Next Article in Journal
Effect of B2O3 on the Structure, Properties and Antibacterial Abilities of Sol-Gel-Derived TiO2/TeO2/B2O3 Powders
Previous Article in Journal
Indoor Air Pollutant (Toluene) Reduction Based on Ultraviolet-A Irradiance and Changes in the Reactor Volume in a TiO2 Photocatalyst Reactor
 
 
Font Type:
Arial Georgia Verdana
Font Size:
Aa Aa Aa
Line Spacing:
Column Width:
Background:
Article

Enhanced Release of Calcium Ions from Hydroxyapatite Nanoparticles with an Increase in Their Specific Surface Area

by
Urszula Szałaj
1,2,*,
Agnieszka Chodara
3,
Stanisław Gierlotka
1,
Jacek Wojnarowicz
1 and
Witold Łojkowski
1
1
Laboratory of Nanostructures, Institute of High Pressure Physics, Polish Academy of Science, Sokolowska 29/37, 01-142 Warsaw, Poland
2
Faculty of Materials Engineering, Warsaw University of Technology, Wołoska 41, 02-507 Warsaw, Poland
3
Leyton Poland Ltd., Wspólna 70, 00-687 Warsw, Poland
*
Author to whom correspondence should be addressed.
Materials 2023, 16(19), 6397; https://doi.org/10.3390/ma16196397
Submission received: 3 September 2023 / Revised: 21 September 2023 / Accepted: 22 September 2023 / Published: 25 September 2023
(This article belongs to the Topic Advances in Biomaterials)

Abstract

:
Synthetic calcium phosphates, e.g., hydroxyapatite (HAP) and tricalcium phosphate (TCP), are the most commonly used bone-graft materials due to their high chemical similarity to the natural hydroxyapatite—the inorganic component of bones. Calcium in the form of a free ion or bound complexes plays a key role in many biological functions, including bone regeneration. This paper explores the possibility of increasing the Ca2+-ion release from HAP nanoparticles (NPs) by reducing their size. Hydroxyapatite nanoparticles were obtained through microwave hydrothermal synthesis. Particles with a specific surface area ranging from 51 m2/g to 240 m2/g and with sizes of 39, 29, 19, 11, 10, and 9 nm were used in the experiment. The structure of the nanomaterial was also studied by means of helium pycnometry, X-ray diffraction (XRD), and transmission-electron microscopy (TEM). The calcium-ion release into phosphate-buffered saline (PBS) was studied. The highest release of Ca2+ ions, i.e., 18 mg/L, was observed in HAP with a specific surface area 240 m2/g and an average nanoparticle size of 9 nm. A significant increase in Ca2+-ion release was also observed with specific surface areas of 183 m2/g and above, and with nanoparticle sizes of 11 nm and below. No substantial size dependence was observed for the larger particle sizes.

1. Introduction

Over the past years, the incidence of diseases and injuries of the skeletal system has significantly increased worldwide. These conditions are caused by the aging of the population, as well as by congenital defects, sports, traffic injuries, and other diseases [1,2,3]. The aim of tissue engineering is to accelerate bone-tissue regeneration and to enable the filling of bone defects with natural bone when bone cannot be regenerated by natural means. Treatments using biological agents, stem cells, biomimetic scaffolds, or suitable implants provide increasingly effective and reliable strategies for creating bone tissue and regenerating large defects, thus improving the quality of patients’ lives [3].
Bone tissue is composed of 60% inorganic constituents (mainly nanohydroxyapatite), 30% the organic constituent (proteins), and 10% water [4]. Natural nanohydroxyapatite contains numerous impurities in the form of potassium, magnesium, strontium, sodium, chloride, fluoride, and carbonate [4]. The second inorganic constituent of bone is whitlockite (Ca18Mg2(HPO4)2(PO4)12) [5,6]. The organic part of bone is composed mainly of type I collagen (ca. 90%) and non-collagen proteins [7,8]. Synthetic calcium phosphates, e.g., hydroxyapatite (HAP, Ca10(PO4)6(OH)2), calcium α-, and β-triphosphate (TCP, Ca3(PO4)2), are the subjects of continuing interest in the field of tissue engineering due to their high chemical similarity to natural hydroxyapatite [9,10,11,12]. Recent research has focused on the artificial production of nano-HAP that is as close as possible to natural HAP in terms of structure [13]. Numerous studies have confirmed the biocompatibility of nano-HAP and its usefulness in bone-tissue regeneration [14,15,16,17,18,19,20,21,22,23,24,25,26].
Calcium in the form of free ions or bound complexes plays a key role in many biological functions. The amount of calcium in the adult body is, on average, 1000 g. This element plays a key role in the mineralization of the skeleton and in other biological processes [27]. Calcium (Ca2+) is an intracellular messenger that controls several cellular processes, such as cell proliferation, gene transcription, and muscle contraction. The signals of Ca2+ involve a number of homeostatic and sensory mechanisms. These Ca2+ signals may induce the expression of genes that are related to bone-cell proliferation in cells [28]. In vitro studies have shown that the calcium contained in bone-regrowth scaffolds supports the increased adhesion, proliferation, and differentiation of osteoblastic MG-63 cells. Further, calcium signals promote osteoblast function through calmodulin, and through the activation of extracellular-signal-regulated kinase 1/2 (ERK1/2) and of the intracellular signaling pathway, which is important in regulating the cell cycle (PI3K/Akt pathways) [28,29,30]. In addition, calcium signals from the endoplasmic reticulum (ER) and the activation of calcineurin cause the nuclear factor of activated T cells, especially in the introduction of the IL-2 or IL-4 gene (NFAT2)’s dephosphorylation and osteoclastic gene expression [31]. The expression and control of osteoclastic genes indicates the role of calcium in bone resorption and homeostasis [31,32]. In vivo studies have shown that Ca2+ ions released from HA/TCP-composite scaffolds increase bone formation in rat calvarial bone defects [33]. In addition, Ca2+-coated titanium implants resulted in increased bone density and osteointegration in a sheep-tibial-bone model [28,30,31,32]. Thus, the control of the Ca2+-ion release to induce bone-tissue repair is necessary for the appropriate application of calcium-phosphate materials in tissue engineering and regenerative medicine.
The effect of particle size on particle solubility and bioavailability has been documented by many researchers [12,33,34,35,36,37,38,39,40,41,42]. This relationship is exploited mainly in pharmacy for the purpose of increasing the bioactivity of drugs [33,34,35,36,37,38]. A reduction in the size of drug particles to nanometer size increases the total effective surface area and dissolution rate. Moreover, a reduction in the particle size leads to a decrease in the thickness of the diffusion layer surrounding the drug particles, resulting in an increase in the concentration gradient [37]. Regarding hydroxyapatite, the sizes of HAP nanoparticles affect the degradation rate of this material [43].
The microwave hydrothermal synthesis (MHS) of hydroxyapatite nanoparticles (HAP NPs), hereinafter referred to as GoHAP, which was recently developed in our laboratory, makes it possible to control their size with nanometric precision in the range of 9 to 50 nm [13]. Its advantages include the possibility of obtaining HAP NPs that meet the requirements of purity for medical applications. This is made possible by taking advantage of microwave heating [44] and the lack of harmful by-products of the synthesis [13]. The literature [45,46,47,48,49,50,51,52,53,54,55,56,57,58,59,60,61] exhaustively describes and discusses the advantages and disadvantages of the methods of nanohydroxyapatite synthesis, e.g., for chemical precipitation, hydrothermal methods, hydrolysis, sol–gel methods, microwave irradiation, chemical vapor, the combustion technique, the pyrolysis technique, and solid-state and mechanochemical methods. If, as producers, we wish to evaluate the quality of the obtained HAP NPs for applications in bone-tissue regeneration, it is most important that the applied method makes it possible to obtain a product that is repeatable in terms of size, size distribution, shape, crystallinity, chemical purity, and phase purity [53].
In this paper, we take advantage of our original HAP-synthesis technology [13,62] to precisely determine the effects of GoHAP size and specific surface area on the Ca2+ concentration in a PBS dispersion. If the relationship between the amount of ions released, the size of the specific surface area, and the sizes of the hydroxyapatite nanoparticles is known, it is possible to program hydroxyapatite resorption [42] and create biodegradable bone grafts [53,63], or layers on titanium implants [64,65,66]. In addition, it is possible to control the degradation rate and the bone-growth-stimulation potential of biodegradable electrospun membranes coated with HAP NPs [67,68], as well as titanium implants coated with HAP NPs. The control of the resorption rate of HAP NPs may be the key to achieving a balance between the rate of material degradation and accelerated bone-tissue regeneration. This will make it possible to tailor the sizes of nanoparticles to physicians’ requirements in specific applications.

2. Materials and Methods

2.1. Materials and Synthesis of Nanoparticles

The MHS-synthesis procedure is based on the method described in [13]. Calcium hydroxide (pure Ca(OH)2, CHEMPUR, Piekary Śląskie, Poland) and orthophosphoric acid (85% solution H3PO4, analytically pure, CHEMPUR, Piekary Śląskie, Poland) were used for the synthesis. The synthesis was carried out in deionized water (0.06 μS/cm) purified by a water double-deionization system (HLP 20 UV deionizer, Hydrolab, Straszyn, Poland, and Ultra Toc/UV/UF, Hydrolab, Straszyn, Poland).
The GoHAP type 1 was obtained by a precipitation reaction:
10 Ca(OH)2 + 6 H3PO4 → Ca10(PO4)6(OH)2 + 18 H2O
The amount of each component was adjusted to obtain calcium-deficient HAP (CDHAP) with a Ca/P ratio of 1.51. In the next step, the obtained suspension was poured into a Teflon vessel with a volume of 270 cm3, which was closed tightly and inserted into the high-pressure chamber of the homemade MSS2 microwave reactor in the batch mode (IHPP PAS (Warsaw, Poland), ITeE-PIB (Radom, Poland), ERTEC (Wroclaw, Poland) [69]), as described in [13]. After switching on the high-power magnetrons, microwave energy was delivered to the vessel using a waveguide. The temperature was calculated from the vapor–liquid equilibrium for water [69].
The power of the magnetrons was set to 3 kW. Time was counted from the moment the power was switched on. After reaching the pre-set pressure, the mean magnetron’s power was adjusted to keep the pre-selected pressure for a programmed time. Table 1 shows the values of up-heating time, total time, and pressure.
For each set of parameters, we produced 6 samples with a weight of ca. 7 g. The reaction products consisted of GoHAP particles and water only. The powders were separated from water by freeze-drying for 72 h (Lyovac GT-2, SRK Systemtechnik GmbH, Riedstadt, Germany).

