*3.1. Microstructures*

Figure 1 shows SEM images of starting powders of Ti6Al4V and EB, and ball-milled powders of Ti6Al4V-EB composite powders as a function of the EB weight fraction (wt.%); 0.05, 0.5, and 5 wt.%. The composites containing EB of 0.05, 0.5, and 5 wt.% were expressed as T-0.05EB, T-0.5EB, and T-5EB, respectively. According to the images, the shape of the ball-milled powders was insignificantly changed, but a rough surface iwa shown due to the ball-milling process. During the ball-milling process, physical changes were induced by the impact between the balls and the powders accompanying shear force. Therefore, the size of the powders was slightly increased from 32 to 45 μm, because certain cold welding could occur between the balls [30]. In the process, nanoscale EB powders were attached to the Ti6Al4V powder surface and then gradually embedded in the Ti6Al4V matrix. Accordingly, EB powders were not observed on the Ti6Al4V powder surface in all composite powders as shown in Figure 1d,f,h. From the elemental analysis using EDS, the key chemical composition was maintained as shown in Figure 1i. The main chemical composition of the hydroxyapatite, such as Ca, P, and O, increased as the EB contents increased.

Figure 2 presents the SEM images and EDS mapping of SPSed Ti6Al4V and the Ti6Al4V-EB composites with the weight fractions of EB 0.05, 0.5, and 5 wt.%. As shown in the images, α-Ti with the hexagonal closed packed (HCP) and β-Ti with the body-centered cubic (BCC) phases are clearly shown as dark grey and bright grey, respectively, and a volume fraction of β-Ti phase (*f*β) is displayed in each of the SEM images. The chemical compositions for each phase from EDS analysis are shown in Table 1. As a reference, defects such as pores and voids were not significantly found in the Ti6Al4V sintered specimen, and the presence and amount of β-Ti were 12.6%. The sintered composites having a porosity below 1% and relative density above 99% on average, excluding Ti6Al4V-5EB composites. Ti6Al4V-5EB composites showed a relative density of 95%, which affects elastic modulus and the hardness results.


**Table 1.** EDS analysis of the samples with the phase fraction of α-Ti and β-Ti phase from the SEM images of Figure 2.

**Figure 1.** *Cont*.

**Figure 1.** SEM images of starting materials and composite powders as a function of their equine bone (EB) contents; (**a**) Ti6Al4V; (**b**) EB; (**<sup>c</sup>**,**d**) T-0.05EB; (**<sup>e</sup>**,**f**) T-0.5EB; and (**g**,**h**) T-5EB. (**i**) Average chemical composition of Ca, P, and O in (**d**), (**f**), and (**h**).

**Figure 2.** SEM images and EDS analysis of ball-milled Ti6Al4V-EB composites under different weight fraction of the EB; (**a**) T-0.05Ed; (**b**) T-0.5EB and (**c**) T-5EB.

For all cases of Ti6Al4V-EB composites, well-distributed EB and α + β morphology were observed. Regarding the distribution of Al and V, the corresponding concentration ratios for α-Ti and β-Ti were found to be approximately 3:2 and 3:4, respectively (in Figure 3e) [31]. The V concentration in the Ti matrix increased according to the β-Ti phase transformation. The significance in *f*β is closely related to elastic modulus, which is an important factor as a biomedical material to attain low elastic modulus, because beta phase has a lower elastic modulus than alpha phase [32–34]. The stress shielding effect can occur according to high elastic modulus; therefore, a decrease in elastic modulus is important for biomedical implants [34–37]. The stress shielding effect is defined as the stress transfer between the bone and implant materials which is not homogeneously caused by a difference in elastic modulus. For instance, when the load is applied, the implant with a high elastic modulus mostly absorbs all of the tensile and compressive stresses and bending moments, which were previously applied to the bone; resultantly, osteoporosis or bone loss can occur [36,38,39].

ǂǂǂǂǂǂǂǂ

**Figure 3.** SEM images and EDS analysis of spark plasma sintered (SPSed) Ti6Al4V and Ti6Al4V-EB composites under different weight fraction of the EB; (**a**) Ti6Al4V; (**b**) T-0.05EB; (**c**) T-0.5EB; and (**d**) T-5EB (*f* is a fraction of β-Ti phase).

