doping increased the physical, mechanical and bioactivity properties of HA/SA biocomposite.

Keywords

Bioceramics composite sintering Vickers hardness hydroxyapatite

Introduction

Hydroxyapatite (HA) (Ca₁₀(PO₄)₆(OH)₂) has been widely used as a bone replacement in orthopaedics implants due to its structure that are similar composition to the human bone. Besides that, extensive studies have been indicated that HA is biocompatible to hard tissues [1]. HA material has been attracted due to the promising bioactive materials for use in biomedical applications. Although, HA is a preferred material, it has several limitation, which is poor mechanical properties especially in load-bearing area due to it brittle characteristic [2 -5]. In addressing these problems, this research has been done by focusing on the improvement of densification, morphology and mechanical properties. The improvement in mechanical properties has been taken place by forming of composite. The combination of HA and SA done to form a composite samples [6].

In recent years, natural polymers such as sodium alginate, chitosan, gelatin, collagen and cellulose have gained attention in biomedical application as these materials are biodegradable, biocompatible and non-toxic. Among natural polymers, SA is a natural polysaccharide derived from sea algae and is useful to treat the loss bone and defect bone. In addition, the pure SA have some deficiencies such as lack of interaction and poor mechanical strength. The main objective of this study is to evaluate the effects of magnesium on the density and mechanical properties of HA/SA composites. The combination of composites material containing SA and inorganic material has gained interest due to possess good bonding, bone regeneration and oesteoconductivity [7 -14].

Bone is composed of 30% organic and 70% inorganic phase which are responsible for mechanical resistance. In the HA structure, the ions can be replaced with the carbonate groups while the calcium ions can be substituted by divalent ions such as Mg²⁺ [4,11,12]. The functions of bone is to provide the mechanical strength and structural framework.

Sintering process is also important as it displayed the grain growth. Furthermore, the increment in Mg doping results in grain growth and porosity reduction. This finding are reported similar to Ramesh et al. [11]. The sintering process is applied in HA/SA to densifying and to produce the complete ceramic composite matrix materials. The sintering process convert this product into sodium carbonate that is important in regeneration of bone implants. However, these material are not enough to be used in load-bearing area. As a solution, a metal- doped biocomposite material is used in this research where Mg was doped into HA/SA biocomposite to enhance the physical, mechanical properties and analyse the bioactivity of HA/SA and to make it widely used in medical fields. In addition, the bone mineral also contains trace ions like Na⁺, Mg²⁺ and which are known to play an important role in implants process. The sintering process could produce the magnesium, sodium and carbonate that is the important trace element found in human bone.

Magnesium ions are one of the essential ions that presence in human body and the deficiency of Mg content can led to defects in bone growth resulting a decrease of osteoblast and osteoclastic activities. Mg ions have well known as one of the substitutes or dopants for calcium in HA structure. The doping with low concentration of Mg can influence the morphology and also improve the osteoblast adhesion if compared to HA/SA. However, the higher doping of Mg content lead to defect in properties of HA [3,4,12]. Suchanek et al. [15] reported that, the Mg doping has better biocompatibility and bioactivity properties. This is widely used in orthopaedic and denstal applications.

The main objective of this study is to develop the effects of Mg doping into HA/SA on densification, microstructure and mechanical properties. In order to improve the physical and mechanical properties of HA/SA, various content of Mg ion were employed in this study. The effects of Mg content on HA/SA properties are further investigated. Undeniably, the results of this study are crucial because this composite material could improve the mechanical stability, increase tissue regeneration in cells, particularly in load-bearing areas, and enhance the bioactivity of Mg-doped HA/SA. As, there is no studies conducted on Mg-doped HA/SA, the present study focused on sintering process that enhanced the physical, mechanical and biocompatibility of sintered sample and intended to determine the effect of Simulated Body Fluid (SBF) on the Mg-doped HA/SA properties.