2.2. Characterization of Nanoparticles

The powder X-ray diffraction (XRD) data were collected by the PANalytical X’Pert Pro diffractometer using monochromatic Cu Kα1 radiation and the PIXcel position-sensitive detector. The measuring range was 10–80° and the step was 0.03°.
Scherrer’s formula was used to determine the mean crystallite size [70]. The shapes of the crystallites were considerably anisotropic, and, therefore, their lengths and widths were determined by the analysis of the XRD peak width for the 002 and 300 Bragg reflections.
The specific surface area (SSA) was examined by the Brunauer–Emmett–Teller (BET) isotherm method with the use of a Gemini 2360 surface analyzer (V 2.01, Micromeritics®, Norcross, GA, USA), in accordance with ISO 9277:2010 [71].
The density (DEN) was examined using a helium pycnometer (AccuPyc II 1340, Micromeritics®, Norcross, GA, USA), in accordance with ISO 12154:2014 [72].
Before the SSA and DEN measurements, samples were dried in a VacPrep 061 desorption station (Micromeritics®, Norcross, GA, USA) for a period of 2 h at 150 °C in vacuum (0.05 mbar).
The mean diameter of GoHAP, also known as the Sauter mean diameter (SMD), was calculated based on the SSA and DEN measurements using Equation (2), with the assumption of a spherical shape:
S M D = A S S A · 10 18 · D E N · 10 21   nm
where SMD is the Sauter mean diameter of the nanoparticle (nm), A is the shape factor, equal to 6 for the sphere, SSA is the specific surface area (m2/g), and DEN is the density (g/cm3). This method of determining the SMD of GoHAP was described previously [73].
The specific-surface-area and density tests were carried out in a laboratory [74], working in accordance with PN-EN ISO/IEC 17025:2018-02 [75].
Transmission-electron microscopy (TEM) imaging was carried out using a JEOL JEM 2000EX apparatus with a beam with 200 keV of energy. Images were recorded on photographic plates and then processed into digital form using a NIKON LS-8000 ED scanner (Nikon, Tokyo, Japan). The powder samples were deposited on a 3-millimeter-diameter copper grid covered with a perforated carbon membrane, catalog symbol S147-4H, from Agar Scientific (Essex, UK). Observations were made using bright- and dark-field imaging. The size distribution of the NPs was determined by the bright field and the dark field based on the theoretical model, assuming spherical particles with a log-normal size distribution. The diameters were determined for at least 130 particles in each sample, and a histogram of the number of particles with diameters in the given range of values was created. The average particle size was calculated as an arithmetic mean using Excel software, version 2308 (Microsoft, Warsaw, Poland).

2.3. Determination of the Amount of Ca2+ Ions Released

For this purpose, as well as to determine the chemical composition of the produced samples, inductively coupled plasma—optical emission spectrometry (ICP-OES) with induction in argon plasma (iCAP 6000series, Thermo Scientfic, Cambridge, United Kingdom) was used. The samples for the tests were prepared as follows.
The nanopowders were dried for 12 h at (100 ± 2 °C). Next, 0.1 g of each type of powder was weighed and placed in plastic containers with a volume of 50 mL. Subsequently, 20 mL of the phosphate-buffered saline (PBS) solution, pH = 7.4 ± 0.1 (Sigma Aldrich, Saint Louis, MS, USA), was added to the containers using a pipette. The sealed plastic containers were placed in a water bath (Heating Bath B-491, BUCHI, Flawil, Switzerland) at 37 ± 1 °C. The batch was shaken in the longitudinal motion at 2 Hz. Ion-concentration analyses using the ICP-OES technique were carried out on the following days: 1, 3, 7, 9, 14. For each time point, analyses of 2 samples of each type were carried out.
The PBS samples for the ICP-OES tests were collected by filtering the particles from the suspension.
To determine the chemical composition, 0.2 g each of GoHAP type 1–type 6 was collected from the containers and transferred to the Teflon (polytetrafluoroethylene) container of the microwave mineralizer (Magnum II, Ertec, Wroclaw, Poland). Next, 20 mL of HCL and 4 mL of HNO3 were added. After 10 min of treatment at a power of 800 W, the powder sample was dissolved. An ICP analysis was then carried out to determine the content and ratio of Ca2+, PO43− ions, and trace elemental impurities: Mg, Si, Al, Fe, Na, Mn. The procedure was repeated for each powder sample.
An analysis of pH and conductivity of the PBS buffer was performed using a pH meter (SevenExcellennce, Multiparameter, Mettler Toledo, Greifensee, Switzerland). An InLab Expert Pro-ISM pH electrode with a built-in temperature sensor was mounted to the instrument to measure pH, and a conductivity probe, InLab 731-ISM Cond (Mettler Toledo, Greifensee, Switzerland), was used to measure conductivity. The temperature of the buffers was maintained at 35 °C using an electric heater (babyono, Natural Nursing, Poznan, Poland). The pH electrode was calibrated with Mettler Toledo technical buffers at three pH points: 4.03, 7.00, and 9.01, respectively. Measurements were made by immersing the electrode in the PBS solutions, which were prepared in the same manner as for the ICP-OES measurements. After each measurement, the electrode was rinsed with deionized water with a conductivity of 0.06 μS/cm and treated in a double deionization system (HLP 20 UV, Hydrolab, Straszyn, Poland, and Ultra Toc/UV/UF, Hydrolab, Straszyn, Poland).

3. Results

3.1. Synthesis of Nanoparticles

The MSS2 reactor allows the rapid heating of reactants, a rapid pressure increase, fast cooling, and, thus, excellent control of the reaction time [69]. Figure 1 shows the pressure–time plots for the GoHAP synthesis. Table 1 lists the produced GoHAP types depending on the process parameters. Time was counted from the moment the magnetron was switched on and the start of the power delivery. The inset in Figure 1 reveals that there was a delay of ca. 43 s between the moment power was switched on to the moment when the pressure sensors of the reactor registered an increase in pressure. Thus, after 43 s, the pressure started to exceed the atmospheric pressure, indicating that the temperature exceeded 100 °C. The heating rate was in the range of 1–2 °C/s.
The operation of the magnetrons was switched from full power to the pulsed mode so that the mean power could be adjusted to keep a constant pressure after the heating time. This allowed the nanoparticles to grow. For GoHAP type 2, the heating was interrupted after 55 s and the power was switched off. The heating time was 60 s for the GoHAP type 3. To shorten the time needed to produce larger particles, the pressure and temperature were raised. The particle-growth phase was 500 s for the GoHAP type 4, 480 s for the GoHAP type 5, and 1000 s for the GoHAP type 6. The unique characteristics of the microwave hydrothermal technology using the MSS2 reactor in terms of time and pressure control are evident.

3.2. Characterization of Nanoparticles

The characteristics of the GoHAP nanomaterial—the specific surface area, density, and particle size of the produced GoHAP hydroxyapatite—are presented in Table 2 and Table 3. When combining information from Table 1 and Table 3, it can be seen that an increase in the process pressure and temperature or time leads to an increase in the particle size.
Figure 2 presents the particle-size distributions of the GoHAP particles, which were obtained from the analysis of the TEM images. In line with the increase in the synthesis temperature and in the synthesis time, the increase in the average particle size and in the size distribution is visible. One should note the results of the sizes of the Type 2 and Type 3 samples: despite the identical average particle size (13 nm), these samples had different particle-size distributions.
Figure 3 shows a correlation between the density and the specific surface area. The greater the mean particle size (and, thus, the smaller the specific surface area), the greater the density.
Figure 4 shows X-ray-diffraction spectra for all the GoHAP nanoparticles studied. The diffraction peaks correspond to pure hydroxyapatite. The decrease in the width of the diffraction peaks indicates the increasing mean size of the GoHAP nanoparticles.
Table 4 shows the Ca/P ratio for the GoHAP type 1–type 6 nanoparticles. The ICP-OES analysis showed that they consisted of calcium and phosphorus, in a ratio of 1.52 ± 0.01. In addition, minor contamination with Mg, Si, Al, Fe, Na, and Mg was observed (Table 5). The ratio of calcium to phosphorus indicates that the produced nanoparticles represent calcium-deficient hydroxyapatite (CDHA). The ion-substituted CDHA has Na+, K+, Mg2+, Sr2+ for Ca2+, CO32− for PO43− or HPO42−, and F, Cl, CO32− for OH-, and with water it forms biological apatite —the main inorganic part of animal and human bone in normal and pathological calcifications [76,77].
The total impurity content was 0.40 ± 0.06 wt.%. The differences in impurity content between the GoHAP types were less than 15% of the total impurity content.
The TEM images (Figure 5 and Figure 6) obtained using the bright- and dark-field techniques showed differences between the shapes and sizes of the GoHAP type 1–type 6 nanoparticles. The GoHAP type 1 nanoparticles had the smallest sizes and a needle-like shapes. A very similar shape was obtained for the GoHAP Type 2 particles. As the nanoparticles grew, they took on increasingly spherical shapes (GoHAP type 3–type 6).

