Major: Theoretical and Physical Chemistry Code: 62 440119gust.edu.vn/media/26/uftai-ve-tai-day26088.pdf · ELEMENTS CO-DOPED HYDROXYAPATITE ON 316L STAINLESS STEEL APPLICATION IN

VIETNAM ACADEMY OF SCIENCE AND TECHNOLOGY

GRADUATE UNIVERSITY SCIENCE AND TECHNOLOGY

-----------------------

Vo Thi Hanh

SYNTHESIS AND CHARACTERIZATION OF TRACE

ELEMENTS CO-DOPED HYDROXYAPATITE ON 316L

STAINLESS STEEL APPLICATION IN BONE IMPLANT

Major: Theoretical and Physical Chemistry

Code: 62 440119

SUMMARY OF DOCTORAL THESIS IN CHEMISTRY

Hanoi – 2018

The thesis has been completed at:

Department of Corrosion and Protection of Metals - Institute for

Tropical Technology - Vietnam Academy of Science and Technology

Scientific Supervisors:

Assoc. Prof. Dr. Dinh Thi Mai Thanh, Institute for Tropical Technology -

Vietnam Academy of Science and Technology

Reviewer 1:

Reviewer 2:

Reviewer 3:

The thesis was defended at Evaluation Council held at Graduate

University of Science and Technology - Vietnam Academy of Science

and Technology on , 2018.

Thesis can be further referred at:

- The Library of Graduate University of Science and Technology.

- National Library of Vietnam.

1

INTRODUCTION

1. The necessary of the thesis

Nowadays, 316L stainless steel (316LSS), titanium and alloys of titanium

are widely used in orthopedic surgery with the purpose of splinting bone.

Materials made of titanium and titanium alloy have a good mechanical

properties and good biocompatibility but they have a high cost. Therefore, in

Vietnam, to reduce the cost of medical services, most of the splints are made of

316L stainless steel. However, 316L stainless steel could be corroded and

limited the ability of biological compatibility in the biological environment. To

improve these problems, 316LSS is generally coated biomaterials such as

hydroxyapatite (Ca10(PO4)6(OH)2, HAp).

HAp has chemical composition and biological activity similar to the

natural bone. HAp could stimulate the bonding between the host bone to

implant materials and make bone healing ability faster. Moreover, HAp also

protects for the metal surfaces against corrosion and prevents the release of

metal ions from the substrates into the environment.

However, pure HAp has been dissolved in the physiological environment

which may lead to the disintegration of the coatings and affects the implant

fixation. These disadvantages could deal with doping some trace elements in

the HAp structure by replacing Ca2+

ions with cations and substituting OH-

group with anions. In addition, the present of trace element such as magnesium,

sodium, strontium, fluorine, zinc … has also the role to stimulate the new bone

formation and provides minerals for bone cells to grow. Besides, the problem of

postoperative infection should be concerned. Thus, antibacterial elements such

as copper, silver and zinc are also being studied to incorporated into HAp.

Based on the reasons mentioned above, the research topic of thesis is

chosen as following: “Synthesis and characterizations of trace elements co-

doped hydroxyapatite coatings on 316L stainless steel application in bone

implaint”

2. The objectives of thesis:

- Trace elements (sodium, magnesium, strontium, fluorine, copper, silver and zinc) doped NaHAp coatings are synthesized sucessfully on the 316LSS

substrates, separately and simultaneously.

- Research on the physical and chemical characteristics, cytotoxicity and antibacterial ability, biological compatibility of the NaHAp coating doping

some trace elements separately and simultaneously.

3. Research contents of the thesis:

- Investigating and selecting of optimal conditions for the synthesis of NaHAp coatings and NaHAp coatings doping magnesium, strontium and fluorine

separately and simultaneously by cathodic scanning potential method.

2

- Investigating and selecting of optimal conditions for the synthesis of NaHAp coatings doping copper, siliver and zinc separately and simultaneously by

ion exchange method.

- The HAp coatings doping 7 elements simultaneously: Mg, Sr, F, Na, Cu, Ag, Zn are studied to synthesize by the combination two methods:

electrodeposition and ion exchange.

- Studying on the biological activity of materials: 316LSS, NaHAp/316LSS, MgSrFNaHAp/316LSS and HApđt/316LSS in simulated body fluid (SBF)

solution.

- Studying on the cytotoxicity ability of powder: NaHAp, MgSrFNaHAp. - Studying on antibacterial ability of powder: NaHAp, MgSrFNaHAp,

AgNaHAp, CuNaHAp, ZnNaHAp và HApđt.

- Evaluation of the biological compatibility of materials: 316LSS, NaHAp/316LSS, MgSrFNaHAp/316LSS on dog’s body.

CHAPTER 1. OVERVIEW OF HAp AND DOPED HAp

1.1. The properties and synthesized methods of HAp and doped HAp coatings

Some trace elements doped HAp coatings have more advantages than

pure HAp coatings, such as: decrease of the dissolution, increase of the

metabolism, antibacterial ability and compatibility.

HAp coatings is deposited on the substrates by many methods: plasma,

magnetron and electrodeposition … These methods have advantages and

disadvantages. The electrodeposition has an important technology because of

the advantages: the low temperature, easily controlling the coatings thickness,

the high purity, high bonding strength and low cost of the equipment.

Furthermore, it is easy to substitute some trace elements ions (Mg2+

, Na+, K

+,

Sr2+

and F- …) into HAp coatings by addiction M(NO3)n or NaX into the

electrolyte. Dope HAp is producted according to the chemical reaction:

(10-x)Ca2+

+ 6PO43-

+ (2-y)OH- + xM

2+ + yX

- Ca10-x M x(PO4)6(OH)2-yXy

1.2. In vitro and in vivo test of HAp

The compatibility of materials is studied by immersion them in SBF

solution and investigate the formation of apatite on the material surface.

Besides, the compatibility of materials is also studied by in vivio test on the

animal.

1.4. The application of HAp, doped HAp

HAp and doped HAp are used as:

- The medicine of calcium supplements: the composition of HAp contains

a lot of calcium and be absorbed directly without transformation.

- Material for implantation: repair of the teeth and bone defects.

1.5. The situation of HAp research in the country

3

Basic on the overview of HAp and doped HAp, it can be seen that there is

no published report about doped HAp coatings in our country; in the world, the

trace elements doped HAp coatings have been only synthesized separately.

Thus, in this doctoral thesis, some trace elements (sodium, magnesium,

strontium, fluorine, copper, silver and zinc) doped HAp coatings were

synthesized separately and simultaneously. The HAp obtained coatings have

many good properties, such as: decrease of the dissolution and increase of the

metabolism, antibacterial ability and compatibility for HAp coatings.

CHAPTER 2. EXPERIMENT AND RESEARCH METHODS

2.1. Synthesis of doped HAp

2.1.1. By the electrodeposition method (cathodic scanning potential)

2.1.1.1. Electrochemical cells

The electrodeposition was carried out in a three-electrode cell with

316LSS as the working electrode, platinum foil as the counter electrode and a

saturated calomel electrode (SCE) as the reference electrode.

2.1.1.2. Synthesis of NaHAp coatings

- NaHAp coatings were synthesized on the 316LSS by cathodic scanning

potential method in 80 mL solution containing Ca(NO3)2 3×10-2

M + NH4H2PO4 1.8×10

-2 M

and NaNO3 with different concentrations: 4.10

-2 M (DNa1), 6.10

-2

M (DNa2) và 8.10-2

M (DNa3).

