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Study and Optimization of Oxygenated Apatite Obtained by Dissolution-Reprecipitation of Hydroxyapatite in a Solution of Hydrogen Peroxide

  • L. Naanaai
  • K. Azzaoui
  • A. Lamhamdi
  • E. Mejdoubi
  • M. Lakrat
  • S. JodehEmail author
Original Article
  • 92 Downloads

Abstract

The experimental design strategy is applied to study and model the preparation of oxygenated apatite obtained by the dissolution-reprecipitation of hydroxyapatite in a hydrogen peroxide. The oxygenated apatite obtained is characterized by infrared spectroscopy (IR), X-ray diffraction (XRD) and scanning electron microscopy (SEM). The characterization by analyzes (IR, DRX, SEM) shows that the dissolution-repricipitation method gives better results for oxygenated apatite containing molecular oxygen. For this, we studied the influence of the four factors such as the temperature of the reaction medium, the pH, the concentration of hydrogen peroxide and the mass of the attacked hydroxyapatite. As well as their interactions whose goal is to increase the molecular oxygen insertion rate in apatitic tunnel. The orthogonal centered composite plane allowed us to establish an equation that links the insertion rate according to the factors studied, and we obtained an optimum of introduction of 5.06%.

Keywords

Hydroxyapatite Dissolution-reprecipitation Oxygenated apatite Oxygen insertion rate Optimization Plans of experiments 

1 Introduction

Oxygenated phosphocalcic apatites have been used as bone substitutes and antiseptic carrier matrices for many decades. Oxygenated apatite (OA) belongs to the apatite family of the chemical formula Ca10(PO4)6(OH)2(O2). It is characterized by a structure containing hexagonal tunnels along the c-axis in which the inserted molecular oxygen acts as an antibacterial [1]. Indeed, they undergo, after implantation, a process of degradation and dissolution combined with bone neoformation and a progressive release of oxygenated species (O2, O22−), elements responsible for the antiseptic power [2, 3]. They can be used for filling dental canals [4].

Several researchers who worked on oxygenated apatite [5, 6], synthesized carbonated oxygenated apatite based on calcium phosphate from calcium carbonate (CaCO3) and orthophosphorique acid (H3PO4) by precipitation. Another synthetic process used for the preparation of oxygenated apatite from the hydrolysis of brushite in hydrogen peroxide [7]. M. Elgadi et al. [8] have proposed the reaction of phosphoric acid with calcium hydroxide or `with calcium carbonate in oxygenated water. They have inserted 2.48% in weight of oxygen into the apatitic tunnel. Lamhamdi et al. [9] modeled the synthesis of oxygenated apatite prepared by the hydrolysis of brushite in oxygenated medium, the rate of oxygen insertion in this study is 4.5%.

Vandecandelaere et al. studied the preparation of peroxide-doped calcium phosphate apatites—in view of potential uses as bioactive bioceramics with antimicrobial functions, and on their main physico-chemical characteristics [10].

Global, apatite-bearing phosphorites represent unique biogeochemical periods coincident with major transitions in biological evolution and, particularly, marine oxygenation. However, current understanding of such oxygenation is limited by qualitative and not uncommonly contrasting interpretations of the marine redox conditions evidenced by sedimentary isotope and trace element abundances [11].

The hydrolysis of the brushite is very slow and the synthesis by double decomposition exhibits a slightly carbonated apatite: these carbonates can prevent the insertion of oxygen into the apatitic tunnels. The synthesis of oxygenated apatite by the method of dissolution-reprecipitation of hydroxyapatite, has advantages in comparison with other methods of synthesis, but this method is poorly studied.