3.3. Calcium-Ion-Release Results

This study showed a strong effect of the specific surface area of GoHAP on the amount of ions released (Figure 7). The release of calcium ions at the highest concentrations, i.e., ca. 18 mg/L, was observed for the GoHAP type 1 with a specific surface area of 240 m2/g and an average nanoparticle size of 9 nm. As the nanoparticle size increased and the specific surface area decreased, the amount of calcium ions released into the buffer solution decreased. Compared to the GoHAP type 1 (SSA 240 m2/g), a significant decrease in solubility and in the associated calcium-ion release was already observed for the slightly larger GoHAP type 2 nanoparticles, with an average nanoparticle size of 10 nm (183 m2/g), and the GoHAP type 3, with an average nanoparticle size of 11 nm (108 m2/g). The decrease in the amount of released Ca2+ ions was ca. ↓7 mg/L, with a difference in the developed specific surface area of 30 m2 /g (GoHAP type 2) and ca. ↓12 mg/L, and with a difference in the developed specific surface area of 57 m2/g (GoHAP type 3) compared to the GoHAP type 1. Small amounts of Ca2+-ion release, ranging from ca. 2.3 mg/L to 4.1 mg/L, were observed for the GoHAP type 4–6 nanoparticles. Therefore, it seems that there is a threshold nanoparticle size at 11 nm and a threshold SSA at 108 m2/g, above which Ca2+-ion release becomes size-independent.
In each of the GoHAP types studied, the Ca2+-ion concentration stabilized after 1 day in the buffer solution (Figure 7). After this time, nearly constant levels of calcium-ion release were observed, which indicates that an equilibrium state was achieved.
The changes in the conductivity and pH of the solution were correlated with the amount of calcium ions released (Figure 8 and Figure 9). The largest increase in conductivity was observed when the GoHAP type 1 nanoparticles with the smallest sizes and the highest specific surface areas were dissolved (Figure 8). The increase in conductivity decreased as the specific surface area of the GoHAP decreased.
During the calcium-ion release tests, the pH increased steadily, depending on the development of the specific surface area, and maintained the following relationship: highest specific surface area—highest pH; lowest development of specific surface area of nanoparticles—lowest pH (Figure 9). However, the differences in the pH values of the solutions during the solubility testing of the GoHAP type 1 (240 m2/g) and type 6 (51 m2/g) nanoparticles were small and accounted for about 0.05 points on the pH scale.

4. Discussion

4.1. Structural and Chemical Characterization

The studies of the GoHAP nanostructure confirmed a gradual increase in the particle size in line with the increase in time and in pressure and temperature of the microwave synthesis. In addition, the particle shape transformed gradually from a plate or a needle to an ellipsoid. The possible mechanism underlying the change in the shapes of GoHAP particles was described in the paper by Kozerozhets et al. [78]. Furthermore, the pycnometric density of the particles gradually increased in line with the time and with the pressure and temperature of the synthesis. The obtained GoHAP nanoparticle samples had a lower density than the theoretical density of hydroxyapatite, which is 3.15 g/cm3 [79,80]. This correlation was previously found for zirconia (ZrO2) nanoparticles [81] and doped and undoped zinc oxide (ZnO) nanoparticles [82,83,84]. It is attributed to the effect of the surfaces of nanoparticles on their mean density. Even nanoparticles with the maximum possible degree of crystallinity have dangling bonds on their surfaces, to which -OH groups in oxides may attach [81]. In addition to the presence of hydroxides, another reason for the density–size correlation is that the thickness of the amorphous phase on the nanoparticle surface decreases as the size of the nanoparticles increases [83]. In a pycnometer study [85,86], such surfaces contributed to the volume occupied by the nanoparticle, so that the mean density measured decreased as the specific surface area increased.
The hydroxyapatite unit cell (both synthetic and natural) usually displays a hexagon crystal system, with a P63/m space group [87,88], even if the occurrence of monoclinic HAP is well-known [89]. The crystal structure of the GoHAP type 1 sample was identical to that of human bone [90], while the crystal structure of the GoHAP type 6 sample was identical to that of human tooth enamel [13]. The XRD data show that the particles had non-spherical shapes. The smallest, type 1 and type 2, had platelet shapes, with the larger surface parallel to the (100) planes. The aspect ratio varied for the particles with SSA values equal to or above 183 m2/g in the range of 1.7–4.2.
The differences between the sizes, as measured by means of the XRD method and calculated from the SSA, were not significant, because the SSA delivered a mean value, while the XRD data depended on the axis selected for the analysis. Further, in the case of SSA, the smallest particles may have delivered the highest contribution to the surface area, while in the case of XRD studies, they may have disappeared into the background due to very broad peaks.
When comparing the results of the crystallite sizes calculated from the XRD data with the results of the average particle sizes calculated from the TEM images, it was found that they were virtually identical or fell within the standard deviation. This means that monocrystalline hydroxyapatite was obtained in all the samples (one particle was built of one crystallite). The results of the TEM imaging are consistent with those of the XRD analysis—for the smallest particles, especially type 1 and type 2, the shape was platelet- or needle-like, while for the larger particles, the shape was close to spherical.
The structural characterizations showed that the MHS method permits the production of a high-purity nanomaterial with a constant chemical composition as a function of size. The density increases in line with the increase in size, due to the decreasing fraction of atoms situated on the surface. The smallest particles, with SSA values below 183 m2/g, have a highly non-equilibrated elongated shape, which is characteristic of particles formed in a short time. For longer times or higher temperatures, the aspect ratio of the nanoparticles decreases, tending towards greater sphericity.
Taking all these results into consideration, it is justified to regard SSA as the main characteristic parameter of the nanomaterial, on which all other properties, size, shape, and density, depend.
Regarding the chemical composition, no SSA effect was detected. The calcium/phosphorus ratio for calcium-deficient hydroxyapatite (CDHA) (Ca10x(HPO)4x(PO)46−x(OH)2−x (0 < x < 1) is 1.5–1.67. As shown in Table 3, x = 1.52, the calcium/phosphorus ratio corresponds to human HAP. A calcium-deficient composition was selected specifically for this study, so that the synthetic GoHAP would mimic the natural nano-HAP as much as possible.
The total impurity content, as shown in Table 4, was 0.40 ± 0.06 wt.%. The differences in the impurity content between the various GoHAP types were less than 15% of the total impurity content.

4.2. Ion-Release Study

For standard commercial hydroxyapatite with particle sizes much larger than those of nanomaterials, the most important parameters are the molar Ca/P ratio, basicity/acidity, and solubility. The lower the Ca/P molar ratio, the more acidic the powder and the more water-soluble the calcium orthophosphate [76,77]. In this respect, hydroxyapatite is one of the most stable calcium phosphates.
However, as the present study shows, a reduction in the sizes of HAP nanoparticles results in active nanoparticles that release calcium ions, i.e., the stability of the particles decreases. They are therefore bioactive and potentially biodegradable. For nanoparticles smaller than 39 nm immersed in PBS, calcium-ion release is observed, but solubility increases significantly for a threshold size lower than 11 nm. The greatest differences in nanoparticle solubility were observed in the nanoparticles with specific surface areas in the range of 240–183 m2/g (Figure 10).
Such properties are sought and explored in the field of nanotechnology, in which the chemical composition of the material is constant, but the size of the particles or the crystallites of the material are variable. This was the case in the present paper. The specific surface area is defined as the external surface area of a substance per unit mass of that substance. This ratio depends on the parameters of size and shape; the smaller the solid and the more needle-like its shape, the greater the specific surface area. In nanoparticles with dimensions of several nm, large fractions of atoms are situated on the surface and, therefore, their free energy is high compared to large particles [91,92] (Figure 11).
The concentration of Ca2+ ions in PBS depends on the SSA value, and not on time. This effect can be explained in terms of an equilibrium between the liquid phase and the specific surface of the solid phase, rather than its mass. This effect is in line with the treatment of the specific surface area (per unit volume or per unit weight) as an independent thermodynamic variable. We regard SSA as the crucial parameter. This is because the sizes of nanoparticles are difficult to determine both with microscopy methods and with XRD methods, especially for complicated shapes, and when there is a size distribution. A focus on the specific surface area as a key parameter is described in multiple papers [93,94]. The effect of the specific surface area’s value (above 183 m2/g) on the solubility of HAP nanoparticles, which we observed, was explained in the paper by Fu et al. [95]. It should be noted that Fu et al. [95] assumed spherically shaped nanocrystals for the purpose of their calculations. The calculated results of the thermodynamic properties of the surfaces showed that the limiting size (diameter) of the nanocrystals was 20 nm. When the size was less than 20 nm, the effect of the particle size on the thermodynamic properties of the surface increased and deviated from linear variation. Spherical HAP nanoparticles measuring 20 nm have specific surface areas of ca. 100 m2/g and the assumption of a diameter of less than 20 nm (specific surface area above 100 m2/g) was confirmed by our four samples, i.e., from GoHAP type 1 (240 m2/g) to Go HAP type 4 (108 m2/g). Fu et al. [95] also discovered that an important factor, in addition to the specific surface area value itself, in the thermodynamic properties is the shape of the nanocrystals. With an identical equivalent diameter of particles, the more the shape deviates from sphere, the stronger the thermodynamic properties of the surface (absolute value) [95]. The shape-criterion and the specific-surface-area values were displayed only by the GoHAP type 1 and GoHAP type 2 samples, in which we observed significantly increased hydroxyapatite solubility. To the best of our knowledge, our study is the first to report the effect of the specific surface area’s value on nanohydroxyapatite solubility [39,96,97,98,99,100,101,102,103,104,105,106,107,108,109,110,111,112,113,114,115,116]. It must be underlined that the novelty of our paper in relation to the papers reported previously [39,96,97,98,99,100,101,102,103,104,105,106,107,108,109,110,111,112,113,114,115,116] is in the values of the specific surface area of the HAP-NP samples that we used in the solubility tests (from 51 m2/g to 240 m2/g). A good comparative example is the paper by Tang et al. [39], who examined the size effects in the dissolution of hydroxyapatite for HAP-NP samples with specific surface areas of 24.2 m2/g, 32.4 m2/g, and 55.1 m2/g. The dissolution studies the authors carried out for the different undersaturations lasted only 100 min. Our tests lasted 11 days and referred to the state of an excess amount of solid HAP NPs in the suspension relative to the solubility product (which was significantly above the state of equilibrium). Tang et al. [39] discovered that in unsaturated biological environments, there is a metastable HAP phase that depends on the effects of particle sizes, resulting in the self-inhibition of dissolution, or even the suppression of the dissolution reaction.
If the ion concentration, the volume of the PBS, and the weights of the nanoparticles are known, it is possible to calculate the amount of dissolved GoHAP. For the highest Ca2+-ion concentration, 18 mg/l, the weight of the dissolved hydroxyapatite was 0.9 mg, i.e., 0.9% of the sample. This is a considerable amount, which dissolved in just one day in the PBS. It is plausible that for each sample, the smallest particles underwent dissolution. In the further studies, we will examine the effect of storage in the solution on the nanostructure of the particles.
Further, an effect of the surface development on the pH and the conductivity of the solution was observed. The changes in the pH and conductivity ranged from ca. 3.24 to ca. 3.29 with an increase in the SSA value. Although the calcium-ion concentration stabilized after one day, the conductivity values stabilized after three days. However, the conductivity can hardly be correlated with calcium release only, as it depends on the overall composition of the solution, with a range of ions present.
Regarding the practical implications of this study for the development of nano-HAP as a material to enhance bone regeneration, there are two contradictory trends to be considered. On one hand, the larger the specific surface area, the greater the activity of the particles in the ion release and, possibly, in the biodegradation. On the other hand, the thermodynamic stability of these particles is limited, as a very high SSA value relates to high energy per unit weight or volume. This may limit the application of these materials because the shelf time would be short. It seems, from the present study, that the optimal SSA value is between 180 m2/g and 200 m2/g. For these values, the aspect ratio of the particles decreases to a stable level, so that the shape of the particles does not change in a significant way with time and possible further SSA decreases. On the other hand, the calcium-ion release remains at a high level. The MHS technology makes it possible to tune the particle size in this narrow gap of values.
Appropriate calcium-phosphate homeostasis is essential for normal bone function. In cases of large bone defects, insufficient calcium-ion release can contribute to a lack of or very slow bone-tissue regeneration. On the other hand, excessive calcium release can lead to undesirable tissue calcification. Therefore, it is necessary to find the optimal amount of calcium at which the calcium signal intensifies the induction of gene expression toward bone cells and thereby accelerates bone-tissue regeneration, which will be the subject of our future work.