- NaHAp coatings were synthesized under following conditions as

follows: the different scanning potential ranges: 0 to -1.5, 0 to -1.7, 0 to -1.9

and 0 to -2.1 V/SCE; reaction temperatures: 25, 35, 50, 60 and 70 oC; pH = 4.0,

4.5, 5.0 and 5.5; scanning time: 1, 3, 5, 7 and 10; scanning rate: 3, 4, 5, 6 and 7

mV/s.

2.1.1.3. Synthesis of Mg2+

, Sr2+

or F- doped NaHAp coatings (ĐNaHAp)

ĐNaHAp were deposied at 50 oC in 80 mL solution containing at the Table

2.1 and under following conditions: the different scanning potential ranges: 0 to

-1.5, 0 to -1.7, 0 to -1.9 and 0 to -2.1 V/SCE; scanning time: 1, 3, 5, 7 and 10;

scanning rate: 3, 4, 5, 6 and 7 mV/s.

Table 2.1. Chemical composition of the electrolyte

ĐNaHAp Notation Chemical composition

MgNaHAp

DMg1 DNa2+ Mg(NO3)2 1x10-4

M


M


M


M

SrNaHAp DSr1 DNa2 + Sr(NO3)2 1x10

-5 M

DSr2 DNa2 + Sr(NO3)2 5x10-5

M

4


, Sr2+

and F- co-doped NaHAp coatings (MgSrFNaHAp)

MgSrFNaHAp were synthesized in 80 mL solution containing at DNa2 +

NaF 2.10-3

M + Sr(NO3)2 5.10-5 M + Mg(NO3)2 1.10

-3 M and under following

conditions as follows: the different scanning potential ranges: 0 to -1.5, 0 to -

1.7, 0 to -1.9 and 0 to -2.1 V/SCE; reaction temperatures: 25, 35, 50, 60 and 70 oC; scanning time: 3, 4, 5, 6, 7 and 10; scanning rate: 3, 4, 5, 6 and 7 mV/s.

2.1.2. By the ion exchange method

Preparing material of NaHAp/316LSS: NaHAp coatings were synthesized

on the 316LSS substrates by cathodic scanning potential method in the otimal

condiction: the scanning potential range of 0 to -1.7 V/SCE, the reaction

temperatures of 50 oC, the scanning time of 5 and the scanning rate of 5 mV/s

in 80 mL DNa2 solution.

2.1.2.1. Synthesis of Cu2+

, Ag+ or Zn

2+ doped NaHAp coatings

Material of NaHAp/316LSS with mass of 2.45x10-3

g was immersed in 4

mL M(NO3)n solutions with variable concentration showed on Table 2.2 and at

different time immersions: 0; 2.5; 5; 10; 20; 30; 60 and 80 minutes at room

temperature.

Table 2.2. The initial concentration of Mn+

(mol/L)

M(NO3)n Concentration (mol/L)

Cu(NO3)2 0.005 0.01 0.02 0.05 0.1

AgNO3 0.0012 0.0022 0.005 0.01 -

Zn(NO3)2 0.01 0.05 0.1 0.15 -


, Ag+ and Zn

2+ co-doped NaHAp coatings

CuAgZnHAp coatings was synthesized by the way: immersion the

material of NaHAp/316LSS about 30 minutes at room temperature in 4 mL

solutions containing simultaneously: Cu(NO3)2 0.02 M + AgNO3 0.001 M +

Zn(NO3)2 0.05 M.

2.1.3. Synthesis of Mg2+

, Sr2+

, Na+, Cu

2+, Ag

+, Zn

2+ and F

- co-doped HAp

coatings (HApđt)

- Preparing material of MgSrFNaHAp/316LSS: MgSrFNaHAp coatings

were synthesized on the 316LSS substrates by cathodic scanning potential

method in the otimal condiction: the scanning potential range of 0 to -1.7

V/SCE; reaction temperatures of 50 oC; scanning time of 5; scanning rate of 5


M


M

FNaHAp

DF1 DNa2 + NaF 5x10-4

M


M


M

5

mV/s in 80 mL the solution containing: DNa2 + NaF 2.10-3

M + Sr(NO3)2 5.10-5

M + Mg(NO3)2 1.10-3

M.

- HApđt coatings was synthesized by the way: immersion the material of

MgSrFNaHAp/316LSS about 30 minutes at room temperature in 4mL solutions

containing simultaneously: Cu(NO3)2 0.02 M + AgNO3 0.001 M + Zn(NO3)2

0.05 M.

2.2. Research method

2.2.1. Electrochemical method

Methods of scanning potential, potential applied, open circuit potential

and electrochemical impedance spectra which were carried out on AUTOLAB

equipment at Institute for tropical Technology.

2.2.2. Ion exchange method

Ion exchange was done by immersing the meterial of NaHAp/316LSS or

MgSrFNaHAp/316LSS in solution containing Mn+

with different concentrations.

2.2.3. Coatings characterization

The composition and structure of doped HAp obtained coatings were

analyzed by the method: IR, XRD, SEM, AFM, EDX (or AAS or ICP-MS),

UV-VIS.

Physical properties of the coatings was determined by: mass, thickness,

adhesion strength. The dissolution behavior of the coatings were studied by

measuring the concentration of Ca2+

dissolved from the coatings and iron

released from 316LSS substrates when the samples immersed into the 0.9 %

NaCl solution or SBF solution.

2.2.5. In vitro and in vivo Test

2.2.5.1. Invitro test in simulated body fluid (SBF) solutions

The in vitro tests in SBF solution investigated by the apatite formed

ability and the protection substrates ability of meterials and using the method:

open circuit potential (OCP), electrochemical impedance measurements at the

OCP and the polarized Tafel curves.

2.2.5.2. Cytotoxicity ability test

The safety and biocompatibility of NaHAp and MgSrFNaHAp powder

were tested on fibroblasts cells by two methods: the Trypan Blue and the MTT.

2.2.5.3. Antibacterial ability test

The antibacterial ability of NaHAp, MNaHAp, MgSrFNaHAp and

HApđt powder were tested on three strains: E.faecalis, E.coli, C.albicans và

P.aerugimosa by the disk diffusion agar method.

2.2.5.4. In vivo test

Healthy dogs are divided to 3 groups, each group of 6 dogs, which are

implanted with 3 splint made of: 316LSS, NaHAp/316LSS and

6

MgSrFNaHAp/316LSS by two methods: implantation the materials on the thigh

and on the femur. The material compatibility is evaluated by observation of the

situation the incision, the general images and the microscope images at transplant

location.

CHAPTER 3. RESULTS AND DISCUSSION

3.1. Synthesis and characterization of doped HAp coatings

3.1.1. Electrodeposition of doped HAp coatings

3.1.1.1. NaHAp coatings

a. The cathodic polarization curve

The cathodic polarization curve of 316LSS substrates at the potential

range 0 ÷ -2.1 V/SCE are shown in Figure 3.1. With this potential range, there

are several electrochemical reactions, such as:

2H+ + 2e

- H2

(3.1)

O2 + 2H2O + 4e- 4OH

- (3.2)

2 4H PO + 2e

-

3

4PO + H2 (3.3)

2 2 4H PO+ 2e

- 2

2

4HPO + H2 (3.4)

22

4HPO + 2e

- 2

3

4PO + H2 (3.5)

3NO + 2H2O + 2e 2NO

+ 2OH

- (3.6)

2H2O + 2e- H2 + 2OH

- (3.7)

2 4H PO

+ OH-

2

4HPO + H2O (3.8)

2

4HPO + OH

-

3

4PO + H2O (3.9)

10(Ca2+

, Na+) + 6

3

4PO + 2OH

− → (Ca, Na)10(PO4)6(OH)2 (3.10)

-2.2 -2.0 -1.8 -1.6 -1.4 -1.2 -1.0 -0.8 -0.6 -0.4 -0.2 0.0-6.0

-5.5

-5.0

-4.5

-4.0

-3.5

-3.0

-2.5

-2.0

-1.5

-1.0

-0.5

0.0

i (m

A/c

m2)

E (V/SCE)

Figure 3.1. The cathodic polarization curve of 316LSS substrates in the

DNa2 solution

b. Effect of Na+ concentration

The ratio of (Ca+0.5Na+Mg)/P in all obtained coatings samples at DNa1,

DNa2 and DNa3 solutions is the same the ratio of Ca/P in the natural bone

(1.67) (Table 3.1). However, to reach the Na/Ca ratio similar to its in natural

bone, the deposited NaHAp coatings in the DNa1 and DNa2 solution are

suitable. Therefore, DNa2 was chosen for the next experiments.