The objective of this work is to develop the dissolution-reprecipitation method by using the strategy of the experience plans for that, we have synthesized oxygenated apatite by the method of dissolution-reprecipitation of hydroxyapatite in a solution of hydrogen peroxide. The product obtained is characterized by the different techniques: infrared spectroscopy (IR), X-ray diffraction (XRD) and scanning electron microscopy (SEM). Then we did the modeling by the strategy of experience plans by studying the effects of temperature T, the pH of the reaction medium, the concentration of hydrogen peroxide and the mass of hydroxyapatite on the rate of insertion of oxygen into the apatite tunnels.

2 Experimental Methods

2.1 Preparation of Oxygenated Apatite

The synthesis of oxygenated apatite is made by several methods [5, 7, 12, 13, 14]. First, the hydroxyapatite was prepared by the double decomposition method by mixing a calcium salt [Ca (NO3)2·4H2O (99%)] and a phosphate salt [(NH4)2·HPO4 (99%)] in a ratio Ca/P = 1.67. All reagents and solvents used in this work have been purchased from Aldrich and used without further purification. We used the dissolution-reprecipitation method. This method consists in attacking the hydroxyapatite powder with hydrogen peroxide according to the following reaction:
$$ \text{Ca}_{10} \left({\rm PO}_{4}\right)_{6}\left({\rm OH}\right)_{2} \xrightarrow{ \begin{subarray}{ll} 1) {\rm HCl}\\ 2) {{\rm H}_{2}{\rm O}_{2}}\\3) {\rm NH}_{4}{\rm OH}\\ \end{subarray}} {\rm Ca}_{\rm 10} \left({\rm PO}_{4} \right)_{6} \left({\rm OH} \right)_{2} {\rm O}_{2} $$
The structural approach of the studied product is carried out by Fourier transform infrared spectroscopy (FTIR). The transmission spectra of the glasses were recorded in a frequency range of 400–4000 cm−1 at room temperature. Sample pellets were prepared by mixing and grinding a small amount of the product powder with spectroscopically grade dry powder KBr and then compressing the mixtures to form thin pellets for analysis as shown in (Tables 1, 2).
Table 1

List of factors for experiments

  

Levels

 

Coded variables Xi

X1, X2, X3, X4

− 2

− 1

0

+ 1

+ 2

Δxi

Natural variables xi

x1 = m(g)

1

1,5

2

2.5

3

0,5

x2 = pH

9

9.5

10

10.5

11

0,5

x3 = % H202

5

10

15

20

25

5

x4 = T °C

25

35

45

55

65

10

Table 2

Matrix of experiments and experimental results

m(g)

