## 2 Materials and Methods
### Characterization of the HAp-Zirconia Nanocomposite  
The obtained powder was characterized using an X-ray diffraction (XRD) Xpert powder diffractometer (PAN Analytical, Eindhoven, The Netherlands) with Cu-k\(\alpha\) radiation of 1.5406 A at a scan rate of 5 \({}^{\circ}\)/min in the scan range of 20-80\({}^{\circ}\) to determine the characteristic phase formation of the HAp-Zirconia composite. Thermal analyses such as thermogravimetry (TG) and differential scanning calorimetry (DSC) analyses of the prepared samples were performed from room temperature to 1200 \({}^{\circ}\)C at a heating rate of 10 \({}^{\circ}\)C/min using the NETZSCH STA 449F3 instrument (Yokohama, Japan). The morphological distribution of the prepared composite was determined using a transmission electron microscope (TEM, JEOL JEM 2100, Freising, Germany) and a field emission scanning electron microscope (FESEM, SIGMA HV (Carl Zeiss, Jena, Germany) with a Bruker Quantax 200-Z10 EDS detector, (Billerica, MA, USA)).  
Green pellets of the HAp-Zirconia (HZ1) composite with dimensions of 12 mm in diameter and 2 mm in thickness were prepared using a hydraulic pellet press with an applied load of 12 MPa. No binder was added in the preparation of the composite pellet. The hardness and Young's modulus of the prepared composite were determined using an Ubi 1 Scanning Quasistatic Nanoindenter (TI-700; Hystrion Inc., Billerica, MA, USA) with a Berkovich pyramidal indenter. The surface area, pore size, and pore size distribution were measured using a BELSORP Max (Microtae BEL, Osaka, Japan) utilizing the Bruner-Emmet-Teller (BET) and Barrett-Joyner-Haiden (BJH) techniques, and the experiments were performed under liquid nitrogen (\(-\)196 \({}^{\circ}\)C). The total pore volume was derived from the quantity of nitrogen adsorbed at P/P\({}_{0}\)-0.99, while the surface area was estimated at a relative pressure ranging from 0.1 to 0.5 using the BET method. Before examination of the surface area, the samples were warmed at 250 \({}^{\circ}\)C for 2 h under a vacuum to eliminate the interlayer humidity and the volatile compounds of the samples.  
The sintering process was carried out in an atmosphere of air. The samples were placed in an alumina crucible and transferred to a muffle furnace. The heating rate for the sintering process was maintained at 5 \({}^{\circ}\)/min and the system was allowed to cool down to room temperature automatically. The MG-63 human osteosarcoma cell lines were obtained from the National Centre for Cell Science (NCCS), Pune, India, and used in the cytotoxicity investigation utilizing the MTT assay. The cells were cultured in Eagle's minimal essential medium with 10% fetal bovine serum (FBS). In a CO\({}_{2}\) incubator, the cells were kept at 37 \({}^{\circ}\)C, 5% CO\({}_{2}\), 95% air, and 100% relative humidity. The cultures were switched every week, while the medium was switched twice a week. The cells were treated with different  
\begin{table}
\begin{tabular}{c c c} \hline
**Sample Code** & **HAp (Weight \%)** & **Zirconia (Weight \%)** \\ \hline Pure HAp & 100 & 0 \\ Pure zirconia & 0 & 100 \\ HZ1 & 95.01 & 4.99 \\ HZ2 & 90.49 & 9.51 \\ HZ3 & 86.39 & 13.61 \\ HZ4 & 82.64 & 17.36 \\ \hline \end{tabular}
\end{table}
Table 1: The presence of HAp and zirconia content in the as-prepared samples.  
concentrations (12.5, 25, 50, 100, and 200 \(\upmu\)g) of the composite powder for 48 h. Cells treated without nanoparticles were used as a control. The percentage of cell viability was analyzed by the MTT assay as mentioned in our previous report [30].