7

Table 3.1. The component of elements of NaHAp deposited on 316L SS in

DNa1, DNa2 and DNa3 solutions

DD Weigh (%)

Na / Ca (0.5 Na+ Ca)/ P P Ca Na

DNa1 17.25 36.09 0.32 0.0155 1.63

DNa2 16.80 33.20 1.50 0.0785 1.61

DNa3 16.60 33.09 2.20 0.1156 1.58

4000 3500 3000 2500 2000 1500 1000 500

Tra

nm

ista

nce (

a.u

)

Wave number (cm-1)

447

566

603

874

1384

1641

3441

PO

43-

PO

43-

CO

32-

CO

32-

OH

-

H2O

1036

10 20 30 40 50 60 70

Inte

nsit

y

degree

1

111

11

1NaHAp

HAp (NIST)

(a)

2

3

2

11

1

1. HAp; 2. CrO.FeO.NiO; 3. Fe

Figure 3.2. IR spectra and XRD patterns of NaHAp deposited in DNa2

solution

Both IR spectra and XRD patterns of NaHAp deposited in DNa2 solution

exhibit that NaHAp coatings have crystals structure and single phase of HAp

(Figure 3.2).

c. Effect of the scanning potential range

The charge, mass, thickness and adhesion strength of NaHAp coatings at

the different potential ranges show that the thickness and adhesion strength of

NaHAp coatings reaches the maximum value at potential range of 0 ÷ -1.7

V/SCE (Table 3.2). Thus, the potential range 0 to -1.7 V/SCE is chosen for

NaHAp coatings electrodeposition.

Table 3.2. The variation of charge, mass, thickness and adhesion strength

of obtained NaHAp coatings at the different scanning potential ranges

Scanning potential ranges

(V/SCE)

Charge

(C)

Mass

(mg/cm2)

Thickness

(µm)

Adhesion

(MPa)

0 ÷ -1.5

0.41 1.00 3.2 -

0 ÷ -1.7

3.23 2.45 7.8 7.2

0 ÷ -1.9

4.29 1.82 5.8 7.1

0 ÷ -2.1

6.57 1.67 5.3 7.0

d. Effect of electrodeposition temperature

The SEM images of NaHAp coatings deposited in DNa2 at different

temperatures show that the temperature have an effected on the morphology of

obtained coatings.

The XRD diffraction data of NaHAp coatings at the different temperatures

are shown in Figure 3.4. The typical peaks of the 316LSS substrates were

8

observed in all samples. At 25 and 35 oC, the obtained coatings is mostly

dicalcium phosphate dehydrate (CaHPO4.2H2O, DCPD) with the typical peaks

at 2 12o and 24

o. With temperature from 50°C, the peaks of DCPD are not

detected and there are only characteristic peaks of HAp phase at 2 26o (002),

32o (211), 33

o (300), 46

o (222) and 54

o (004). Thus, 50

oC is chosen to prepare

NaHAp coatings.

Figure 3.3. The SEM images of deposited NaHAp coatings at different

temperatures

10 20 30 40 50 60 70degree

Inte

nsity

350C

250C2

500C

600C

1. HAp; 2. DCPD

3. CrO.FeO.NiO; 4. Fe

1 1

3

43

2

111

Figure 3.4. XRD patterns of NaHAp deposited at different temperatures

e. Effect of pH

Results of mass and thickness of NaHAp coatings with pH solusions from

4.0 to 5.5 show on table 3.3. The results indicate that their values reaches the

highest value at pH0=4.5. Thus, pH0 is chosen for NaHAp coatings

electrodeposition.

Table 3.3. The variation of mass and thickness of obtained NaHAp coatings at

pH solutions difference

pH 4.0 4.5 5.0 5.5

Mass of NaHAp coatings (mg/cm2) 2.05 2.43 1.54 1.31

Thickness of NaHAp coatings (µm) 6.55 7.80 4.92 4.19

g. Effect of the scanning times

The charge, mass, thickness and adhesion strength of NaHAp coatings at

the different scanning times show that the thickness and adhesion strength of

NaHAp coatings are highest at 5 scanning times (Table 3.4).


of obtained NaHAp coatings at the different scanning times

Scanning times

(times)

Charge

(C)

Mass

(mg/cm2)

Thickness

(µm)

Adhesion

(MPa)

9

1 0.74 0.52 1.6 -

3 2.21 1.50 4.7 7.2

5 3.23 2.45 7.8 7.2

7 4.07 1.27 4.1 6.3

10 5.20 1.05 3.4 6.0

The SEM images of obtained NaHAp coatings show that: they have slate

shapes with large size at 3 scanning times; plate shapes and denser with the size of

150×25 nm at 5 scanning times; both slate and plate shapes at 7 scanning times

(figure 3.5).

Based on the above results, 5 scanning times is selected for NaHAp

coatings electrodeposition.

Figure 3.5. The SEM images of NaHAp coatings deposited at different

scanning times

h. Effect of the scanning rate

The thickness of obtained NaHAp coatings is highest at 5 mV/s scanning

rates (Table 3.5) so it is chosen to deposite the HAP coatings.


of obtained NaHAp coatings at the different scanning rates

Scanning rates

(times)

Charge

(C)

Mass

(mg/cm2)

Thickness

(µm)

Adhesion

(MPa)

3 5.09 1.95 6.2 6.2

4 4.11 2.15 6.9 6.5

5 3.23 2.45 7.8 7.2

6 2.21 1.27 4.1 7.8

7 1.85 0.93 3.0 10.6


, Sr2+

or F- doped NaHAp coatings (ĐNaHAp)

a. Effect of concentration

The cathodic polarization curve of 316LSS substrates in the different

solusions shows on figure 3.6. Concentration increasing of Mg2+

, Sr2+

or F- ion

in the solution leads improving ionic strength of the electrolyte and the

reducting speed of NO3- to OH

- raise. Thus, the cathodic current density

increases.