5. Conclusions

Hydroxyapatite nanoparticles with the following average sizes were obtained with the use of the original method of microwave hydrothermal synthesis: 39 nm (51 m2/g), 29 nm (67 m2/g), 19 nm (108 m2/g), 11 nm (183 m2/g), 10 nm (211 m2/g), and 9 nm (240 m2/g). By varying the temperature and synthesis time, microwave hydrothermal synthesis makes it possible to precisely tune the specific surface area, shape, and density of hydroxyapatite nanoparticles, while keeping their chemical composition constant. A threshold specific surface area of 183 m2/g (11 nm) was found; above this threshold, the solubility of hydroxyapatite nanoparticles in phosphate-buffered saline increases significantly. Particles with optimal properties for application as bone-graft materials should have a specific surface area value in the range of 180–200 m2/g. The calcium release from the nanoparticles immersed in phosphate-buffered saline increased strongly above this specific surface area value. This effect can be exploited to produce bioactive hydroxyapatite. The nanoparticle size is therefore crucial when designing materials for bone-tissue regeneration.

Author Contributions

Conceptualization, U.S. and A.C.; methodology, U.S., A.C. and S.G.; formal analysis, U.S.; investigation, U.S., A.C. and S.G.; writing—original draft preparation, U.S. and J.W.; writing—review and editing, U.S., W.Ł. and J.W.; supervision, W.Ł. All authors have read and agreed to the published version of the manuscript.

Funding

The research studies were conducted on equipment funded by the project Center for Preclinical Research and Technology—CePT I (POIG.02.02.00-14-024/08), financed by the European Regional Development Fund “Innovative Economy” for years 2007–2013 and CePT II (RPMA.01.01.00-14-8476/17-04) from Regional Operational Programme of the Mazowieckie Voivodeship 2014–2020”. The present study results are the scope of the Ph.D Thesis of Urszula Szałaj.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Not applicable.

Acknowledgments

We would like to thank Kamil Sobczak (Faculty of Chemistry, Biological and Chemical Research Centre, University of Warsaw, Poland) for the TEM measurements.

Conflicts of Interest

The authors declare no conflict of interest.

