Dye removal, antibacterial properties, and morphological behavior of hydroxyapatite doped with Pd ions

2.1 Synthesis of Pd-HAP

Calcium chloride dihydrate [CaCl₂·2H₂O], diammonium hydrogen phosphate [(NH₄)₂HPO₄], and palladium chloride [PdCl₂] were purchased from LOBA, India, and they were utilized without further purifications. Then HAP was prepared using the co-precipitation method using appropriate molar ratios as the following equation:

In summary, different contributions of [CaCl₂·2H₂O] were used including 0.5, 0.48, 0.46, 0.44 and 0.42 M to be dissolved in 100 mL of distilled water, individually and 0.02, 0.04, 0.06 or 0.08 M of PdCl₂ were added into the accompanied Ca solution. On the other hand, 0.3 M of [(NH₄)₂HPO₄] was prepared in 100 mL as a fixed solution for each Ca. Then, (P) solution was added drop wisely into the analog (Ca + Pd) solution, whereas continuous stirring is working. The pH value was kept ad 11.0 ± 0.1. After 1 h of stirring, the solutions were aged for 24 h for precipitation, then they were washed using distilled water, and dried 50–60 °C.

2.2 Characterizations

X-ray diffraction (XRD) patterns were recorded on a (Pertpro, Cu k_α1 radiation, λ = 1.5404 Å, 45 kV, 40 mA, USA) diffractometer. The taken range at 2 $\theta$  = 5–60° and the average crystalline size of different Pd content was calculated using the well-known Debye–Scherrer equation (Mansour et al., 2017c). To confirm the functionality of Pd-HAP, the FT-IR spectrum (Model no. AUP1200343) was used, 500–4000 cm⁻¹ is the controlled range. Thermal stability of the 0.0Pd-HAP, 0.6Pd-HAP, and 1.0Pd-HAP was determined in the temperature range of 20–1000 °C (DTG 60, Shimadzu), and the thermal analysis of the samples was performed in an N₂ inert gas atmosphere with a heating rate of 5 °C/min. The surface area was measured by BET analysis (Micromeritics 3Flex equipment Tristar II 3020). The surface texture and roughness parameters were measured using a field emission scanning electron microscope (FESEM) and scan the surface roughness with an operating voltage fixed at 20 kV (model: FEG250 - QUANTA, Netherlands). Micrographs of different content of Pd-HAP were obtained using FESEM before culturing the cell line and treated using Gwyddion 2.45 software to scan their roughness (Mansour et al., 2017b). 3D images were processed for each composition and the resolution was maintained at 16,000 × 1020 pixels. The edges of the micrograph have been removed to avoid excessive borders. Therefore, the dependence of the roughness parameters on the changes in the compositions was evaluated by the program itself in (nm).

2.3 Antibacterial activity

The performance of Pd-HAP as antibacterial was studied towards gram-positive (S. aureus) and in gram-negative (E. coli) under the same conditions. The recommended initial concentration is 50 mg/ml of each one to be analyzed. The inhibition zone was measured after one day of incubation at 37 °C in the aid of visible-light illumination.

2.4 Photocatalytic set-up

A working solution to verify the MB extraction process was set at 5 mg/L of MB. Then, about 50 mg of the Pd-HAP powder phase was added to each MB beaker containing (10 mL of MB) separately at room temperature. The lighting system used here is a visible light spectrum (100 W) in a sealed box. After illumination, 3 mL of the catalyst/MB solution was removed by centrifugation to measure with a spectrophotometer at measurable time intervals. The (η %) is estimated via the expression (Abdelbar et al., 2018; Abou-Zeid et al., 2018; Salama et al., 2018; Ramadan, 2019):

3.1 Structural investigation

The XRD patterns were used to investigate the phase composition of the powder phase of Pd-HAP, and it was illustrated in Fig. 1 that HAP seems to be crystallized in a single-phase upon the ICCD card no (01-073-0293). The diffraction peaks indicated that composition has a hexagonal symmetry accruing to the space group of P6₃/m. The fact that there are no other phases that were detected through the XRD patterns might indicate that Pd(II) ions have been successfully trapped into HAP lattice.

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Fig. 1

XRD patterns of Pd-HAP at different contents of Pd(II).

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The crystallite size upon Scherrer’s equation was reported in Table 1, displays a decreasing trend with rising the content of Pd(II) starting from 36.7 nm at no additional Pd reaching to 18.6 nm at the highest contribution of Pd(II). Lattice parameters seem to be changed considerably upon the variation of Pd(II) contribution within HAP structure. The a-axis decreases from 9.447 to 9.412, 9.389, 9.361, and 9.368 Å, while the c-axis fluctuates starting from 6.875, 6.885, 6.871, 6.882 and 6.866 Å for 0.0Pd-HAP, 0.4Pd-HAP, 0.6Pd-HAP, 0.8Pd-HAP, and 1.0Pd-HAP, respectively. Moreover, c/a ratio increases when Pd is present with respect to no Pd, which indicates that HAp crystals tend to grow in the c-axis.