pH

% H2O2

T °C

% O2

X1

x1

X2

x2

X3

x3

X4

x4

1

− 1

1.5

− 1

9.5

− 1

10

− 1

35

3.79

2

+ 1

2.5

− 1

9.5

− 1

10

− 1

35

3.54

3

− 1

1.5

+ 1

10.5

− 1

10

− 1

35

3.31

4

− 1

1.5

− 1

9.5

+ 1

20

− 1

35

2.78

5

− 1

1.5

− 1

9.5

− 1

10

+ 1

55

4.3

6

+ 1

2.5

+ 1

10.5

− 1

10

− 1

35

2.53

7

+ 1

2.5

− 1

9.5

+ 1

20

− 1

35

2.78

8

+ 1

2.5

− 1

9.5

− 1

10

1

55

4.93

9

− 1

1.5

+ 1

10.5

+ 1

20

− 1

35

4.55

10

− 1

1.5

+ 1

10.5

− 1

10

+ 1

55

4.65

11

− 1

1.5

− 1

9.5

+ 1

20

+ 1

55

3.03

12

+ 1

2.5

+ 1

10.5

+ 1

20

− 1

35

2.9

13

+ 1

2.5

+ 1

10.5

− 1

10

+ 1

55

3.59

14

+ 1

2.5

− 1

9.5

+ 1

20

+ 1

55

3.61

15

− 1

1.5

+ 1

10.5

+ 1

20

+ 1

55

3.41

16

+ 1

2.5

+ 1

10.5

+ 1

20

+ 1

55

3.39

17

0

2

0

10

0

15

0

45

3.16

18

0

2

0

10

0

15

0

45

3.16

19

0

2

0

10

0

15

0

45

3.16

20

0

2

0

10

0

15

0

45

3.16

21

0

2

0

10

0

15

0

45

3.16

22

0

2

0

10

0

15

0

45

3.16

23

0

2

0

10

0

15

− 2

25

2.27

24

0

2

0

10

0

15

+ 2

65

3.66

25

0

2

0

10

− 2

5

0

45

3.23

26

0

2

0

10

+ 2

25

0

45

2.02

27

0

2

0

9

0

15

0

45

4.04

28

0

2

+ 2

11

0

15

0

45

3.92

29

− 2

1

0

10

0

15

0

45

5.06

30

+ 2

3

0

10

0

15

0

45

4.19

The X-ray powder diffraction patterns of the above product were recorded using Cu-Ka radiation. The material to be analyzed is finely ground, homogenized and the mass composition is determined. Therefore, it is also called the diffraction method of the powder.

Scanning Electron Microscopy (SEM) is an electron microscopy technique capable of producing high resolution images of the surface of a sample using the principle of electron-matter interactions. Sample preparation is binding. They must be dehydrated and then undergo treatment to become a driver. The sample is then placed on the slide.

2.2 Experimental Study

This study aims to represent the synthesis of oxygenated apatite from the so-called dissolution-reprecipitation method of hydroxyapatite in the presence of hydrogen peroxide. To determine the optimal conditions for the synthesis of oxygenated apatite. We used the experimental design method. Using the orthogonal composite plane, a descriptive mathematical model has been developed which expresses the response (%O2) as a function of temperature T, pH, % H2O2 and the mass of hydroxyapatite. The equation of the model is then written, according to the four factors, in the following form:
$$ \begin{aligned} {\text{Y}} =& \, \% {\text{ O}}_{ 2} = {\text{ a}}_{0} + {\text{ a}}_{ 1} *{\mathbf{X}}_{{\mathbf{1}}} + {\text{ a}}_{ 2} *{\mathbf{X}}_{{\mathbf{2}}} \\ & + {\text{ a}}_{ 3} *{\mathbf{X}}_{{\mathbf{3}}} + {\text{ a}}_{ 4} *{\mathbf{X}}_{{\mathbf{4}}} \\ & + {\text{ a}}_{ 1 1} * \, ({\mathbf{X}}_{{\mathbf{1}}} )^{{\mathbf{2}}} + {\text{ a}}_{ 2 2} * \, ({\mathbf{X}}_{{\mathbf{2}}} )^{{\mathbf{2}}} \\ & + {\text{ a}}_{ 3 3} * \, ({\mathbf{X}}_{{\mathbf{3}}} )^{{\mathbf{2}}} + {\text{ a}}_{ 4 4} * \, \left( {{\mathbf{X}}_{{\mathbf{4}}} } \right)^{{\mathbf{2}}} \\ & + {\text{ a}}_{ 1 2} * \, \left( {{\mathbf{X}}_{{\mathbf{1}}} *{\mathbf{X}}_{{\mathbf{2}}} } \right) \, + {\text{ a}}_{ 1 3} * \, \left( {{\mathbf{X}}_{{\mathbf{1}}} *{\mathbf{X}}_{{\mathbf{3}}} } \right) \\ & + {\text{ a}}_{ 1 4} *\left( {{\mathbf{X}}_{{\mathbf{1}}} * \, {\mathbf{X}}_{{\mathbf{4}}} } \right) \, + {\text{a}}_{ 2 3} * \, \left( {{\mathbf{X}}_{{\mathbf{2}}} *{\mathbf{X}}_{{\mathbf{3}}} } \right) \, \\ & + {\text{ a}}_{ 2 4} * \, ({\mathbf{X}}_{{\mathbf{2}}} * \, {\mathbf{X}}_{{\mathbf{4}}} ) + {\text{ a}}_{ 3 4} * \, ({\mathbf{X}}_{{\mathbf{3}}} * \, {\mathbf{X}}_{{\mathbf{4}}} ) \\ \end{aligned} $$
With:
  • Y: response function (% O2)

  • ai, aii and aij: the coefficients of the mathematical model.