10

In the potential range of 0 ÷ -1,7 V/SCE, the reducted reactions are listed

at section 3.1.1. Then, doped HAp coatings (MgNaHAp, SrNaHAp or FNaHAp)

is producted on the cathode substrates according to the chemical reaction:

10(Ca2+

,Mg2+

,Na+) + 6PO4

3− + 2OH

− → (Ca,Mg,Na)10(PO4)6(OH)2 (3.12)

10(Ca2+

,Sr2+

,Na+) + 6PO4

3− + 2OH

− → (Ca,Sr,Na)10(PO4)6(OH)2 (3.13)

(Ca,Na)10(PO4)6(OH)2 + xF- + xH

+ (Ca,Na)10(PO4)6(OH)2-xFx + xH2O (3.14)

-1.8 -1.6 -1.4 -1.2 -1.0 -0.8 -0.6 -0.4 -0.2 0.0

-6

-5

-4

-3

-2

-1

0 (a)

DMg4

DMg3

DMg2

DMg1

DNa2

i (m

A/c

m2)

E (V/SCE) -1.8 -1.6 -1.4 -1.2 -1.0 -0.8 -0.6 -0.4 -0.2 0.0

-4

-3

-2

-1

0 (b)

DSr4

DSr3

DSr2

DSr1

DNa2

i (m

A/c

m2)

E (V/SCE) -1.8 -1.6 -1.4 -1.2 -1.0 -0.8 -0.6 -0.4 -0.2 0.0

-6

-5

-4

-3

-2

-1

0(c)

DF3

DF2

DF1

DNa2

i (m

A/c

m2)

E (V/SCE)

Figure 3.6. The cathodic polarization curve of 316LSS substrates in the

solusions with different concentrations of Mg2+

(a), Sr2+

(b) và F- (c) ions

Table 3.6 shows that the concentration of doped ions in the solution

increases leading to the increase of their components and the atomic ratios of

X/Ca in obtained coatings. However, to reach the X/Ca similar to its in natural

bone, the soluion of DMg1, DMg2, DMg3, DSr1 or DSr2 are suitable to

deposite the MgNaHAp or SrNaHAp coatings. The ratio of F/Ca is smaller than

its in natural bone but the extension of F- concentration is more than 2.10

-3 M

leading to precipitation of CaF2 in the solution. Therefore, DMg3 or DSr2 or

DF3 is chosen to deposite the MgNaHAp or SrNaHAp or FNaHAp coatings.

Table 3.6. The element components of obtained coatings at diffrent

solutions

Solutions Weigh (%)

Ca P Na Mg Sr F

DMg1 34.12 17.18 1.21 0.06 - -

DMg2 35.20 17.90 1.13 0.12 - -

DMg3 34.60 18.10 1.20 0.20 - -

DMg4 34.20 18.50 1.10 0.40 - -

DSr1 34.19 17.25 1.27 - 1.74.10-4

-

DSr2 34.72 17.96 1.22 - 3.68.10-4

-

DSr3 33.34 17.46 1.16 - 6.30.10-4

-

DSr4 33.32 17.36 1.12 - 1.00.10-3

-

DF1 38.40 18.90 1.15 - - 1.01

DF2 37.20 18.01 1.50 - - 1.30

DF3 33.10 16.80 1.90 - - 1.55

11

Table 3.7. The atomic ratios of X/Ca, Y/P and the formula of

ĐNaHAp coatings

DD Na/Ca X/Ca Y/ P The formula (expectation)

DMg1 0.062 2.90x10-3

1.59 Ca9.403Mg0.027Na0.570(PO4)6(OH)2

DMg2 0.056 5.70x10-3

1.58 Ca9.438Mg0.052Na0.510(PO4)6(OH)2

DMg3 0.060 9.60x10-3

1.54 Ca9.378Mg0.086Na0.536(PO4)6(OH)2

DMg4 0.056 1.95x10-2

1.50 Ca9.352Mg0.168Na0.480(PO4)6(OH)2

DSr1 0.065 1.74x10-4

1.64 Ca9.403Sr0.002Na0.595(PO4)6(OH)2

DSr2 0.061 3.68x10-4

1.59 Ca9.447Sr0.003Na0.549(PO4)6(OH)2

DSr3 0.0605 6.30x10-4

1.57 Ca9.457Sr0.006Na0.537(PO4)6(OH)2

DSr4 0.049 1.00x10-3

1.58 Ca9.469Sr0.009Na0.521(PO4)6(OH)2

DF1 0.052 5.50x10-2

1.66 Ca9.508Na0.492(PO4)6(OH)1.477F0.523

DF2 0.070 7.40x10-2

1.67 Ca9.326Na0.674 (PO4)6(OH)1.293F0.707

DF3 0.099 9.90x10-2

1.67 Ca9.085Na0.915(PO4)6(OH)1.097F0.903

(X/Ca = Mg/Ca or Sr/Ca or F/Ca; Y/P = (0,5Na+ Ca + Mg + Sr)/P)

b. The effect of scanning potential range

Table 3.8 shows that the scanning potential range of 0 ÷ -1.7 V/SCE for

MgNaHAp + SrNaHAp and at 0 ÷ -1.8 V/SCE for FNaHAp, the mass, thickness

and adhension strength of obtained ĐNaHAp coatings are best. Thus, the

scanning potential range of 0 ÷ -1.7 V/SCE is selected to synthesize the MgNaHAp

+ SrNaHAp coatings and 0 ÷ -1.8 V/SCE for FNaHAp coatings.


of deposited ĐNaHAp coatings at the different scanning potential ranges

ĐNaHAp

Scanning

potential ranges

(V/SCE)

Charge

(C)

Mass

(mg/cm2)

Thickness

(µm)

Adhesion

(MPa)

MgNaHAp

0 ÷ -1.5

0.42 1.21 5.5 7.3

0 ÷ -1.7

3.56 2.63 8.1 7.2

0 ÷ -1.9

4.52 1.96 6.3 7.1

0 ÷ -2.1

6.85 1.41 4.5 7.0

SrNaHAp

0 ÷ -1.5

0.31 1.12 5.2 7.4

0 ÷ -1.7

3.51 2.35 7.6 7.3

0 ÷ -1.9

4.32 1.91 6.1 7.1

0 ÷ -2.1

6.69 1.45 4.7 7.0

FNaHAp

0 ÷ -1.6

0.50 1.40 4.2 7.6

0 ÷ -1.7

3.63 2.40 7.8 7.1

0 ÷ -1.8

4.25 2.90 8.3 6.9

0 ÷ -1.9

4.97 1.80 5.4 5.5

12

c. The effect of the scanning times

The results of the mass and thickness of ĐNaHAp coatings at the

different scanning times show on Table 3.9. The mass, thichness and adhesion

strength of deposited ĐNaHAp coatings at 5 scanning times are higher than

othes scanning times so 5 scanning times is chosen to deposited the ĐNaHAp

coatings.


of deposited ĐNaHAp coatings at the different scanning times

ĐNaHAp Scanning

times

Charge

(C)

Mass

(mg/cm2)

Thickness

(µm)

Adhesion

(MPa)

MgNaHAp

1 0.76 0.57 1.6 -

3 2.40 1.72 5.5 7.3

5 3.51 2.63 8.1 7.2

7 4.61 1.41 4.5 6.3

10 6.33 0.98 3.1 5.7

SrNaHAp

1 0.56 0.37 1.2 -

3 2.13 1.51 5.2 10.0

5 3.51 2.35 7.6 7.3

7 4.02 1.51 4.8 7.5

10 4.98 1.12 3.7 5.2

FNaHAp

1 0.78 0.62 1.8 -

3 2.61 1.80 5.6 7.4

5 3.82 2.40 7.8 7.1

7 5.14 1.52 4.9 6.1

10 6.96 1.26 4.1 5.8

d. Characterization of ĐNaHAp coatings

The analysed results of IR spectra, XRD patterns and SEM images show

that deposited ĐNaHAp coatings have crystals structure and single phase of

HAp (Figure 3.7) and the morphology change because of the presence of Mg,

Sr or F into NaHAp coatings (Figure 3.8).