Experimental method

Sample preparation

Mg-HA/SA was prepared by using the precipitation method. With respect to this method, the commercial powder with various weight percentages of magnesium hydroxide Mg(OH)₂ (Q-Rec) powder at 0.5 wt-%, 1.0 wt-% and 1.5 wt-% were doped with hydroxyapatite (HA) Ca₁₀(PO₄)₆(OH)₂ (Emory,99.5%)/sodium alginate (SA) NaC₆H₇O (Sigma Aldrich). The ratio of HA is 99.5 while SA is at about 0.5. Then, the HA/SA and Mg-doped HA/SA powder were compacted by the uniaxial pressing at 19.9 MPa for five minutes and then sintered at 1300°C, at 2°C/min for two hours. Next, the sintered samples were coated with a platinum layer to induce conductivity for FESEM analysis. Subsequently, the morphology and elementary analysis of sintered samples were evaluated by using Field Emission Electron Microscopy (FESEM), (JSM model JEOL, Japan) at accelerating voltage of 10 kV to 15 kV and energy dispersive X-ray analysis (EDS). Also, the density of sintered samples was determined by Archimedes Principle using distilled water. Additionally, the samples were polished with polishing cloth using diamond paste and thermal etched at 1250°C for 30 min at 2°C min^–1 to reveal the grain boundary. The grain sizes were revealed by using Intercept method. Straight lines were drawn at the diagonal of the microstructure and the intercept lines between grain boundaries were measured. The measurement must be taken at two lines that were drawn at diagonal and the average values were recorded. Finally, Vickers Microhardness was applied to measure the hardness of the sintered samples. 0.2 HV load with dwell time of 10 s are employed in this polished samples to produce the indentation. Five indenter tests were applied on the surface of sample, and the average value was recorded. Thus, 5 samples were tested for each samples.2 mg of each specimen samples was immersed in 40 mL simulated body fluid (SBF). The SBF solution had similar composition and concentration that are similar to human blood plasma and was prepared by dissolving NaCl, KCl, K₂HPO₄, 3H₂O, MgCl2.6H₂O, CaCl₂, and Na₂SO₄ reagents in deionised water according the kokubo procedure, and pH of solution adjusted at 7.4 by hydrochloric acid, HCl. The samples were kept in SBF for 36.5°C for 1, 7 and 14 days. The apatite formation was observed by FESEM analysis.

Results and discussion

Densification

Figure 1, shows that the Mg increased the relative density of HA/SA until 1.0 wt-% at about 92% and experienced a slight reduction at 1.5 wt-% around 89% of Mg content. Based on these results, it could be deduced that the increment of relative density was governed by the presence of Mg ions. Clearly, the composition of 1.0 wt-% of Mg doping was sufficient to enhance the densification and hardness. Interestingly, at 1.0 wt-% of Mg doping, the grain growth became large while the porosity underwent reduction. Conversely, the sintered samples of HA/SA exhibited low density due to the presence of large pores as their behaviours could be observed in FESEM microstructure. Overall, the presence of Mg-doping significantly influenced the density and porosity of the sample. In connection with this result, a past research by Muralithran and Ramesh also highlighted that a higher sintering temperature could reduce the porosity level [16]. For instance, in this particular study, 1.0 wt-% of Mg-HA/SA samples produced better densification than the other composition ratios. This is most probably due to the fine particle size which influences the increase of particles size when corresponded with a higher sintering temperatures as outlined by Ramesh et al. [11]. Based on a previous research conducted by Ramesh and Ou et al. [11,17] it was recorded that the densification at 1.5 wt-% of Mg showed a decrease in density. With respect to this study, this outcome could most probably be attributed to the process of elimination of pore along the grain boundaries that exist in the green compact samples. As a matter of fact, the decrease in density was attributed to the HA/SA limit. Furthermore, the initial sintering accelerated the growth of grain boundaries to ensure the isolated pores are kept confined within the grain structure. On the other hand, the sintering temperature at 1300°C displayed grain growth within the range of 3.7 µm. Inevitably, the increment in Mg content resulted in grain growth and pore reduction. Thus, this outcome supports the finding of Ramesh et al. [11] which claimed that density corresponds positively with the Mg content and sintering temperature, which stimulates grain growth. There is the larger grain growth that led to decrease in porosity where Figure 1(b) shows the HA/SA has higher porosity. However, due to Mg doping the porosity decrease from 0.5 wt-% and 1.0 wt-% but increase at 1.5 wt-% of Mg. This results are relative to microstructure that presented in Figure 3 demonstrate the well consistent of Mg-doped HA/SA at sintering temperature 1300°C.