The two sites of Ca(II) are two perspective crystallographic positions to be replaced partially with Pd(II) ions. The coordination of Ca(1) is shaped as a tricapped trigonal prism and possesses the main effect towards the c-axis, while the Ca(2) is configured in a distorted octahedron and affects into a-axis (Jiang et al., 2002;66.). The ionic substitution of Pd(II) into either Ca(1) or Ca(2) induces a crystallographic distortion due to the difference in both ionic radii and the folds coordination between Ca(II) and Pd(II). The ionic radius of Pd(II) is around ions (0.86 Å), while it tends to be larger for Ca(II) with a value of (0.99 Å). The Pd(II) ion possesses a coordination number of (6), while the available coordinations for Ca(II) are (7) and (9) for Ca(2) and Ca(1), respectively. Therefore, to minimize lattice distortion, Pd(II) is hypothesized to replace Ca(2) sites rather than Ca(1). Due to the crystallographic position of Ca(2) through the ab plan, it might provoke a distortion in a-axis, therefore, the variation in values of a-axis matches well with the hypothetical notes. Furthermore, it was reported that the carbonate ${(CO}_3^{2-})$ group tends to replace phosphate (B type) or hydroxyl (A-type) sites individually or simultaneously (AB type). However, the AB type might be considered as the most energetically preferred crystallized orientation of HAP crystal upon Wu et al. (2020a). The theoretical density increases from 3.140 to 3.197 g/cm³ for the lowest and the highest Pd(II) contents owing to the addition of molar mass of heavy ions into the calculations.

3.2 FTIR

The FTIR spectra of the Pd-HAP compositions are illustrated in Fig. 2, whereas the characteristic bands are reported in Table 2. The bands of 472.8 cm⁻¹ indicate the bending vibrational mode (ν₂) of O–P–O, while the bands of 567.5 and 603.4 cm⁻¹ refer to the formation of bending mode (ν₄) belong to ${\mathrm{P}\mathrm{O}}_4^{3-}$ (Mansour et al., 2015; Ahmed et al., 2014). The band of 1038.4 cm⁻¹ is assigned to the asymmetric stretching (ν₃) of ${\mathrm{P}\mathrm{O}}_4^{3-}$ (Ahmed et al., 2019b, 2018). Furthermore, the bands of 1639.3 and 3844.3 cm⁻¹ indicate the absorbed OH— groups on the lattice surface. On the other hand, the bands of 874.3, 1427.3, and 1456.4 cm⁻¹ indicate the presence of vibrations of ν₂ and ν₃ belonging to CO₃²⁻ groups, respectively (Ahmed et al., 2020c, 2019a; Afifi et al., 2020). The high affinity of CO₃²⁻ ions to be absorbed through HAP structure among the synthesis process via the CO₂ from the ambient atmosphere, may introduce a facile and spontaneous way for an ionic replacement. This ionic substitution promotes crystallographic disordering within the unit cell, and thus plunge the crystallographic orientation through the lattice. The deterioration of lattice regularity induces a high rate of degradation and lowering of thermal and chemical stability, besides encouraging chemical activity towards the atomic milieu.

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Fig. 2

FTIR spectra of HAP modified with different additive concentration of Pd(II).

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3.3 Morphological features

The surface morphology of Pd-HAP depends strongly on the components’ variation as obvious in Fig. 3(a)–(d). At zero addition of Pd(II), pristine HAP seems to be formed in agglomerated rod shapes with lengths in the range of 63.4–110.3 nm, and diameters about 8.3 nm. The rods are distributed in non-oriented grains, while porosity content tends to be high. Fig. 3(b) shows that the surface of 0.4Pd-HAP was configured as aggregated rods with dimensions of (44.1–59.7) × 9.1 nm for length × diameter, respectively. The composition of 0.6Pd-HAP was formed with rods of lengths around 49.1–100.4 mm and diameters of 5.1 nm. The roughness tends to be higher, while porosity is lower than the former compositions. The highest contribution of Pd(II) into HAP seems to be shaped as a rough rod with lengths of 43.4–70.5 nm and diameters of 6.5 nm, and porosity content seems to be higher than the former ones. It could be noticed that the grain size of Pd-HAP is reduced with the rising of Pd(II) contribution. This trend is assigned to the crystallographic disordering that is provoked upon the additional ions. Therefore, controlling ionic substitution is a good tool to control shapes and grains size distribution.