4000 3500 3000 2500 2000 1500 1000 500

Wave number (cm-1)

Tra

nm

ista

nce (

a.u

)

565

1390

1035

3445

H2O

NaHAp

MgNaHAp

437

602

874

1645

FNaHAp

SrNaHAp

F-

PO

4

3-

CO

3

2-

PO

4

3-

CO

32-

OH

-

10 15 20 25 30 35 40 45 50 55 60 65 70

MgNaHAp

NaHAp

SrNaHAp

In

ten

sit

y

degree

FNaHAp

3

2

2

1111

1


Figure 3.7. IR spectra and XRD patterns of ĐNaHAp coatings

13

Figure 3.8. SEM images of NaHAp and ĐNaHAp coatings


, Sr2+

and F- co-doped NaHAp coatings (MgSrFNaHAp)

a. The effect of the scanning potential range

The MgSrFNaHAp coatings is produced on the cathode substrates

according to the chemical reaction:

10(Ca2+

,Na+,Mg

2+,Sr

2+) + 6

3

4PO

+ 2OH- (Ca,Na,Mg,Sr)10(PO4)6(OH)2 (3.15)

(Ca,Na,Mg,Sr)10(PO4)6(OH)2 + x F- + x H

+ (Ca,Na,Mg,Sr)10(PO4)6(OH)2- xFx

+ xH2O (3.16)

The change of charge, mass, thickness and adhesion strength of

MgSrFNaHAp coatings with the different potential ranges shows on Table 3.10.

The mass and thickness of MgSrFNaHAp coatings are highest at 0 ÷ -1,7

V/SCE.

Table 3.10. The variation of charge, mass, thickness and adhesion

strength of deposite MgSrFNaHAp coatings with the different scanning

potential ranges

Scanning potential

ranges (V/SCE)

Charge

(C)

Mass

(mg/cm2)

Thickness

(µm)

0 ÷ -1.5

1.13 1.01 3.1

0 ÷ -1.7

4.32 3.17 8.9

0 ÷ -1.8 5.08 2.54 7.8

0 ÷ -1.9

5.92 1.95 5.9

0 ÷ -2.1

7.84 1.47 4.2

SEM images of MgSrFNaHAp coatings deposited with different potential

ranges are presented in Fig. 3.9. With potential ranges of 0 ÷ -1.7 và 0 ÷ -1.8

V/SCE, the deposited coatings are denser with cylinder shapes.

Therefore, the potential ranges of 0 ÷ -1,7 V/SCE is selected to

electrodeposited MgSrFNaHAp coatings.

Figure 3.9. SEM images of MgSrFNaHAp coatings deposited with the

potential ranges: (a) 0 ÷ -1,5; (b) 0 ÷ -1,7; (c) 0 ÷ -1,8; (d) 0 ÷ -1,9 (V/SCE)

14

b. Effect of electrodeposition temperature

At 25 and 35 oC, the deposited coatings are mostly DCPD. With higher

temperature, the peaks of DCPD are not detected and there are only peaks of

HAp phase (Figure 3.10a). Thus, 50 oC is chosen to prepare MgSrFNaHAp

coatings.

c. The effect of the scanning times

The XRD patterns indicate that the phase of deposited coatings at one

scanning times is only DCPD without HAp. At 3 scanning times, it appears the

phase of HAp but DCPD is still the mainly component. From 5 scanning times,

the obtained coatings have single phase of HAp (Figure 3.10b). Thus, 5

scanning times is chosen for MgSrFNaHAp coatings electrodeposition.

d. Effect of the scanning rate

Figure 3.10c presents the XRD patterns of MgSrFNaHAp coatings

deposited at different scanning rates. Both XRD patterns exhibit the

hydroxyapatite phase with the typical peaks at 2 of 32o and 26

o. However, with

the scanning rate at 6, 7 mV/s, there are also appear peaks of DCPD at 2 of

12o. Thus, scanning rate 5 mV/s is chosen to deposite of MgSrFNaHAp

coatings.

10 20 30 40 50 60 70

degree

Inte

nsi

ty

25 0C

35 0C

50 0C

60 0C

(a)

111431

2

1 111

1 1. HAp; 2. DCPD; 3. CrO.FeO.NiO; 4. Fe

70 0C

10 20 30 40 50 60 70

Inte

nsi

ty

degree

1 lÇn quÐt

3 lÇn quÐt

2

5 lÇn quÐt

7 lÇn quÐt

(b)

10 lÇn quÐt1

1 13

43

11

1. HAp; 2. DCPD; 3. CrO.FeO.NiO; 4. Fe

10 20 30 40 50 60 70

degree

Inte

nsit

y

2

7 mV/s

2

4 mV/s

5 mV/s

6 mV/s

(c)

1 31 1

1. HAp; 2. DCPD; 3. CrO.FeO.NiO; 4. Fe

43

11

3 mV/s

Figure 3.10. XRD patterns of deposited coatings with the change:

(a) temperature, (b) scanning time, (c) scanning rate

e. Characterization of MgSrFNaHAp coatings

The EDX spectra of MgSrFNaHAp coatings shows that: the presence of 7

main elements doped in HAp including: Ca, O, P, Mg, Na, F and Sr (Table

3.11). These results have been used to calculate the atomic ratios of M/Ca, (Ca

+ M)/P (Table 3.12). The ratios suggest that the components of the elements in

the coatings are similar to component of mineral phase in natural bone .

Table 3.11. The element component of MgSrFNaHAp coatings deposited

on 316L SS

Elements O Ca P Na Sr Mg F

Weigh (%) 39.34 32.65 15.76 0.56 0.03 0.14 1.50

Atomic (%) 68.20 18.00 11.20 0.99 0.01 0.13 1.47

15

Table 3.12. The atomic ratios of M/Ca and M/P in MgSrFNaHAp

coatings and in natural bone

M/Ca (M: Na, Mg, Sr, F) MgSrFNaHAp In natural bone

Na/ Ca 8.8×10-2

0.102

Mg/Ca 1.2×10-3

6.7×10-3

÷ 1.7×10-2

Sr/ Ca 8.9×10-4

2.7×10-4

÷ 9.8×10-4

F/ Ca 1.3×10-2

0.024 ÷ 0.15

(0,5 Na + Mg + Sr + Ca)/P 1.664 -

SEM and AFM images of deposited coatings are shown in Figure 3.11. At

the same conditions, MgSrFNaHAp coatings with the presence of Mg, Sr, F

are high density, uniform and has a rod shape, while HAp coatings has a plate

shape. The roughness value (Ra) of MgSrFNaHAp coatings is less 2 times than

its of NaHAp coatings.

Figure 3.11. SEM (a) and AFM (b) images of NaHAp and MgSrFNaHAp coatings

3.1.2. Synthesis of doped HAp by ion exchange method


, Ag+ or Zn

2+ doped NaHAp coatings

a. Effects of M2+

concentration

For the ion exchange between HAp and Cu2+

, the initial concentration of

Cu2+

increases from 0.005 M ÷ 0.02 M, the ion exchange capacity rises rapidly.

When the concentration of Cu2+

was elevated to 0.05 M and 0.1 M, the capacity

altered slightly, as the ion exchange process had reached trace of the

equilibrium. Therefore, 0.02 M Cu2+

solution is used to synthesize CuHAp

coatings (Table 3.13).

For ion exchange between HAp coatings with Ag+ and Zn

2+ ions, ion

exchange capacity increased simutaneously as Ag+ and Zn

2+ concentration

increased. Xray diffraction of the obtained samples after ion exchange are

presented in Fig. 1. With concentration of Ag+

from 0.001 M to 0.005 M and

Zn2+

from 0.01 M to 0.1 M, all the samples have crystals structure and single

phase of HAp. Contrary, with the Ag+ concentration of 0.01 M, the obtained

coatings have mainly the phase of Ag3PO4. Therefore, 0.001 M Ag+ and 0.05 M

Zn2+

solutions are used to synthesize AgHAp and ZnHAp coatings.