Vickers micro hardness

Figure 2 shows the hardness of various concentration of Mg doped HA/SA compared with HA/SA sintered samples. The figure indicates that the increment of Mg concentration increased the hardness of samples as compared with pure HA/SA. Tan et al. [18] reported that, the increase in hardness are due to doping of Mg ions. At 1.5 wt-% of Mg doping, there was a significant decrease in both density and hardness due to the increase in porosity. In summary, the increase in Mg content at certain limit could affect both density and hardness. Additionally, the average grain size of Mg-doped HA/SA was within the range of 1–6.5 µm at 1300°C, whereas for 0.5 wt-% and 1.5 wt-% of Mg doping, the average grain size dropped to 0.49–6.27 µm and 0.57–3.98 µm respectively. From these results, it could be concluded that there is a correlation between hardness and grain size in which critical grain size may undergo diminution with respect to the hardness of Mg-doped HA/SA. Mg-doped HA/SA has high density and mechanical properties compared to HA/SA. The hardness of Mg-doped HA/SA increased with the grain growth. The hardness results are relative to the densification process where the increase in density lead to decrease in porosity and resulting the grain growth. Based on the observation, the grain growth increase the hardness of Mg-doped HA/SA at certain composition. It can be summaries that the hardness of Mg-doped HA/SA are very much controlled by density. The increase in both properties leads to decrease in porosity. Niakan et al. [19] reported that the amount of porosity can be suit in the biomedical application that require higher density and hardness.

Microstructural evaluation

Figure 3(a,b) shows the FESEM microstructure of HA/SA biocomposites with different amounts of Mg doping. Therefore, it was speculated that the Mg-doped HA/SA composites consisting small and large grains that could increase the grain growth. Essentially, the grain growth was discovered to enlarge as a result of the increase in amount of Mg doping in composite samples from 0.5 wt-% to 1.5 wt-% as compared with pure HA/SA. Also, further augmentation in the weight percentages of magnesium led to the decrement of the grain size of Mg-doped HA/SA. For instance, one of the notable grain growths was recorded from the size of 1–6.5 µm at 1300°C. Besides, the grain size of 1.0 wt-% of Mg doping showed the grain size was much smaller when compared to 0.5 wt-% and 1.5 wt-%. For instance, the grain size for 1.0 wt-% of Mg doping increased from 0.56–2.68 µm when tested at sintering temperatures of 1000–1300°C. On the other hand, the grain sizes for 0.5 wt-% and 1.5 wt-% of Mg doping exhibited an increase from 0.49–6.27 µm and 0.57–3.98 µm respectively at similar temperatures. Evidently, these results prove that Mg has been successfully incorporated into the material. Among the various composition ratios, 1.0 wt-% was identified to be most uniform and practical for bone implant. Note that, HA/SA samples demonstrate high porosity. The distribution of HA/SA is uniform as compared with 0.5 wt-% to 1.0 wt-% of Mg content. The findings indicates that the incorporation of Mg content into HA/SA significantly reduces the porosity of sintered samples. The decrease in the porosity value of HA/SA composites samples are due to the Mg doping content.

Bioactivity of SBF

Figure 4 shows the FESEM micrograph of HA, HA/SA and Mg-doped HA/SA composites after two weeks soaking in SBF. It was found that the ability of apatite formation of Mg-doped HA/SA is increase. The bioactivity of Mg-doped HA/SA on bone implant expected more bioactive due to the existence of Mg. The doping of Mg into HA/SA are mimic to natural bone composition. The Mg doping at 1.0 wt-% are expected to have more excellent bioactive and biocompatibility compared to HA/SA and HA. This material is more suitable to be used compared than HA as a bone substitute in implantation process [20 -22]. It was suggested by Nabiyouni et al. [23] that Mg ions promote the compatibility and proliferation rate of implant by in-vitro test.

Conclusion

In this research, magnesium ions were successfully incorporated in the HA/SA samples. Primarily, the effects of Mg doping studied were mainly focused on physical, mechanical and bioactivity properties with respect to the microstructure analyses. Notably, the maximum density of sintered samples was achieved at 1.0 wt-% of Mg. The increase in Mg content leads to enhance the mechanical properties of HA/SA samples. The morphology, physical, mechanical properties and bioactivity of HA/SA biocomposite are promising for the applications in tissues engineering. In summary, the maximum relative density and hardness were achieved at 92% and 4.11 GPa respectively at 1.0 wt-% of magnesium doping. The aim of this study was to examine the influence of doping process on the morphology of HA/SA and the influence of Mg ions on bone regeneration process. Furthermore, the doping of Mg ions in HA/SA could improve the biocompatibility and mechanical properties.

Footnotes

Disclosure statement

No potential conflict of interest was reported by the authors.

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