References

  1. Gimigliano, F.; Resmini, G.; Moretti, A.; Aulicino, M.; Gargiulo, F.; Gimigliano, A.; Liguori, S.; Paoletta, M.; Iolascon, G. Epidemiology of Musculoskeletal Injuries in Adult Athletes: A Scoping Review. Medicina 2021, 57, 1118. [Google Scholar] [CrossRef] [PubMed]
  2. Sebbag, E.; Felten, R.; Sagez, F.; Sibilia, J.; Devilliers, H.; Arnaud, L. The world-wide burden of musculoskeletal diseases: A systematic analysis of the World Health Organization Burden of Diseases Database. Ann. Rheum. Dis. 2019, 78, 844–848. [Google Scholar] [CrossRef] [PubMed]
  3. Kanczler, J.M.; Wells, J.A.; Gibbs, D.M.R.; Marshall, K.M.; Tang, D.K.O.; Oreffo, R.O.C. Chapter 50-Bone Tissue Engineering and Bone Regeneration. In Principles of Tissue Engineering, 5th ed.; Lanza, R., Langer, R., Vacanti, J.P., Atala, A., Eds.; Academic Press: San Diego, CA, USA, 2020; pp. 917–935. ISBN 9780128184226. [Google Scholar] [CrossRef]
  4. Feng, X. Chemical and Biochemical Basis of Cell-Bone Matrix Interaction in Health and Disease. Curr. Chem. Biol. 2009, 3, 189–196. [Google Scholar] [CrossRef]
  5. Cheng, H.; Chabok, R.; Guan, X.; Chawla, A.; Li, Y.; Khademhosseini, A.; Jang, H.L. Synergistic interplay between the two major bone minerals, hydroxyapatite and whitlockite nanoparticles, for osteogenic dierentiation of mesenchymal stem cells. Acta Biomater. 2018, 69, 342–351. [Google Scholar] [CrossRef] [PubMed]
  6. Batool, S.; Liaqat, U.; Hussain, Z.; Sohail, M. Synthesis, Characterization and Process Optimization of Bone Whitlockite. Nanomaterials 2020, 10, 1856. [Google Scholar] [CrossRef] [PubMed]
  7. Morgan, E.F.; Barnes, G.L.; Einhorn, T.A. The bone organ system: Form and function. In Osteoporosis, 3rd ed.; Marcus, R., Feldman, D., Nelson, D.A., Rosen, C.J., Eds.; Academic Press: San Diego, CA, USA, 2007; pp. 3–25. ISBN 978-0-12-370544-0. [Google Scholar]
  8. Zhu, W.; Robey, P.G.; Boskey, A.L. The Regulatory Role of Matrix Proteins in Mineralization of Bone. In Osteoporosis, 3rd ed.; Marcus, R., Feldman, D., Nelson, D.A., Rosen, C.J., Eds.; Academic Press: San Diego, CA, USA, 2007; pp. 191–240. ISBN 978-0-12-370544-0. [Google Scholar]
  9. Dorozhkin, S.V. Bioceramics of calcium orthophosphates. Biomaterials 2010, 31, 1465–1485. [Google Scholar] [CrossRef]
  10. Hench, L.L. Bioceramics. J. Am. Ceram. Soc. 1998, 81, 1705–1728. [Google Scholar] [CrossRef]
  11. Suchanek, W.; Yoshimura, M. Processing and properties of hydroxyapatite-based biomaterials for use as hard tissue replacement implants. J. Mater. Res. 1998, 13, 94–117. [Google Scholar] [CrossRef]
  12. Ojeda, E.; García-Barrientos, Á.; Martínez de Cestafe, N.; Alonso, J.M.; Pérez-González, R.; Sáez-Martínez, V. Nanometric Hydroxyapatite Particles as Active Ingredient for Bioinks: A Review. Macromol 2022, 2, 20–29. [Google Scholar] [CrossRef]
  13. Kuśnieruk, S.; Wojnarowicz, S.; Chodara, A.; Chudoba, T.; Gierlotka, S.; Lojkowski, W. Influence of hydrothermal synthesis parameters on the properties of hydroxyapatite nanoparticles. Beilstein J. Nanotechnol. 2016, 7, 1586–1601. [Google Scholar] [CrossRef]
  14. Ielo, I.; Calabrese, G.; De Luca, G.; Conoci, S. Recent Advances in Hydroxyapatite-Based Biocomposites for Bone Tissue Regeneration in Orthopedics. Int. J. Mol. Sci. 2022, 23, 9721. [Google Scholar] [CrossRef] [PubMed]
  15. Rogowska-Tylman, J.; Locs, J.; Salma, I.; Woźniak, B.; Pilmane, M.; Zalite, V.; Wojnarowicz, J.; Kędzierska-Sar, A.; Chudoba, T.; Szlązak, K.; et al. In Vivo and in Vitro Study of a Novel Nanohydroxyapatite Sonocoated Scaffolds for Enhanced Bone Regeneration. Mater. Sci. Eng. C 2019, 99, 669–684. [Google Scholar] [CrossRef] [PubMed]
  16. Cavalcante, M.D.; de Menezes, L.R.; Rodrigues, E.J.D.; Tavares, M.I.B. In vitro characterization of a biocompatible composite based on poly (3-hydroxybutyrate)/hydroxyapatite nanoparticles as a potential scaffold for tissue engineering. J. Mech. Behav. Biomed. 2022, 128, 105138. [Google Scholar] [CrossRef] [PubMed]
  17. Kavasi, R.-M.; Coelho, C.C.; Platania, V.; Quadros, P.A.; Chatzinikolaidou, M. In Vitro Biocompatibility Assessment of Nano-Hydroxyapatite. Nanomaterials 2021, 11, 1152. [Google Scholar] [CrossRef]
  18. Cintra, C.C.V.; Ferreira-Ermita, D.A.C.; Loures, F.H.; Araújo, P.M.A.G.; Ribeiro, I.M.; Araújo, F.R.; Valente, F.L.; Reis, E.C.C.; Costa, A.C.F.M.; Bicalho, S.M.C.M.; et al. In vitro characterization of hydroxyapatite and cobalt ferrite nanoparticles compounds and their biocompatibility in vivo. J. Mater. Sci. Mater. Med. 2022, 33, 21. [Google Scholar] [CrossRef]
  19. Biazar, E.; Daliri, J.M.; Heidari, S.K.; Navayee, A.D.; Kamalvand, M.; Sahebalzamani, M.; Royanian, F.; Shabankhah, M.; Farajpour, F.L. Characterization and biocompatibility of hydroxyapatite nanoparticles extracted from fish bone. J. Bioeng. Res. 2020, 2, 10–19. [Google Scholar] [CrossRef]
  20. Turon, P.; Del Valle, L.J.; Alemán, C.; Puiggalí, J. Biodegradable and Biocompatible Systems Based on Hydroxyapatite Nanoparticles. Appl. Sci. 2017, 7, 60. [Google Scholar] [CrossRef]
  21. Torres, E.C.L.; De Sousa, E.M.B.; Cipreste, M.F. A Brief Review on Hydroxyapatite Nanoparticles Interactions with Biological Constituents. J. Biomater. Nanobiotechnol. 2022, 13, 24–44. [Google Scholar] [CrossRef]
  22. Borkowski, L.; Jojczuk, M.; Belcarz, A.; Pawlowska-Olszewska, M.; Kruk-Bachonko, J.; Radzki, R.; Bienko, M.; Slowik, T.; Lübek, T.; Nogalski, A.; et al. Comparing the Healing Abilities of Fluorapatite and Hydroxyapatite Ceramics in Regenerating Bone Tissue: An In Vivo Study. Materials 2023, 16, 5992. [Google Scholar] [CrossRef]
  23. Nalesso, P.R.L.; Vedovatto, M.; Gregório, J.E.S.; Huang, B.; Vyas, C.; Santamaria-Jr, M.; Bártolo, P.; Caetano, G.F. Early In Vivo Osteogenic and Inflammatory Response of 3D Printed Polycaprolactone/Carbon Nanotube/Hydroxyapatite/Tricalcium Phosphate Composite Scaffolds. Polymers 2023, 15, 2952. [Google Scholar] [CrossRef]
  24. George, S.M.; Nayak, C.; Singh, I.; Balani, K. Multifunctional Hydroxyapatite Composites for Orthopedic Applications: A Review. ACS Biomater. Sci. Eng. 2022, 8, 3162–3186. [Google Scholar] [CrossRef] [PubMed]
  25. Oliveira, H.L.; Da Rosa, W.L.O.; Cuevas-Suárez, C.E.; Carreño, N.L.V.; Da Silva, A.F.; Guim, T.N.; Dellagostin, O.A.; Piva, E. Histological Evaluation of Bone Repair with Hydroxyapatite: A Systematic Review. Calcif. Tissue. Int. 2017, 101, 341–354. [Google Scholar] [CrossRef] [PubMed]
  26. Bal, Z.; Kaito, T.; Korkusuz, F.; Yoshikawa, H. Bone regeneration with hydroxyapatite-based biomaterials. Emergent Mater. 2020, 3, 521–544. [Google Scholar] [CrossRef]
  27. Peacock, M. Calcium metabolism in health and disease. Clin. J. Am. Soc. Nephrol. 2010, 5 (Suppl. S1), S23–S30. [Google Scholar] [CrossRef]
  28. O’Neill, E.; Awale, G.; Daneshmandi, L.; Umerah, O.; Lo, K.W.-H. The roles of ions on bone regeneration. Drug Discov. Today 2018, 23, 879–890. [Google Scholar] [CrossRef] [PubMed]
  29. Liu, D.; Genetos, D.C.; Shao, Y.; Geist, D.J.; Li, J.; Ke, H.Z.; Turner, C.H.; Duncan, R.L. Activation of extracellular-signal regulated kinase (ERK 1/2) by fluid shear is Ca2+- and ATP-dependent in MC3T3-E1 osteoblasts. Bone 2008, 42, 644–652. [Google Scholar] [CrossRef] [PubMed]
  30. Danciu, T.E.; Adam, R.M.; Naruse, K.; Freeman, M.R.; Hauschka, P.V. Calcium regulates the PI3K-Akt pathway in stretched osteoblasts. FEBS Lett. 2003, 536, 193–197. [Google Scholar] [CrossRef]
  31. Foskett, J.K.; White, C.; Cheung, K.-H.; Mak, D.-O.D.; Lacruz, R.S.; Habelitz, S.; Wright, J.T.; Paine, M.L.; Hohendanner, F.; DeSantiago, J.; et al. Inositol trisphosphate receptor Ca2+ release channels. Physiol. Rev. 2007, 87, 593–658. [Google Scholar] [CrossRef]
  32. Lim, S.S.; Chai, C.Y.; Loh, H.-S. In vitro evaluation of osteoblast adhesion, proliferation and differentiation on chitosan-TiO2 nanotubes scaffolds with Ca2+ ions. Mater. Sci. Eng. C Mater. Biol. Appl. 2017, 76, 144–152. [Google Scholar] [CrossRef]
  33. Seol, Y.-J.; Park, J.Y.; Jung, J.W.; Jang, J.; Girdhari, R.; Kim, S.W.; Cho, D.-W. Improvement of Bone Regeneration Capability of Ceramic Scaffolds by Accelerated Release of Their Calcium Ions. Tissue. Eng. Part A 2014, 20, 2840–2849. [Google Scholar] [CrossRef]
  34. Sun, J.; Wang, F.; Sui, Y.; She, Z.; Zhai, W.; Wang, C.; Deng, Y. Effect of particle size on solubility, dissolution rate, and oral bioavailability: Evaluation using coenzyme Q10 as naked nanocrystals. Int. J. Nanomed. 2012, 7, 5733–5744. [Google Scholar] [CrossRef]
  35. Saentho, A.; Wisawapipat, W.; Lawongsa, P.; Aramrak, S.; Prakongkep, N.; Wantana, K.; Christl, I. Speciation and pH- and particle size-dependent solubility of phosphorus in tropical sandy soils. Geoderma 2022, 408, 115590. [Google Scholar] [CrossRef]