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Fig. 3

FESEM micrographs of as-synthesized Pd-HAP; (a) 0.0pd-HAP, (b) 0.4Pd-HAP, (c) 0.6Pd-HAP and (d) 1.0Pd-HAP.

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The progression of surface roughness of Pd-HAP is shown in Fig. 4(a)–(d), while the roughness parameters are reported in Table 3. It is noticed that the roughness average (R_a) increases from 21.7 reaching 35.3 nm upon the substitution contribution. The root means square roughness (R_q) seems to follow the behavior of (R_a). On the other hand, the maximum height of the roughness (R_t) displays higher values than the maximum roughness valley depth (R_v) for the conjugated compositions. The roughness behavior contains two main contributors: peaks and notches. While R_t refers to the peaks, R_v denotes to the notches. Therefore, the higher values of R_t implies that the Pd-HAP seems to be conspicuous, which introduce a physical way to interact with the ambient atmosphere (Ahmed et al., 2020b, 2020a, 2020f). The adhesion with the surrounding environment is a vital property to activate the serial interaction for versatile applications including drug delivery, antibacterial agency, and water filtration (Ahmed et al., 2020b, 2020a, 2020c). Consequently, the roughness manipulation, and thus the activity is controlled via the compositional tailoring.

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Fig. 4

Development of surface roughness upon the variation of Pd(II) contributions through Pd-HAP; (a) 0.0pd-HAP, (b) 0.4Pd-HAP, (c) 0.6Pd-HAP and (d) 1.0Pd-HAP.

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3.4 Thermal stability

The thermogravimetric analysis has been done for Pd-HAP at different concentrations of Pd(II) up to 1000 °C in ambient air as shown in Fig. 5. It is noticed that the rate of weight loss seems to be high for all composition starting from room temperature (R. T.) up to around 70 °C, whereas the compositions lost about 3.7% of their initial weights. The weight loss in this stage is referred to as the release of the absorbed water molecules. Therefore, all curves seem to be identical in the first stage.

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Fig. 5

Thermogravimetric analysis of Pd-HAP upon the contributions of Pd(II) through the powder phase.

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The second stage continues upon 418 °C, and obvious that the loss rate increases significantly upon the content of Pd(II), where the powdered compositions of 0.0Pd-HAP, 0.6Pd-HAP, and 1.0Pd-HAP loss around 9.2, 15.7, and 14.6% from their weights at 418 °C, respectively. In this stage, the compositions tend to be degenerated to release hydroxyl ions, which is the initial stage before transforming to calcium oxide phases. The presence of Pd(II) through the crystal provokes high distortion, and thus promote a high tendency to destabilize, besides the high specific surface area, which encourages thermal decomposition in lower relative temperatures. Moreover, the third stage represents a plateau up to 1000 °C, whereas the samples lose around 10.4, 17.5, and 15.6% from their initial weight at 1000 °C for 0.0Pd-HAP, 0.6Pd-HAP, and 1.0Pd-HAP, respectively. The highest weight loss was recorded at 0.6Pd-HAP to be 17.5%, which might indicate relative thermal stability for the as-prepared compositions.

3.5 BET isotherm

The minimization of the usage doses of an antibacterial agent is pivotal for reducing the environmental loading. The increase of the effective surface could permit the use of low concentrations of the same composition, which could be accomplished by reducing the size of particles. The calculated surface area was around 28.3, 42.0, and 63.4 m²/g for 0.0Pd-HAP, 0.6Pd-HAP, and 1.0Pd-HAP, respectively as mentioned in Table 4. The total pore volume increases from 19 × 10⁹ to 43 × 10⁹ m³/g, while the average pore radius grows from 1.82 reaching 1.97 nm for the lowest and the highest Pd(II) contributions. The average values of pore size indicate that porosity could be classified as micro-porous compositions. As noticed from Fig. 6, the N₂ adsorption/desorption isotherms behaviors of Pd-HAP denote the slope of the absorbance curve increases from the low to the high Pd(II) concentrations. This behavior matches well with the results from XRD and FESEM that Pd(II) acting as a crumbling factor within HAP crystals, and thus might reduce grains size, increase porosity, and consequently promote activity. Furthermore, it could be noticed from the obtained hysteresis loops that incorporation of Pd(II) ions into HAP lattice has caused a transition of microporous structure. Thus, the surface area was enlarged upon the additional Pd(II).

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Fig. 6

The N₂ adsorption/desorption isotherms behaviors of Pd-HAP upon the changing of Pd(II) ions.