The coefficients of the mathematical model are determined from the JMP software, as well as all the statistical analyzes and graphs have been made using this software. We performed 30 experimental trials, including 6 central points using the experimental design of the orthogonal centered composite plan strategy. In each experiment, the levels of the selected factors were varied simultaneously. The concentration is determined by attacking oxygenated apatite with perchloric acid, which destroys the apatite network. The release of oxygen and carbon dioxide is done by adding perchloric acid to the powder. Carbon dioxide is trapped thanks to a solution of soda. The volume of oxygen is directly measured by means of a graduated tube filled with mercury as shown in (Table 3).
Table 3

Analysis of variance

Source

Degrees of freedom

Somme of squares

Middle square

Rapport F

Model

14

14.871238

1.06223

19.3812

Residual

15

0.822108

0.05481

Prob > F

Total

29

15.693347

 

0.0001*

*Significant

3 Results and Discussions

Figure 1 shows the FTIR infrared absorption spectrum of the oxygenated apatite synthesized by the dissolution-reprecipitation method of hydroxyapatite, calcined at 300 °C. Indeed, the spectrum is dominated by the typical PO4 bands: asymmetric mode at 1046–1087 cm−1, symmetric stretching mode at 962 cm−1 and bending mode at 600–650 cm−1, the band at 3465 cm−1 corresponds to the trace of water incorporated to the structure, wide band around 1610 cm−1 can be attributed to bending mode (OH), Carbonate bands (CO32−) which are characteristic of air contamination are also observed at 1420–1465 cm−1 [15, 16]. These bands indicate that the synthesized AO is in a weakly crystalline apatite form and contains a small amount of carbonates.
Fig. 1

Infrared spectrum of oxygenated apatite, obtained by dissolution-reprecipitation of hydroxyapatite calcined at 300 °C

The X-ray diffraction patterns (Fig. 2) represent the calcined AO at 300 °C. It appears from this diagram (wide and diffuse lines) that the synthesized product has evolved into a poorly crystallized apatite pure phase.
Fig. 2

X-ray diffraction spectrum of the AO calcined at 300 °C

The morphological observation of the prepared apatite was performed by scanning electron microscopy (SEM). To minimize load effects and achieve high resolution, we worked under low voltage (20 kV). Before being analyzed, the powders are deposited on an adhesive patch of carbon and metallic gold. The images obtained make it possible to visualize the morphology and the distribution of the grains (Fig. 3). We note that the particle size of apatite is of the micrometric type.
Fig. 3

Scanning electron microscopy of oxygenated apatite heated to 300 °C

3.1 Statistical Analyzes and Interpretations

3.1.1 Analysis of the Variance

The value of the experimental factor is (Fexp = 19.3812) is greater than that of the theoretical factor (F0.01 (14; 15) = 3.563). The value of the theoretical factor is obtained using the Fisher-Snedecor table. With Fα (v1, v2) is the function of Snedecor at v1 and v2 degrees of freedom for a probability of α. In our case (α = 0.01, v1 = 14, v2 = 15). The analysis of the variance allowed us to conclude that the changes in the responses of the inserted oxygen are due to variations of different factors.