  36. Jinno, J.; Kamada, N.; Miyake, M.; Yamada, K.; Mukai, T.; Odomi, M.; Toguchi, H.; Liversidge, G.G.; Higaki, K.; Kimura, T. Effect of particle size reduction on dissolution and oral absorption of a poorly water-soluble drug, cilostazol, in beagle dogs. J. Control Release 2006, 111, 56–64. [Google Scholar] [CrossRef] [PubMed]
  37. Babu, V.R.; Areefulla, S.; Mallikarjun, V. Solubility and dissolution enhancement: An overview. J. Pharm. Res. 2010, 3, 141–145. [Google Scholar]
  38. Dinh, H.T.T.; Tran, P.H.L.; Duan, W.; Lee, B.-J.; Tran, T.T.D. Nano-sized solid dispersions based on hydrophobic-hydrophilic conjugates for dissolution enhancement of poorly water-soluble drugs. Int. J. Pharm. 2017, 533, 93–98. [Google Scholar] [CrossRef] [PubMed]
  39. Tang, R.; Wanga, L.; Nancollas, G.H. Size-effects in the dissolution of hydroxyapatite: An understanding of biological demineralization. J. Mater. Chem. 2004, 14, 2341–2346. [Google Scholar] [CrossRef]
  40. Jassim, Z.E.; Rajab, N.A. Review on preparation, characterization, and pharmaceutical application of nanosuspension as an approach of solubility and dissolution enhancement. J. Pharm. Res. 2018, 12, 771–774. [Google Scholar]
  41. Dizaj, S.M.; Vazifehasl, Z.H.; Salatin, S.; Adibkia, K.H.; Javadzadeh, Y. Nanosizing of drugs: Effect on dissolution rate. Res. Pharm. Sci. 2015, 10, 95–108. [Google Scholar]
  42. Hecq, J.; Deleers, M.; Fanara, D.; Vranckx, H.; Amighi, K. Preparation and characterization of nanocrystals for solubility and dissolution rate enhancement of nifedipine. Int. J. Pharm. 2005, 299, 167–177. [Google Scholar] [CrossRef]
  43. Smoleń, D.; Chudoba, T.; Gierlotka, S.; Kędzierska, A.; Łojkowski, W.; Sobczak, K.; Święszkowski, W.; Kurzydowski, K.J. Hydroxyapatite Nanopowder Synthesis with a Programmed Resorption Rate. J. Nanomater. 2012, 2012, 841971. [Google Scholar] [CrossRef]
  44. Wojnarowicz, J.; Chudoba, T.; Lojkowski, W. A Review of Microwave Synthesis of Zinc Oxide Nanomaterials: Reactants, Process Parameters and Morphologies. Nanomaterials 2020, 10, 1086. [Google Scholar] [CrossRef] [PubMed]
  45. Mohd Pu’ad, N.A.S.; Abdul Haq, R.H.; Mohd Noh, H.; Abdullah, H.Z.; Idris, M.I.; Lee, T.C. Synthesis method of hydroxyapatite: A review. Mater. Today Proc. 2020, 29, 233–239. [Google Scholar] [CrossRef]
  46. Gui, X.; Peng, W.; Xu, X.; Su, Z.; Liu, G.; Zhou, Z.; Liu, M.; Li, Z.; Song, G.; Zhou, C.; et al. Synthesis and application of nanometer hydroxyapatite in biomedicine. Nanotechnol. Rev. 2022, 11, 2154–2168. [Google Scholar] [CrossRef]
  47. DileepKumar, V.G.; Sridhar, M.S.; Aramwit, P.; Krut’ko, V.K.; Musskaya, O.N.; Glazov, I.E.; Reddy, N. A review on the synthesis and properties of hydroxyapatite for biomedical applications. J. Biomater. Sci. Polym. Ed. 2022, 33, 229–261. [Google Scholar] [CrossRef]
  48. Latocha, J.; Wojasinski, M.; Sobieszuk, P.; Ciach, T. Synthesis of hydroxyapatite in a continuous reactor: A review. Synthesis of hydroxyapatite in a continuous reactor: A review. Chem. Process Eng. 2018, 39, 281–293. [Google Scholar] [CrossRef]
  49. Baskaran, T.; Mohammad, N.F.; Md Saleh, S.S.; Mohd Nasir, N.F.; Mohd Daud, F.D. Synthesis Methods of Doped Hydroxyapatite: A Brief Review. J. Phys. Conf. Ser. 2021, 2071, 012008. [Google Scholar] [CrossRef]
  50. Haider, A.; Haider, S.; Han, S.S.; Kang, I.-K. Recent advances in the synthesis, functionalization and biomedical applications of hydroxyapatite: A review. RSC Adv. 2017, 7, 7442. [Google Scholar] [CrossRef]
  51. Alorku, K.; Manoj, M.; Yuan, A. A plant-mediated synthesis of nanostructured hydroxyapatite for biomedical applications: A review. RSC Adv. 2020, 10, 40923. [Google Scholar] [CrossRef]
  52. Ferraz, M.P.; Monteiro, F.J.; Manuel, C.M. Hydroxyapatite nanoparticles: A review of preparation methodologies. J. Appl. Biomater. Biomech. 2004, 2, 74–80. [Google Scholar]
  53. Munir, M.U.; Salman, S.; Ihsan, A.; Elsaman, T. Synthesis, Characterization, Functionalization and Bio-Applications of Hydroxyapatite Nanomaterials: An Overview. Int. J. Nanomed. 2022, 17, 1903–1925. [Google Scholar] [CrossRef]
  54. Burdusel, A.-C.; Neacsu, I.A.; Birca, A.C.; Chircov, C.; Grumezescu, A.-M.; Holban, A.M.; Curutiu, C.; Ditu, L.M.; Stan, M.; Andronescu, E. Microwave-Assisted Hydrothermal Treatment of Multifunctional Substituted Hydroxyapatite with Prospective Applications in Bone Regeneration. J. Funct. Biomater. 2023, 14, 378. [Google Scholar] [CrossRef] [PubMed]
  55. Koutsopoulos, S. Synthesis and characterization of hydroxyapatite crystals: A review study on the analytical methods. J. Biomed. Mater. Res. 2002, 62, 600–612. [Google Scholar] [CrossRef] [PubMed]
  56. Gani, M.A.; Budiatin, A.S.; Lestari, M.L.A.D.; Rantam, F.A.; Ardianto, C.; Khotib, J. Fabrication and Characterization of Submicron-Scale Bovine Hydroxyapatite: A Top-Down Approach for a Natural Biomaterial. Materials 2022, 15, 2324. [Google Scholar] [CrossRef] [PubMed]
  57. Qi, M.-L.; He, K.; Huang, Z.-N.; Shahbazian-Yassar, R.; Xiao, G.-Y.; Lu, Y.-P.; Shokuhfar, T. Hydroxyapatite Fibers: A Review of Synthesis Methods. JOM 2017, 69, 1354–1360. [Google Scholar] [CrossRef]
  58. Shavandi, A.; Bekhit, A.E.-D.A.; Sun, Z.F.; Ali, A. A Review of Synthesis Methods, Properties and Use of Hydroxyapatite as a Substitute of Bone. J. Biomim. Biomater. Biomed. Eng. 2015, 25, 98–117. [Google Scholar] [CrossRef]
  59. Firdaus Hussin, M.S.; Abdullah, H.Z.; Idris, M.I.; Abdul Wahap, M.A. Extraction of natural hydroxyapatite for biomedical applications—A review. Heliyon 2022, 8, e10356. [Google Scholar] [CrossRef]
  60. Radulescu, D.-E.; Vasile, O.R.; Andronescu, E.; Ficai, A. Latest Research of Doped Hydroxyapatite for Bone Tissue Engineering. Int. J. Mol. Sci. 2023, 24, 13157. [Google Scholar] [CrossRef]
  61. Prasad, P.S.; Marupalli, B.C.G.; Das, S.; Das, K. Surfactant-assisted synthesis of hydroxyapatite particles: A comprehensive review. J. Mater. Sci. 2023, 58, 6076–6105. [Google Scholar] [CrossRef]
  62. Method for of Producing Nano-Plates of Synthetic Hydroxyapatite and Nanopowder Comprising a Synthetic Hydroxyapatite Nano-Plates. Patent, Exclusive Right Number-Pat.235292. Available online: https://api-ewyszukiwarka.pue.uprp.gov.pl/api/collection/2f1684e27bedc23db247cb2a29fe4b67 (accessed on 15 September 2023).
  63. Alves Cardoso, D.; Jansen, J.A.; Leeuwenburgh, S.C.G. Synthesis and application of nanostructured calcium phosphate ceramics for bone regeneration. J. Biomed. Mater. Res. Part. B 2012, 100B, 2316–2326. [Google Scholar] [CrossRef]
  64. Singh, J.; Chatha, S.S.; Singh, H. In vitro assessment of plasma-sprayed reinforced hydroxyapatite coatings deposited on Ti6Al4V alloy for bio-implant applications. J. Mater. Res. 2022, 37, 2623–2634. [Google Scholar] [CrossRef]
  65. Ritwik, A.; Saju, K.K.; Vengellur, A.; Saipriya, P.P. Development of thin-film hydroxyapatite coatings with an intermediate shellac layer produced by dip-coating process on Ti6Al4V implant materials. J. Coat. Technol. Res. 2022, 19, 597–605. [Google Scholar] [CrossRef]
  66. Kubicki, G.; Leshchynsky, V.; Elseddawy, A.; Wisniewska, M.; Maev, R.G.; Jakubowicz, J.; Sulej-Chojnacka, J. Microstructure and Properties of Hydroxyapatite Coatings Made by Aerosol Cold Spraying-Sintering Technology. Coatings 2022, 12, 535. [Google Scholar] [CrossRef]
  67. Higuchi, J.; Fortunato, G.; Woźniak, B.; Chodara, A.; Domaschke, S.; Męczyńska-Wielgosz, S.; Kruszewski, M.; Dommann, A.; Łojkowski, W. Polymer Membranes Sonocoated and Electrosprayed with Nano-Hydroxyapatite for Periodontal Tissues Regeneration. Nanomaterials 2019, 9, 1625. [Google Scholar] [CrossRef]
  68. Higuchi, J.; Klimek, K.; Wojnarowicz, J.; Opalińska, A.; Chodara, A.; Szalaj, U.; Dąbrowska, S.; Fudala, D.; Ginalska, G. Electrospun Membrane Surface Modification by Sonocoating with HA and ZnO:Ag Nanoparticles-Characterization and Evaluation of Osteoblasts and Bacterial Cell Behavior In Vitro. Cells 2022, 11, 1582. [Google Scholar] [CrossRef] [PubMed]
  69. Majcher, A.; Wiejak, J.; Przybylski, J.; Chudoba, T.; Wojnarowicz, J. A novel reactor for microwave hydrothermal scale-up nanopowder synthesis. Int. J. Chem. React. Eng. 2013, 11, 361–368. [Google Scholar] [CrossRef]
  70. Holzwarth, U.; Gibson, N. The Scherrer equation versus the ‘Debye-Scherrer equation’. Nat. Nanotechnol. 2011, 6, 534. [Google Scholar] [CrossRef]
  71. ISO 9277; 2010-Determination of the Specific Surface Area of Solids by Gas Adsorption—BET Method. ISO: Geneva, Switzerland, 2023. Available online: https://www.iso.org/standard/44941.html (accessed on 24 August 2023).
  72. ISO 12154; 2014-Determination of Density by Volumetric Displacement—Skeleton Density by Gas Pycnometry. ISO: Geneva, Switzerland, 2023. Available online: https://www.iso.org/standard/54731.html (accessed on 24 August 2023).