16

Table 3.13. Ion exchange capacity and the formula of MHAp

Ion Concentration

Mn+

(M) Q (mmol/g)

The formula

MNaHAp (expectation)

Cu2+

0.005 0.065 Ca9.278Na0.722 Cu0.065(PO4)6(OH)2

0.01 0.117 Ca9.162Na0.722 Cu0.116(PO4)6(OH)2

0.02 0.166 Ca9.113Na0.722 Cu0.165(PO4)6(OH)2

0.05 0.204 Ca9.076Na0.722 Cu0.202(PO4)6(OH)2

0.1 0.216 Ca9.064Na0.722 Cu0.214(PO4)6(OH)2

Ag+

0.001 0.259 Ca9.021Na0.722 Ag0.257(PO4)6(OH)2

0.002 0.374 Ca8.907Na0.722 Ag0.371(PO4)6(OH)2

0.005 0.569 Ca8.714Na0.722 Ag0.564(PO4)6(OH)2

0.01 2.470 -

Zn2+

0.01 0.499 Ca8.783Na0.722 Zn0.495(PO4)6(OH)2

0.05 1.248 Ca8.040Na0.722 Zn1.238(PO4)6(OH)2

0.1 3.858 Ca5.452Na0.722 Zn3.826(PO4)6(OH)2

10 15 20 25 30 35 40 45 50 55 60 65

a

Inte

nsity

b

d

c

f

e

111

3

32

2

degree

1 24

444

4

4

4

4

1. HAp; 2. CrO.FeO.NiO; 3. Fe; 4. Ag3PO

4

g

Figure 3.12. XRD patterns of the obtained samples after ion exchange

between HAp and solution containing: 0.01 M Zn2+

(a), 0.05 M Zn2+

(b), 0.1 M

Zn2+

(c) and 0.001 M Ag+ (d), 0.002 M Ag

+(e), 0.005 M Ag

+ (f), 0.01 M Ag

+ (g)

b. Effect of contact time

The change in ion exchange capacity folowing the contact time are

presented in Fig. 2. The results show that: after 10 minutes contact with Ag+ ion

and after 30 minutes contact with Cu2+

or Zn2+

, the ion exchange capacity has

reached equilibrium trace (figure 3.13); if the contact time is longer, this value

changes light. Thus, the contact time is selected to synthesize CuNaHAp or

ZnNaHAp of 30 minutes and AgNaHAp coatings of 10 minutes.

17

0 10 20 30 40 50 60 70 80 900.12

0.13

0.14

0.15

0.16

0.17

0.18

0.19

NaHAp + Cu2+

0,02M

Q (

mm

ol C

u2+/g

NaH

Ap

)

Time (min) 0 10 20 30 40 50 60 70 80 90

0.30

0.35

0.40

0.45

0.50

0.55

0.60

Time (min)

NaHAp + Ag+0,001M

Q (

mm

ol A

g+/g

NaH

Ap

)

0 10 20 30 40 50 60 70 80 90

0.6

0.8

1.0

1.2

1.4

1.6

Time (min)

NaHAp + Zn2+

0,05M

Q (

mm

ol Z

n2+/g

NaH

Ap

)

Figure 3.13. The change in ion exchange capacity of the HAp coatings

with Mn+

solutions

c. Characterization of CuNaHAp, AgNaHAp, ZnNaHAp coatings

4000 3500 3000 2500 2000 1500 1000 500

Tra

nm

ista

nce

(a.

u)

Wave number (cm-1)

1643

3430

CuHAp

AgHAp

ZnHAp

NaHAp

H2O PO

43-

PO

43-

CO

32-

OH

-

565602

1034

1390

10 20 30 40 50 60

degree

Inte

nsit

y

AgNaHAp

CuNaHAp

NaHAp

ZnNaHAp

(a)1

11

11

32

2

1


Figure 3.14. IR spectra and XRD patterns of NaHAp and MNaHAp

coatings

Both IR spectra and XRD patterns of MNaHAp coatings exhibit that

NaHAp coatings have crystals structure and single phase of HAp (Figure 3.14).

SEM images of HAp and MHAp coatings show that with the present of

Cu, Ag, Zn in HAp structure, the morphology changes from plate shape of HAp

to rod shape of CuHAp; fiber shape of AgHAp and coral-shape of ZnHAp

(Figure 3.15).

Figure 3.15. SEM images of NaHAp and MnaHAp coatings


, Ag+ and Zn

2+ co-doped NaHAp coatings

The ion exchange capacity of NaHAp coatings with the solution

containing simultaneously: Cu2+

0.02 M + Ag+ 0.001 M + Zn

2+ 0.05 M at 30

minutes is smaller than its with the solution containing separately: Cu2+

0.02 M

or Ag+ 0.001 M or Zn

2+ 0.05 M (Table 3.14).

Table 3.14. The ion exchange capacity and the fomular of CuAgZnNaHAp coatings

Ion Concentration M

n+

(M) Q (mmol/g)

The formula

MNaHAp (expectation)

Cu2+

0.02 0.121 Ca8.550Na0.722 Cu0.121

18

Ag+ 0.001 0.207 Ag0.208Zn1.121(PO4)6(OH)2

Zn2+

0.05 1.117

Results of IR spectra, XRD patterns and SEM images demonstrate that

CuAgZnNaHAp obtained coatings have crystals structure with slate shape and

single phase of HAp (Figure 3.16).

4000 3000 2000 1000

Wave number (cm-1)

Tra

nm

ista

nce

(a.

u)

(a)

565

602

1390

1034

1643

3432

(b)

10 15 20 25 30 35 40 45 50 55 60 65In

ten

sit

y

degree

(a)

(b)

2

32

2 11

11

1

1


Figure 3.16. IR spectra and XRD patterns of NaHAp (a) and

CuAgZnNaHAp (b) coatings and SEM images of CuAgZnNaHAp coatings

3.1.3. Synthesis of Mg2+

, Sr2+

, Na+, Cu

2+, Ag

+, Zn

2+ and F

- co-doped HAp

coatings (HApđt)

The EDX spectra of HApđt coatings obtained shows that: There are the

presence of 10 main elements doped in HAp, including: Ca, O, P, Mg, Na, F,

Sr, Cu, Ag and Zn with their components listed at table 3.15. The atomic ratios

of X/Ca and (0,5Na+Ca+Mg+Sr+Cu+0,5Ag+Zn)/P (symbol Z/P) are calculated

and show on Table 3.16. To compare with component of mineral phase in

natural bone, the element components of Mg, Sr, F and Na in the coatings are

similar to but this values of Cu, Ag and Zn and Na are higher to increase the

antibacterial ability of the coatings.

Table 3.15. The element component of HApđt coatings

Elements O P Ca Na Mg Sr F Cu Ag Zn

Weigh (%) 29.01 14.67 52.83 0.15 0.04 0.03 1.07 0.18 0.39 1.06

Atomic (%) 49.17 12.63 35.82 0.18 0.05 0.008 1.53 0.08 0.1 0.44

Table 3.16. The atomic ratios of X/Ca and Z/P in HApđt coatings and in

natural bone

The atomic

ratios F/Ca Mg/Ca Sr/Ca Na/Ca Cu/Ca Ag/Ca Zn/Ca Z/P

HApđt

coatings 0.0646 2x10

-3 4x10

-4 8x10

-3 3x10

-3 4x10

-3 0.0187 1.65

Natural bone 0.149 0.176 4x10-4

0.102 1x10-4

1x10-6

6x10-4

1.67

The fomular

(expectation) Ca9.005Mg0.019Sr0.004F0.638Cu0.032Ag0.041Zn0.185Na0.074(PO4)6(OH)2

IR spectra, XRD patterns and SEM images of HApđt obtained coatings

demonstrate that they have crystals structure with slate shape and single phase

of HAp (Figure 3.17).