Table 4 summarizes the values of the coefficients of the model as well as the corresponding statistical data. The results obtained in the table above show that the coefficients of the model a2, a44, a13, and a24 have insignificant values.
Table 4

Model coefficient estimates

Coefficients of model

Estimation

Degrees of freedom

Sum of squares

Rapport F

Prob. > F

a0

3.16

a1

− 0.1787

1

0.7668375

13.9915

0.0020*

a2

− 0.0279

1

0.0187042

0.3413

0.5678

a3

− 0.2754

1

1.8205042

33.2165

< 0.0001*

a4

0.3129

1

2.3500042

42.8776

< 0.0001*

a12

− 0.2793

1

1.2488063

22.7854

0.0002*

a13

0.0231

1

0.0085563

0.1561

0.6983

a23

0.2831

1

1.2825563

23.4012

0.0002*

a14

0.1756

1

0.4935063

9.0044

0.0090*

a24

− 0.0768

1

0.0945562

1.7253

0.2088

a34

− 0.2418

1

0.9360562

17.0791

0.0009*

a11

0.36947

1

3.7444074

68.3196

< 0.0001*

a22

0.20822

1

1.1892860

21.6994

0.0003*

a33

− 0.1305

1

0.4672646

8.5256

0.0106*

a44

− 0.0455

1

0.0568360

1.0370

0.3247

*Significant

The model equation for the oxygen insertion rate is written, taking into account the significance level of 1%.
$$ \begin{aligned} {\text{Y}} =& \, \% {\text{ O}}_{ 2} = { 3} . 1 6 { } - \, 0. 1 7 8 7 { }*{\mathbf{X}}_{{\mathbf{1}}} - \, 0. 2 7 5 4 { }*{\mathbf{X}}_{{\mathbf{3}}} \\ & + \, 0. 3 1 2 9*{\mathbf{X}}_{{\mathbf{4}}} + \, 0. 3 6 9 4 { }* \, ({\mathbf{X}}_{{\mathbf{1}}} )^{{\mathbf{2}}} \\ & + \, 0. 20 8 2* \, ({\mathbf{X}}_{{\mathbf{2}}} )^{{\mathbf{2}}} - 0. 1 30 5*\left( {{\mathbf{X}}_{{\mathbf{3}}} } \right)^{{\mathbf{2}}} \\ & - \, 0. 2 7 9 3* \, \left( {{\mathbf{X}}_{{\mathbf{1}}} *{\mathbf{X}}_{{\mathbf{2}}} } \right) \, + \, 0. 1 7 5 6 { }*\left( {{\mathbf{X}}_{{\mathbf{1}}} * \, {\mathbf{X}}_{{\mathbf{4}}} } \right) \, \\ & + 0. 2 8 3 1 { }* \, \left( {{\mathbf{X}}_{{\mathbf{2}}} *{\mathbf{X}}_{{\mathbf{3}}} } \right) - \, 0. 2 4 1 8* \, ({\mathbf{X}}_{{\mathbf{3}}} * \, {\mathbf{X}}_{{\mathbf{4}}} ) \\ \end{aligned} $$
From Fig. 4 (the effect curves), it can be said that the mass of the powder attacked by the acid, has a negative effect at the lower level of the center of interest. On the other hand, at the higher level of the center of interest, it has a positive effect on the oxygen insertion rate. This is due to mass interactions with other factors, such as the temperature and concentration of oxygenated water. At a fixed value of the H2O2 concentration, the oxygen insertion rate will be depleted with the increase of the attacked mass. The increase in temperature favors the precipitation of apatite for high mass values, therefore the insertion rate increases.
Fig. 4

Effect Curves

The pH effect curve shows that the latter has no effect on the oxygen level. This is explained by the range of pH variations, since we are still in a largely basic environment (stability domain of apatite). The effect curve of the concentration of hydrogen peroxide shows that it has negative effects on the oxygen insertion rate, when going from a low level to a high level. This negative effect is due to the deoxygenating of hydrogen peroxide in basic medium, for fairly high values of the pH and the concentration of hydrogen peroxide. Therefore the oxygen insertion is weak. The effect of temperature on the oxygen insertion rate increases as the temperature goes from a low level to a higher level. Indeed, the apatitic structure further traps the oxygen molecules for relatively high temperatures. But this in the experimental field we have previously chosen.