  73. Szałaj, U.; Świderska-Środa, A.; Chodara, A.; Gierlotka, S.; Łojkowski, W. Nanoparticle Size Effect on Water Vapour Adsorption by Hydroxyapatite. Nanomaterials 2019, 9, 1005. [Google Scholar] [CrossRef]
  74. Polish Center for Accreditation, Testing Laboratories. Accreditation Number: AB 1503. Available online: https://www.pca.gov.pl/en/accredited-organizations/accredited-organizations/testing-laboratories/AB%201503,entity.html (accessed on 24 August 2023).
  75. PN-EN ISO/IEC 17025; 2018-02-Polish Version-General Requirements for the Competence of Testing and Calibration Laboratories. ISO: Geneva, Switzerland, 2023. Available online: https://sklep.pkn.pl/pn-en-iso-iec-17025-2018-02p.html (accessed on 24 August 2023).
  76. Legeros, R.Z. Calcium phosphates in oral biology and medicine. Monogr. Oral. Sci. 1991, 15, 1–201. [Google Scholar]
  77. Legeros, R.Z. Biological and Synthetic Apatites in Hydroxyapatite and Related Materials, 1st ed.; Brown, P.W., Constantz, B., Eds.; CRC Press: Boca Raton, FL, USA, 1994; pp. 3–28. [Google Scholar]
  78. Kozerozhets, I.V.; Panasyuk, G.P.; Semenov, E.A.; Avdeeva, V.V.; Danchevskaya, M.N.; Simonenko, N.P.; Vasiliev, M.G.; Kozlova, L.O.; Ivakin, Y.I. Recrystallization of nanosized boehmite in an aqueous medium. Powder Technol. 2023, 413, 118030. [Google Scholar] [CrossRef]
  79. Vdoviaková, K.; Jenca, A.; Jenca, A., Jr.; Danko, J.; Kresáková, L.; Simaiová, V.; Reichel, P.; Rusnák, P.; Pribula, J.; Vrzgula, M.; et al. Regenerative Potential of Hydroxyapatite-Based Ceramic Biomaterial on Mandibular Cortical Bone: An In Vivo Study. Biomedicines 2023, 11, 877. [Google Scholar] [CrossRef]
  80. ISO 13175-3; 2012(en) Implants for Surgery—Calcium Phosphates—Part 3: Hydroxyapatite and Beta-Tricalcium Phosphate Bone Substitutes. ISO: Geneva, Switzerland, 2023. Available online: https://www.iso.org/obp/ui/#iso:std:iso:13175:-3:ed-1:v1:en (accessed on 28 August 2023).
  81. Opalinska, A.; Malka, I.; Dzwolak, W.; Chudoba, T.; Presz, A.; Lojkowski, W. Size-dependent density of zirconia nanoparticles. Beilstein J. Nanotechnol. 2015, 6, 27–35. [Google Scholar] [CrossRef]
  82. Wojnarowicz, J.; Opalinska, A.; Chudoba, T.; Gierlotka, S.; Mukhovskyi, R.; Pietrzykowska, E.; Sobczak, K.; Lojkowski, W. Effect of water content in ethylene glycol solvent on the size of ZnO nanoparticles prepared using microwave solvothermal synthesis. J. Nanomater. 2016, 2016, 2789871. [Google Scholar] [CrossRef]
  83. Kusiak-Nejman, E.; Wojnarowicz, J.; Morawski, A.W.; Narkiewicz, U.; Sobczak, K.; Gierlotka, S.; Lojkowski, W. Size-dependent effects of ZnO nanoparticles on the photocatalytic degradation of phenol in a water solution. Appl. Surf. Sci. 2021, 541, 148416. [Google Scholar] [CrossRef]
  84. Wojnarowicz, J.; Chudoba, T.; Gierlotka, S.; Sobczak, K.; Lojkowski, W. Size Control of Cobalt-Doped ZnO Nanoparticles Obtained in Microwave Solvothermal Synthesis. Crystals 2018, 8, 179. [Google Scholar] [CrossRef]
  85. Tamari, S.; Aguilar-Chavez, A. Optimum Design of Gas Pycnometers for Determining the Volume of Solid Particles. J. Test. Eval. 2005, 33, JTE12674. [Google Scholar] [CrossRef]
  86. Tamari, S.; Aguilar-Chávez, A. Optimum design of the variable-volume gas pycnometer for determining the volume of solid particles. Meas. Sci. Technol. 2004, 15, 1146–1152. [Google Scholar] [CrossRef]
  87. Wilson, R.M.; Elliott, J.C.; Dowker, S.E.P. Rietveld Refinement of the Crystallographic Structure of Human Dental Enamel Apatites. Am. Mineral. 1999, 84, 1406–1414. [Google Scholar] [CrossRef]
  88. Okazaki, M.; Taira, M.; Takahashi, J. Rietveld Analysis and Fourier Maps of Hydroxyapatite. Biomaterials 1997, 18, 795–799. [Google Scholar] [CrossRef]
  89. Ma, G.; Liu, X.Y. Hydroxyapatite: Hexagonal or Monoclinic? Cryst. Growth Des. 2009, 9, 2991–2994. [Google Scholar] [CrossRef]
  90. Adam, M.; Ganz, C.; Xu, W.; Sarajian, H.; Götz, W.; Gerber, T. In vivo and in vitro investigations of a nanostructured coating material–a preclinical study. Int. J. Nanomed. 2014, 9, 975–984. [Google Scholar] [CrossRef]
  91. Shekhawat, D.; Vauth, M.; Pezoldt, J. Size Dependent Properties of Reactive Materials. Inorganics 2022, 10, 56. [Google Scholar] [CrossRef]
  92. Durkan, C. Size Really Does Matter: The Nanotechnology Revolution, 1st ed.; World Scientific Publishing: Singapore, 2019; ISBN 978-1-78634-661-2. [Google Scholar] [CrossRef]
  93. Navrotsky, A. Energetics of nanoparticle oxides: Interplay between surface energy and polymorphism. Geochem. Trans. 2003, 4, 34–37. [Google Scholar] [CrossRef] [PubMed]
  94. Kimmel, G.; Zabicky, J. Stability, Instability, Metastability and Grain Size in Nanocrystalline Ceramic Oxide Systems. Solid State Phenom. 2008, 140, 29–36. [Google Scholar] [CrossRef]
  95. Fu, Q.; Xue, Y.; Cui, Z. Size- and shape-dependent surface thermodynamic properties of nanocrystals. J. Phys. Chem. Solids 2018, 116, 79–85. [Google Scholar] [CrossRef]
  96. Wang, L.; Nancollas, G.H. Calcium orthophosphates: Crystallization and dissolution. Chem. Rev. 2008, 108, 4628–4669. [Google Scholar] [CrossRef]
  97. Wang, L.; Lu, J.; Xu, F.; Zhang, F. Dynamics of crystallization and dissolution of calcium orthophosphates at the near-molecular level. Chin. Sci. Bull. 2011, 56, 713–721. [Google Scholar] [CrossRef]
  98. Porter, A.E.; Patel, N.; Skepper, J.N.; Best, S.M.; Bonfield, W. Comparison of in vivo dissolution processes in hydroxyapatite and silicon-substituted hydroxyapatite bioceramics. Biomaterials 2003, 24, 4609–4620. [Google Scholar] [CrossRef]
  99. Hoppe, A.; Guldal, N.S.; Boccaccini, A.R. A review of the biological response to ionic dissolution products from bioactive glasses and glass-ceramics. Biomaterials 2011, 32, 2757–2774. [Google Scholar] [CrossRef]
  100. Nikolenko, M.V.; Vasylenko, K.V.; Myrhorodska, V.D.; Kostyniuk, A.; Likozar, B. Synthesis of Calcium Orthophosphates by Chemical Precipitation in Aqueous Solutions: The Effect of the Acidity, Ca/P Molar Ratio, and Temperature on the Phase Composition and Solubility of Precipitates. Processes 2020, 8, 1009. [Google Scholar] [CrossRef]
  101. Dorozhkin, S.V. Dissolution mechanism of calcium apatites in acids: A review of literature. World J. Methodol. 2012, 2, 1–17. [Google Scholar] [CrossRef]
  102. Dorozhkin, S.V. Surface reactions of apatite dissolution. J. Colloid Interface Sci. 1997, 191, 489–497. [Google Scholar] [CrossRef] [PubMed]
  103. Gayathri, B.; Muthukumarasamy, N.; Velauthapillai, D.; Santhosh, S.B. Magnesium incorporated hydroxyapatite nanoparticles: Preparation, characterization, antibacterial and larvicidal activity. Arab. J. Chem. 2018, 11, 645–654. [Google Scholar] [CrossRef]
  104. Zhu, Y.; Huang, B.; Zhu, Z.; Liu, H.; Huang, Y.; Zhao, X.; Liang, M. Characterization, dissolution and solubility of the hydroxypyromorphite–hydroxyapatite solid solution [(PbxCa1−x)5(PO4)3OH] at 25 °C and pH 2–9. Geochem. Trans 2016, 17, 2–9. [Google Scholar] [CrossRef]
  105. Prakash, K.H.; Kumar, R.; Ooi, C.P.; Cheang, P.; Khor, K.A. Apparent Solubility of Hydroxyapatite in Aqueous Medium and Its Influence on the Morphology of Nanocrystallites with Precipitation Temperature. Langmuir 2006, 22, 11002–11008. [Google Scholar] [CrossRef] [PubMed]
  106. Larsen, M.J.; Jensen, S.J. The hydroxyapatite solubility product of human dental enamel as a function of pH in the range 4.6–7.6 at 20 °C. Arch. Oral Biol. 1989, 34, 957–961. [Google Scholar] [CrossRef]
  107. Chen, Z.-F.; Darvell, B.W.; Leung, V.W.-H. Hydroxyapatite solubility in simple inorganic solutions. Arch. Oral Biol. 2004, 49, 359–367. [Google Scholar] [CrossRef]
  108. Arcos, D.; Vallet-Regí, M. Substituted hydroxyapatite coatings of bone implants. J. Mater. Chem. B 2020, 8, 1781–1800. [Google Scholar] [CrossRef]
  109. Kuranov, G.; Mikhelson, K.; Puzyk, A. Solubility of Hydroxyapatite as a Function of Solution Composition (Experiment and Modeling). In Processes and Phenomena on the Boundary between Biogenic and Abiogenic Nature; Lecture Notes in Earth System Sciences; Frank-Kamenetskaya, O., Vlasov, D., Panova, E., Lessovaia, S., Eds.; Springer: Cham, Switzerland, 2020. [Google Scholar] [CrossRef]
  110. Kamieniak, J.; Kelly, P.J.; Banks, C.E.; Doyle, A.M. Mechanical, pH and Thermal Stability of Mesoporous Hydroxyapatite. J. Inorg. Organomet. Polym. 2018, 28, 84–91. [Google Scholar] [CrossRef]
  111. Bell, L.C.; Mika, H.; Kruger, B.J. Synthetic hydroxyapatite-solubility product and stoichiometry of dissolution. Arch. Oral Biol. 1978, 23, 329–336. [Google Scholar] [CrossRef]
  112. Bengtsson, Å.; Shchukarev, A.; Persson, P.; Sjöberg, S. A solubility and surface complexation study of a non-stoichiometric hydroxyapatite. Geochim. Cosmochim. Acta 2009, 73, 257–267. [Google Scholar] [CrossRef]