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3.6 Anti-bacterial activity

The ability of Pd-HAP to degenerate bacterial cells was investigated against both E. coli and S. aureus (gram-positive) as illustrated in Fig. 7(a)–(c). The control sample was considered to be a bacterial colony without additional antibacterial substances. The pristine HAP does not show an antibacterial effect, however, elevating Pd(II) contribution induced growing activity, as the inhibition zone increased from 2.2 ± 0.4, 2.6 ± 0.6 mm reaching to the largest area of 4.3 ± 0.9 and 4.5 ± 0.8 mm against both E. coli and S. aureus for 0.4Pd-HAP and 1.0Pd-HAP, respectively. The rising activity of the Pd-HAP could be assigned to the ionic release of Pd(II) through the culture medium. The Pd(II) ions displayed high chemical activity towards bacterial cell walls, which may cause degeneration of bacterial growth (Gholami Derami et al., 2020). The compositions with high values of Pd(II) display significant antibacterial behavior owing to the adequate quantity of Pd(II) that could be released through the media, besides the high dissolution rate accompanied by the high level of ionic substitutions.

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Fig. 7

Antibacterial behavior of Pd-HAP against E. coli and S. aureus.

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3.7 Photocatalytic investigation

HAP displays selective adsorption behavior towards versatile organic matrices; however, its morphological features, optical properties, and structural configuration control its photocatalytic activity. Photodegradation using Pd-HAP was done against MB dye under visible light at R. T as illustrated in Fig. 8(a)–(e). The MB absorption spectra decrease exponentially upon irradiation time. The calculated degradation efficiency (η) of Pd-HAP reached its highest values of 26.35, 38.74, 60.34, 71.76, and 86.4 after 120 min of continuous irradiation for compositions of 0.0Pd-HAP, 0.4Pd-HAP, 0.6Pd-HAP, 0.8Pd-HAP, and 1.0Pd-HAP, respectively. On the other hand, the degeneration of dye is noticed to obey pseudo-first-order kinetics as follows (Shoueir et al., 2019; Omrani and Nezamzadeh-Ejhieh, 2020):

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Fig. 8

UV–vis spectral degradation of MB towards selected (a) 0.6Pd-HAP, (b) 0.8Pd-HAP, (c) 1.0Pd-HAP, (d) kinetic model, and (d) reusability performance.

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The photocatalytic interaction occurs when an incident photon with sufficient energy bombard a surface of a substance with a moderate bandgap. The direct interaction between photon and electron might generate electron-hole pairs (excitations) (Reeta Mary et al., 2018). The created photoelectrons jump up to the conduction band level (C.B), while the conjugated hole moves in the valence band (V.B). Next, the created charge carriers trigger redox serial reactions against dye molecules after their surface of a substance. These reactions may begin with a break of water molecules to generate a free radical involving ( ${\mathit{OH}}^{·-})$ and electron, which is able to induce a formation of superoxide ( $O_2^{·-})$ , besides the hydrogen peroxide anions ( $H{O}_2^{-}$ ). These obtained free radicals are responsible to degrade the targeted dye, whereas the degradation rate depends strongly on the composition of dye and the light source, besides the compositional characteristics of the photocatalyst including its bandgap and surface area (Prabhu et al., 2018).

Furthermore, the configuration of HAP crystal possesses versatile superficial charges; whereas the a-surface is rich with ( ${\mathrm{P}\mathrm{O}}_4^{3-}$ ) anions, while the c-surface is decorated with Ca(II) cations. However, the HAP has a tendency to be crystallized in c-direction; therefore, a-surface is expected to obtain a higher surface area than c-face. Thus, the photogenerated electron through the ( ${\mathrm{P}\mathrm{O}}_4^{3-}$ ) is hypothesized to form a vacancy through HAP surface followed by a free radical generation as illustrated in Fig. 9. Hence, the dye molecules could be absorbed on the surface of HAP. Herein, it might be reported that surface roughness favors the adsorption of the dye matrix via both physical and chemical interactions (Mohseni-Salehi et al., 2018; Shoueir et al., 2020a). The chemical type is encouraged via the high content of crystallographic defects that acts as ionic traps, while the physical one is done by the adhesion on the peaks of grains. The ability to remove the agent to be reused several times represents an important economic issue. Therefore, the reusability of the Pd-HAP has been investigated for 6 cycles and showed that the removal efficiency deteriorated to be about 5.65, 8.14, 6.24, 8.76, and 10.2% for different contents of Pd(II), which indicates moderate stability for these compositions.

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Fig. 9

Schematic diagram of photocatalysis process to degrade MB via Pd-HAP compositions.

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