19

4000 3000 2000 1000

Tra

nm

ista

nce (

a.u

)

Wave number (cm-1)

(a)

565

602

1390

1034

1643

3432

(b)

10 15 20 25 30 35 40 45 50 55 60 65

degree

Inte

nsit

y

3

2

2

1

1

11

1

1

(a)

(b)


Figure 3.17. IR spectra and XRD patterns of NaHAp (a) and HApđt (b)

coatings and SEM images of HApđt coatings

The dissolution behaviors of HApđt, MgSrFNaHAp and NaHAp coatings is

studied by immersions materials in 0.9% NaCl and SBF solutions. For all

samples, the dissolved amount of Ca2+

ions from these coatings increases with

immersion time. However, The dissolution of HApđt coatings is slowest and of

NaHAp is faster at any time (Figure 3.18a). The release concentration of iron

ions from substrates increases according to time for all sample. Because HApđt

coatings play as a barrier to protect the substrates and the dissolution of the

coatings decreases with the presence of the trace elements in HAp structure so

the iron ion release is arranged in order: 316LSS > NaHAp/316LSS >

MgSrFNaHAp/316LSS > HApđt/316LSS.

This suggested that the protect ability for the substrates of the coatings:

HApđt > MgSrFNaHAp > NaHAp.

0 2 4 6 8 10 12 14 16 18

2

3

4

5

6

7

8

Time (days)

a: NaHAp/TKG316L

b: MgSrFNaHAp/TKG316L

c: HAp®t

/TKG316L

c

b

a

Co

ncen

trati

on

Ca

2+(p

pm

)

7 14 21 280

50

100

150

200

Time (days)

a: TKG316L

b: NaHAp/TKG316L

c: MgSrFNaHAp/TKG316L

d: HAp®t

/TKG316L

d

c

b

a

Co

ncen

trati

on

Fe (

pp

b)

Figure 3.18. The release concentration of Ca

2+ (1) and Fe

n+ (2)

3.2. The in vitro and in vivo test

3.2.1. The in vitro test

3.2.1.1. Invitro test in simulated body fluid (SBF) solutions

a. The variation of the pH and the open circuit potential (OCP - Eo) value

With 316LSS sample, the pH solution decreases and Eo tends to increase

during immersion time (Figure 3.19).

20

0 5 10 15 20 25

6.2

6.4

6.6

6.8

7.0

7.2

7.4

7.6

7.8

8.0

8.2

Time (days)

(a): TKG316L

(b): TKG316L/NaHAp

(c): TKG316L/MgSrFNaHAp

(d): TKG316L/HApdt

(d)

(c)

(b)

(a)

pH

0 2 4 6 8 10 12 14 16 18 20 22

-160

-120

-80

-40

0

40

80

120

Time (days)

(a): TKG316L

(b): NaHAp/TKG316L

(c): MgSrFNaHAp/TKG316L

(d): HAp®t

/TKG316L

(d)

(c)

(b)

(a)

E0 (

V/S

CE

)

Figure 3.19. The variation of pH (1) and Eo (2) vs. different immersion

times in SBF solution

With doped HAp/316LSS materials, pH solution and value of Eo have

fluctuated which shows the formation of new apatite crystals or the dissolution

of the coatings in the immersion process. The dissolution HAp causes to

increase pH solution Eo. In the process of forming apatite, OH- as Ca

2+, PO4

3- is

consumed large quantities leading to reduce pH and rise Eo.

b. The electrochemical impedance

During 21 immesion days, the impedance of 316LSS increases, these

values of doped HAp coated 316LSS changes, but they are much higher than

316LSS because of protection ability of coatings and tend to increase which

demonstrates that the rate of the formation is higher than the rate of the

dissolution of the coatings (Figure 3.20).

Moreover, the variations of impedance modulus at 100 mHz frequency

show that the values of impedance modulus of MgSrFNaHAp/316LSS and

HApđt/316LSS material are higher than of NaHAp/316LSS and 316LSS which

indicates that HAp doped with the present of some trate elements have the

protection ability better than NaHAp coatings.

0 1 2 3 4 5

0

1

2

3

4

5

x : 100 mHz

10 days

14 days

17 days

21 days

1 day

3 days

5 days

7 days

Z'' (

.cm

2)

Z' (.cm2)

316LSS

0 1 2 3 4 5 6 7 8 9 10 11 12

0

1

2

3

4

5

6

7

8

9

10

11

12

x : 100 mHz

10 days

14 days

17 days

21 days

1 day

3 days

5 days

7 days

Z'' (

.cm

2)

Z' (.cm2)

NaHAp/316LSS

0 2 4 6 8 10 12 14 16 18 20 22

0

2

4

6

8

10

12

14

16

18

20

22

x : 100 mHz

10 days

14 days

17 days

21 days

1 day

3 days

5 days

7 days

Z''

(

.cm

2)

Z' (.cm2)

MgSrFNaHAp/316LSS

0 2 4 6 8 10 12 14 16 18 20 22 24 26 28

0

2

4

6

8

10

12

14

16

18

20

22

24

26

28

x : 100 mHz

10 days

14 days

17 days

21 days

1 day

3 days

5 days

7 days

HAp®t

/316LSS

Z' (.cm2)

Z'' (

.cm

2)

0 2 4 6 8 10 12 14 16 18 20 22

2

4

6

8

10

12

14

16

18

20

22

24

26

28

30a: 316LSS

b: NaHAp/316LSS

c: MgSrFNaHAp/316LSS

d: HApdt

/316LSS

(a)

(b)

(c)

(d)

IZI (k

.cm

2)

Time (days)

Figure 3.20. Nyquist plots

and the variation of

impedance modulus at 100

mHz versus immersion time

in SBF solution

21

c. The polarized Tafel curves

The presence of trace elements in HAp structure leads to move the

corrosion potential (Ecorr) toward the positive side and reduce the corrosive

current density (icorr) in comparation with 316LSS (Figure 3.21 and Table 3.17).

This indicates that the protection ability for the substrates of doped HAp

coatings is better than that of NaHAp one.

-0.8 -0.6 -0.4 -0.2 0.0 0.2 0.4 0.6-8.5

-8.0

-7.5

-7.0

-6.5

-6.0

-5.5

-5.0

-4.5

-4.0

-3.5(a): 316LSS

(b): NaHAp/316LSS

(c): MgSrFNaHAp/316LSS

(d): HAp®t

/316LSS

dc

b

a

lg(i

), A

/cm

2

E (V/SCE)

Table 3.17. The values of the corrosion current

density (icorr) and the corrosion potential (Ecorr)

of meterials after immersion in SBF solution

Materials Ecorr

(V)

icorr

(µA/cm2)

316LSS -0.424 2.773

NaHAp/316LSS -0.354 0.842

MgSrFNaHAp/316LSS -0.258 0.355

HApđt/316LSS -0.213 0.193

Figure 3.21. The polarized

Tafel curves of materials

d. The SEM images

Figure 3.22 presents SEM images of 316LSS, NaHAp/316LSS,

MgSrFNaHAp/316LSS and HApđt/316LSS before and after immersion in SBF

solution. After immersion, the formation of new apatite crystals is observed on

the surface of all materials.

Figure 3.22. SEM images of materials before (above) and after (under)

21 immersion days in SBF solution

3.2.1.2. Cytotoxicity ability test

Results of the cytotoxicity ability test by Trypan Blue and the MTT method

show that NaHAp or MgSrFNaHAp powder with different concentrations are safe

for fibroblasts and lymphocyte cells.

3.2.1.3. Antibacterial ability test

The results of antibacterial ability test with three strains (P.aerugimosa,

E. coli and E.faecalis) show that: AgHAp and HApđt have good resistance to all

22

of them; CuHAp has a good effect on P. aerugimosa; the HAp and ZnHAp

have no effect on all strains;

3.2.2. In vivo test on the dog

3.2.2.1. Results of implantation at the thigh

At the first 3 days, the wound at implantation area has been edema but not

bleeding. After 1 month, the skin was almost completely covered (Figure 3.23).