  113. Bloebaum, R.D.; Lundeen, G.A.; Bachus, K.N.; Ison, I.; Hofmann, A.A. Dissolution of particulate hydroxyapatite in a macrophage organelle model. J. Biomed. Mater. Res. 1998, 40, 104–114. [Google Scholar] [CrossRef]
  114. Wang, D.; Xie, Y.; Jaisi, D.P.; Jin, Y. Effects of low-molecular-weight organic acids on the dissolution of hydroxyapatite nanoparticles. Environ. Sci. Nano 2016, 3, 768–779. [Google Scholar] [CrossRef]
  115. Capanema, N.S.V.; Mansur, A.A.P.; Carvalho, S.M.; Silva, A.R.P.; Ciminelli, V.S.; Mansur, H.S. Niobium-Doped Hydroxyapatite Bioceramics: Synthesis, Characterization and In Vitro Cytocompatibility. Materials 2015, 8, 4191–4209. [Google Scholar] [CrossRef] [PubMed]
  116. Mocanu, A.; Cadar, O.; Frangopol, P.T.; Petean, I.; Tomoaia, G.; Paltinean, G.-A.; Racz, C.P.; Horovitz, O.; Tomoaia-Cotisel, M. Ion release from hydroxyapatite and substituted hydroxyapatites in different immersion liquids: In vitro experiments and theoretical modelling study. Ion release from hydroxyapatite and substituted hydroxyapatites in different immersion liquids: In vitro experiments and theoretical modelling study. R. Soc. Open Sci. 2020, 8, 201785. [Google Scholar] [CrossRef]
Figure 1. Pressure–time plots for the GoHAP synthesis of size-controlled nanoparticles (GoHAP Type 2–Type 6). Time is counted from the moment the magnetron was switched on and the start of power delivery.
Figure 1. Pressure–time plots for the GoHAP synthesis of size-controlled nanoparticles (GoHAP Type 2–Type 6). Time is counted from the moment the magnetron was switched on and the start of power delivery.
Materials 16 06397 g001
Figure 2. The histogram of the particle-size distribution of GoHAP samples (TEM method): (A) Type 1, (B) Type 2, (C) Type 3, (D) Type 4, (E) Type 5, (F) Type 6.
Figure 2. The histogram of the particle-size distribution of GoHAP samples (TEM method): (A) Type 1, (B) Type 2, (C) Type 3, (D) Type 4, (E) Type 5, (F) Type 6.
Materials 16 06397 g002
Figure 3. Density as a function of specific surface area (A), and mean particle size as a function of specific surface area (B). Experimental and calculated correlation of density as a function of mean particle size based on BET for GoHAP Type 1–Type 6.
Figure 3. Density as a function of specific surface area (A), and mean particle size as a function of specific surface area (B). Experimental and calculated correlation of density as a function of mean particle size based on BET for GoHAP Type 1–Type 6.
Materials 16 06397 g003
Figure 4. X-ray-diffraction-line profiles of GoHAP samples.
Figure 4. X-ray-diffraction-line profiles of GoHAP samples.
Materials 16 06397 g004
Figure 5. TEM images of GoHAP samples: (A1,A2) type 1, (B1,B2) type 2, (C1,C2) type 3. TEM (A1C1) TEM images obtained using bright-field imaging. (A2C2) TEM images obtained using dark-field imaging.
Figure 5. TEM images of GoHAP samples: (A1,A2) type 1, (B1,B2) type 2, (C1,C2) type 3. TEM (A1C1) TEM images obtained using bright-field imaging. (A2C2) TEM images obtained using dark-field imaging.
Materials 16 06397 g005
Figure 6. TEM images of GoHAP samples: (A1,A2) type 4, (B1,B2) type 5, (C1,C2) type 6. (A1C1) TEM images obtained using bright-field imaging. (A2C2) TEM images obtained using dark-field imaging.
Figure 6. TEM images of GoHAP samples: (A1,A2) type 4, (B1,B2) type 5, (C1,C2) type 6. (A1C1) TEM images obtained using bright-field imaging. (A2C2) TEM images obtained using dark-field imaging.
Materials 16 06397 g006
Figure 7. Concentrations of calcium ions released from GoHAP Type 1–Type 6.
Figure 7. Concentrations of calcium ions released from GoHAP Type 1–Type 6.
Materials 16 06397 g007
Figure 8. Conductivity of PBS filtrate during dissolution tests of GoHAP type 1–type 6.
Figure 8. Conductivity of PBS filtrate during dissolution tests of GoHAP type 1–type 6.
Materials 16 06397 g008
Figure 9. The PH of PBS filtrate during the degradation test of GoHAP type 1–type 6.
Figure 9. The PH of PBS filtrate during the degradation test of GoHAP type 1–type 6.
Materials 16 06397 g009
Figure 10. Correlation between the amount of calcium ions released and the specific surface area of GoHAP type 1–type 6.
Figure 10. Correlation between the amount of calcium ions released and the specific surface area of GoHAP type 1–type 6.
Materials 16 06397 g010
Figure 11. Illustration of the effect of the specific surface areas of hydroxyapatite nanoparticles on calcium-ion release. SSA—specific surface area; MPS—mean particle size.
Figure 11. Illustration of the effect of the specific surface areas of hydroxyapatite nanoparticles on calcium-ion release. SSA—specific surface area; MPS—mean particle size.
Materials 16 06397 g011
Table 1. Microwave-synthesis parameters.
Table 1. Microwave-synthesis parameters.
GoHAPHeating Time (s)Total Reaction Time (s)Pressure
(bar)
Temperature
(°C)
Type 1----
Type 255550.2115
Type 360900.3125
Type 41006003130
Type 512060010175
Type 6200120050260
Table 2. GoHAP characterization. Standard deviation is given for each value (±).
Table 2. GoHAP characterization. Standard deviation is given for each value (±).
GoHAPSpecific Surface Area,
as (m2/g)
Skeleton
Density,
ρs ± SD (g/cm3)
Type 12402.89 ± 0.01
Type 22112.90 ± 0.01
Type 31832.92 ± 0.01
Type 41082.98 ± 0.01
Type 5673.05 ± 0.01
Type 6513.05 ± 0.01
Table 3. Comparison of the GoHAP NP sizes with different methods. D(002)—crystallite size for crystal plane 002. D(300)—crystallite size for crystal plane 300. Standard deviation is given for each value (±). d—mean particle size (diameter); SD—standard deviation; SSA—specific surface area; TEM—transmission-electron microscopy.
Table 3. Comparison of the GoHAP NP sizes with different methods. D(002)—crystallite size for crystal plane 002. D(300)—crystallite size for crystal plane 300. Standard deviation is given for each value (±). d—mean particle size (diameter); SD—standard deviation; SSA—specific surface area; TEM—transmission-electron microscopy.
GoHAPTMMean Particle Size Based on TEM Method,
dTEM ± SD (nm)
Mean Particle Size Based on SSA,
dSSA ± SD (nm)
Mean Size of Crystallites Based on Scherrer’s Formula
Length
D(002) ± SD (nm)
Width
D(300) ± SD (nm)
Aspect Ratio
(D(002)/D(300))
Type 18 ± 49 ± 114 ± 75 ± 12.8
Type 213 ± 410 ± 121 ± 125 ± 24.2
Type 313 ± 611 ± 129 ± 1517 ± 71.7
Type 420 ± 919 ± 233 ± 1723 ± 81.4
Type 530 ± 1229 ± 343 ± 2027 ± 101.6
Type 636 ± 1339 ± 451 ± 2432 ± 111.6
Table 4. Calcium-to-phosphate molar ratio of GoHAP Type 1–Type 6. Data obtained from ICP-OES analyses.
Table 4. Calcium-to-phosphate molar ratio of GoHAP Type 1–Type 6. Data obtained from ICP-OES analyses.
GoHAPCalcium
(mol)
Phosphorus
(mol)
Calcium-Phosphorus (Ca/P) Ratio
Type 18.295.441.52
Type 28.095.361.51
Type 39.676.401.51
Type 48.715.741.52
Type 55.383.541.52
Type 69.296.091.53
Table 5. Impurity content of GoHAP type 1–type 6. Data obtained from ICP-OES analyses.
Table 5. Impurity content of GoHAP type 1–type 6. Data obtained from ICP-OES analyses.
GoHAPMagnesium
(wt%)
Silicon
(wt%)
Aluminum
(wt%)
Iron
(wt%)
Sodium
(wt%)
Sodium
(wt%)
Type 10.2250.0530.0190.0210.0820.0009
Type 20.2190.0560.0180.0180.0830.0009
Type 30.2580.0490.0190.0200.0740.010
Type 40.2320.0540.0190.0200.0870.010
Type 50.1440.0430.0130.0140.0730.006
Type 60.2520.0500.0190.0260.0830.010
Mean value0.222 ± 0.0410.051 ± 0.0050.018 ± 0.0020.020 ± 0.0040.080 ± 0.0060.006 ± 0.004
Disclaimer/Publisher’s Note: The statements, opinions and data contained in all publications are solely those of the individual author(s) and contributor(s) and not of MDPI and/or the editor(s). MDPI and/or the editor(s) disclaim responsibility for any injury to people or property resulting from any ideas, methods, instructions or products referred to in the content.

Share and Cite

MDPI and ACS Style

Szałaj, U.; Chodara, A.; Gierlotka, S.; Wojnarowicz, J.; Łojkowski, W. Enhanced Release of Calcium Ions from Hydroxyapatite Nanoparticles with an Increase in Their Specific Surface Area. Materials 2023, 16, 6397. https://doi.org/10.3390/ma16196397

AMA Style

Szałaj U, Chodara A, Gierlotka S, Wojnarowicz J, Łojkowski W. Enhanced Release of Calcium Ions from Hydroxyapatite Nanoparticles with an Increase in Their Specific Surface Area. Materials. 2023; 16(19):6397. https://doi.org/10.3390/ma16196397

Chicago/Turabian Style

Szałaj, Urszula, Agnieszka Chodara, Stanisław Gierlotka, Jacek Wojnarowicz, and Witold Łojkowski. 2023. "Enhanced Release of Calcium Ions from Hydroxyapatite Nanoparticles with an Increase in Their Specific Surface Area" Materials 16, no. 19: 6397. https://doi.org/10.3390/ma16196397

Note that from the first issue of 2016, this journal uses article numbers instead of page numbers. See further details here.

Article Metrics

Back to TopTop