Figure 4 shows the XRD analysis of the starting materials and Ti6Al4V-EB composite before and after SPS to identify the elements and the phase distribution. The major peaks are attributed to α-Ti (marked by black square), β-Ti (marked by green square), and hydroxyapatite (marked by a red square). All peaks are in good agreemen<sup>t</sup> with each standard spectrum from the JCPDS database; α-Ti peaks displayed the (100), (002), (101), (102), and (110) reflection peak around 2θ = 35.4, 38.6, 40.5, 53.4, and 63.7◦, respectively (JCPDS Card No. 00-044-1294 and 00-009-0098). Although the peaks of hydroxyapatite for the composite powder and SPSed composites were shrouded in major peaks of Ti due to a small amount of EB, peaks from 2θ = 32◦ to 2θ = 35.2◦ for hydroxyapatite (JCPDS Card No. 00-009-0432) were detected, as shown in Figure 3. According to the literature, for sintering up to 1200 ◦C, hydroxyapatite can prevent decomposition into metastable phases such as tri-calcium phosphate during the sintering [40]. After SPS at 1000 ◦C, the main peaks were clearly detected for Ti6Al4V-EB composites. Several β-phases within the Ti6Al4V/EB composites, the (110) reflection peak of which is around 2θ = 39.7◦, are displayed in Figure 4b. The XRD peak of the β-Ti phase observed in the T-5EB sample subjected to SPS is shifted to a lower angular position, because more V contents in the matrix, with a lower atomic radius of 0.132 nm, were dissolved in the β-Ti phase. After SPS, while 2θ was increased from 40.48 to 40.51◦, the peak shifted towards a smaller angle θ of 0.03◦, while when 2θ was increased from 38.40 to 38.41◦, the peak shifted towards a smaller angle θ of 0.01◦.

**Figure 4.** XRD analysis of the (**a**) starting powders and the Ti6Al4V-EB composite powders, and (**b**) SPSed composites.

#### *3.2. Mechanical Properties*

To estimate the mechanical characteristics for biomedical implants, Vickers hardness and elastic modulus of the Ti6Al4V-EB composites, a function of EB contents, are shown in Figure 5a. For accuracy, the mean average values of the elastic modulus were calculated from ultrasonication method and nanoindentation curves. The increase in Vickers hardness by means of EB contents is because the hardness of hydroxyapatite contained in EB (nanohardness of 5–6 GPa and Vickers hardness of 460–480 HV) is higher than that of the Ti matrix (nanohardness of ~2 GPa and Vickers hardness of 350–400 HV) [41–44]. In addition, nanoscale hydroxyapatite was uniformly dispersed in the Ti6Al4V matrix, thereby strengthening particle effects on the hardness increment. The hardness increases as the amount of EB added increases; this increment is based on the previous studies which showed that the hardness can be increased by adding a ceramic material such as EB to the Ti matrix. When hydroxyapatite particles from EB are added to the Ti matrix, the hydroxyapatite particles are allowed to act as a barrier to dislocations [29,45]. Therefore, the values of Vickers hardness for Ti6Al4V-EB composites have been shown to range from 503.3 to ~690.1 HV, which is 1.4–2 times higher than that of pure Ti6Al4V [44]. However, the values of elastic modulus have displayed a different trend compared to those of Vickers hardness. In general, elastic modulus values for Ti6Al4V-EB composites, which range from 133.2 to 139.7 GPa, are slightly higher than those of Ti6Al4V (114 GPa); however, it depends on the fraction of β-Ti among the composites (in Figure 2). Therefore, the composite containing the highest *f*β of 19.6% showed the lowest elastic modulus value of 130.5 GPa, rather than the composite containing a *f*β of 13.6% with the 139.7 GPa. In addition, oxygen from EB is a typical stabilizing element for α-Ti—that is, as the amount of EB increased, the fraction of α-Ti having a relatively high elastic modulus was increased due to the stabilizing element. Thus, the elastic moduli of the SPSed composites also increased. Further, the elastic moduli of the SPSed samples were affected by the density; the Ti6Al4V-0.5EB composites showed the lowest elastic moduli due to having a relatively low density of 95%. Consequently, this result showed that the proper selection of process conditions for the fabrication of Ti6Al4V matrix composites could have high strength and suitable elastic moduli for use as biomedical implants. For instance, when the composites are sintered above 1200 ◦C, hydroxyapatite can phase transform to a meta-stable phase such as tricalcium phosphate, which may lead to biodegradation in the human body [40].

**Figure 5.** Mechanical properties of the Ti6Al4V and Ti6Al4V/HB composites. (**a**) Vickers hardness and elastic modulus, and (**b**) stress-strain curves from nanoindentation.