Figure 5 shows the Pareto diagram, which shows the importance of the different factors on the oxygen insertion rate. The interaction between the mass of the attacked hydroxyapatite with itself is the most influential factor on the oxygen insertion rate in the apatitic tunnels. This means that the mass of the hydroxyapatite used only starts to have positive effects at high mass values. This could be explained by the fact that the germination effect takes place only at fairly high hydroxyapatite masses. On the other hand, at low values of the hydroxyapatite mass used, the apatite formed would pass through an intermediate phase (probably octocalcium phosphate) which reduces the insertion of oxygen into the apatite network.
Fig. 5

Pareto diagram

According to the interaction curves (Fig. 6), the insignificant interactions are: the interaction between m(g) and % H2O2 and the interaction between the pH and the temperature of the reaction medium. The significant interactions are: the interaction between the mass (m) of apatite and the concentration of hydrogen peroxide (% H2O2) on the one hand and the pH and T (°C) factors on the other hand. Then we can consider that the insignificant interactions are negligible as shown in (Fig. 7).
Fig. 6

Interaction curves

Fig. 7

Response area at T = 45 °C and % H2O2 = 10

The study of these significant interactions shows that the optimal conditions will be obtained with:
  • at the lower level of interest for the m * pH interaction

  • at higher level of interest for pH *% H2O2 interaction

  • m (g) at level − 1 and T at level + 1 for m * T interaction

  • % H2O2 at level − 1 and T at level + 1 for %H2O2 * T interaction.

The analysis of the isoresponses curves (Fig. 8) shows that the insertion optimum of O2, is obtained under the following conditions: the region of the pH 9–11, the small masses of hydroxyapatite attacked, the small concentration of the hydrogen peroxide and with increasing temperature. Indeed, the increase in temperature implies an increase in the insertion rate for low values of hydroxyapatite. Therefore, the decrease in mass reinforces the positive effect of temperature. According to the isoréponses curves one can simply find the optimum of insertion in the apatitic tunnels of phosphocalcic hydroxyapatite, according to the different factors and the variations of each factor. Thus, these presentations offer wide practical applications in dentistry without the need for experimentation. The confirmatory tests were performed and we obtained % O2 equal to = 5.06% by weight for the optimal operating conditions.
Fig. 8

isoresponse plot of %O2

4 Conclusion

The synthesis of oxygenated apatite by the dissolution-reprecipitation method of hydroxyapatite is poorly studied. For this, we have deepened the study using the strategy of the plans of experiments. The orthogonal composite plane allowed us to establish a second-degree polynomial mathematical equation, which described the variation of the oxygen insertion rate as a function of the different synthesis factors (the mass of the hydroxyapatite attacked by the perchloric acid, the pH of the reaction medium, the temperature of the reaction medium and the concentration of the hydrogen peroxide), taking into account their interactions. The results obtained by this study show that the interaction of the mass of the attacked hydroxyapatite with itself is the most influential factor on the oxygen insertion rate and that the insertion optimum is 5.06%. This study allowed us to have a higher oxygen insertion rate compared to other studies. In addition to this interesting study, it is important to make further analysis to know more about the properties of oxygenated apatite, and to combine this apatite with polymers whose purpose is to have biomedical composites.

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Copyright information

© The Tunisian Chemical Society and Springer Nature Switzerland AG 2019

Authors and Affiliations

  1. 1.Equipe de Physico-Chimie de la Matière CondenséePCMC, Faculté des Sciences de MeknèsMeknesMorocco
  2. 2.Ecole Nationale des Sciences Appliquée Al HoceimaMohamed Premier UniversityOujdaMorocco
  3. 3.Laboratoire du Chimie de Solide Minérale et Analytique, Faculté des SciencesOujdaMorocco
  4. 4.Department of ChemistryAn-Najah National UniversityNablusPalestine

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