Figure 3.24. The woud at implantation area

Microscopic images at implantation area shows that all implanted animals

have the same results. The area of the direct contact with the material is forming

a membrane link. However, some lymphocyte cells appear on 316LSS and

NaHAp/316LSS materials but in contrast, they are completely not observed on

MgSrFHAp/316LSS material.

3.2.2.2. Results of transplantation on the femur

The wound has been edema but not bleeding at the first 3 days and after 1

month, the skin was almost completely covered at the location of transplantation.

Microscopic images at transplantation area shows that:

- After 1 week: all transplanted animals have the same results. At the

transplantation area, many osteoblasts cells are observed but acute inflammation

cells still exist (Figure 3.24).

Figure 3.24. Images of NaHAp/316LSS after 1 week transplantation

- After 1 month: the acute inflammation cells have absented and the

necrosis have not observed on all transplanted materials. There are many

osteoblasts cells on 316LSS and NaHAp/316LSS but still have the lymphocyte

cells (Figure 3.25a). With MgSrFNaHAp/316LSS, the osteoblast concentrate on

the location of the bone edge to form bone (Figure 3.25b) and form a membrane

link attached on the surface of this material (Figure 3.25c).

23

Figure 3.25. Images of the osteoblast near the location of transplanted

materials: 316LSS (a), MgSrFNaHAp/316LSS (b, c) after 1 month

- After 2 months: There are many osteoblasts cells on 316LSS and

NaHAp/316LSS but still have few the lymphocyte cells (Figure 3.26a). The

tissues adhese on the surface of NaHAp/316LSS better than the 316LSS. On the

surface of MgSrFNaHAp/316LSS, the lymphocyte cells have not observed

(Figure 3.26b) and a new bone layer is produced (Figure 3.26c)


materials: 316LSS (a), MgSrFNaHAp/316LSS (b, c) after 2 months

- After 3 months: on all transplanted materials, there are not the

lymphocyte cells (Figure 3.27a, b). On the surface of MgSrFNaHAp/316LSS, a

thick new bone is formed which are rarely found out on 316LSS and

NaHAp/316LSS (Figure 3.27c).


materials: 316LSS (a), MgSrFNaHAp/316LSS (b, c) after 3 months

CONCLUSIONS

1. The optimal conditions is selected to synthesize the NaHAp coatings and NaHAp coatings doping magnesium, strontium and fluorine separately and

simultaneously by cathodic scanning potential method: the scanning

potential range of 0 to -1.7 V/SCE (0 ÷ -1.8 V/SCE for FNaHAp); the

reaction temperatures of 50 oC; the scanning time of 5; the scanning rate of

5 mV/s in DNa2, DMg3, DSr3, DF3 và DNaMgSrF, respectively. The dope

deposited HAp have crystals structure and single phase of HAp with the

thickness about 7,6 ÷ 8,1 µm. The component wt% of the elements Na, Mg,

24

Sr or F in the NaHAp, MgNaHAp, SrNaHAp, FNaHAp coatings are 1.5; 0.2;

6.3x10-4

hoặc 1.55 %, respectively and in the MgSrFNaHAp are 0.56 % Na;

0.14 % Mg; 0.03 % Sr and 1.5 % F.

2. The NaHAp coatings doping copper, siliver and zinc separately and simultaneously are synthesized successfully by ion exchange method and

by the way: immersion the material of NaHAp/316LSS at room

temperature in 4mL solutions containing separately or simultaneously:

Cu(NO3)2 0.02 M, AgNO3 0.001 M and Zn(NO3)2 0.05 M about 30 minutes

(10 minute for AgNaHAp coatings).

3. The HAp coatings doping 7 elements simultaneously: Mg, Sr, F, Na, Cu, Ag, Zn are synthesized successfully by the combination two methods:

electrodeposition and ion exchange. The HApđt obtained coatings have

crystals structure with coral shape and single phase of HAp. The component

wt% of the elements Mg, Sr, F, Na, Cu, Ag and Zn in the coatings are 0,04;

0,03; 1,07; 0,15; 0,18; 0,39 và 1,06 %, respectively. The presents of seven

elements in the coatings lead to decrease the dissolution behaviors and

increase the protect ability for the substrates.

4. The in vitro test in SBF solution by the methods of open circuit potential, electrochemical impedance measurements and the polarized Tafel curves show

that the biological activity and the protect ability of these material follow the

order: HApđt/316LSS > MgSrFNaHAp/316LSS > NaHAp/316LSS > 316LSS.

5. The cytotoxicity ability test by Trypan Blue and the MTT method show that NaHAp or MgSrFNaHAp powder with different concentrations are safe for

fibroblasts cells.

6. The antibacterial ability test with three strains (P.aerugimosa, E. coli and E.faecalis) show that: AgHAp and HApđt have good resistance to all of

them; CuHAp has a good effect on P. aerugimosa; the HAp and ZnHAp

have no effect on all strains.

7. The in vivo test on dog’s body by implantation 3 splints made from: 316LSS, NaHAp /316LSS and MgSrFNaHAp/316LSS at the thigh and on

the femur with tested time from 1 to 3 months show that their biological

compatibility follows the order: MgSrFNaHAp/316LSS > NaHAp /316LSS

> 316LSS.

NEW CONTRIBUTIONS OF THESIS

1. The HAp coatings doping 7 elements simultaneously: Mg, Sr, F, Na, Cu, Ag, Zn are synthesized successfully on 316LSS substrates by the combination

two methods: electrodeposition and ion exchange. The presents of seven

elements in the coatings lead increasing the abilities of the biological

compatibility, the antibactery and the protection for the substrates and

decreasing the dissolution behaviors in comparation with MgSrFNaHAp and

NaHAp coatings.

25

2. The in vivo tests on dog’s body with tested time from 1 to 3 months indicate that MgSrFNaHAp/316LSS with the presents of some trace

elements (Mg, Sr and F) in the coatings lead the biological compatibility

better than NaHAp/316LSS and uncoated 316LSS which is demonstrated

by the forminations a thick new bone on the surface of

MgSrFNaHAp/316LSS after 3 months transplantation.

LIST OF PUBLICATIONS

1. Pham Thi Nam, Nguyen Thi Thom, Nguyen Thu Phuong, Vo Thi Hanh,

Nguyen Thi Thu Trang, Vu Thi Hai Van, Trinh Hoang Trung, Tran Dai

Lam, Dinh Thi Mai Thanh. Electrodeposition of substainable fluoridated

Hydroxylapatite coatings on 316L stainless steel for application in bone

implaint. Green Processing and Synthesis, 5, 499 - 510, 2016 (ISI).

2. Vo Thi Hanh, Pham Thi Nam, Nguyen Thi Thom, Do Thi Hai và Dinh Thi

Mai Thanh. Electrodeposition of sodium doped hydroxyapatite coatings on

316L stailess steel. Vietnam Journal of Chemistry, 55(3), 348 - 354, 2017.

3. Vo Thi Hanh, Pham Thi Nam, Dinh Thi Mai Thanh. Electrodeposition and characterization of strontium hydroxyapatite coatings on 316L stailess steel.

Vietnam Journal of Chemistry, 55(3e12), 346 - 350, 2017.

4. Vo Thi Hanh, Pham Thi Nam and Dinh Thi Mai Thanh. Synthesis ad characterization of copper doped hydroxyapatite on on 316L stailess steel.

HNUE Journal of Science 62(3), 51 - 59, 2017.

5. Vo Thi Hanh, Le Thi Duyen, Do Thi Hai, Pham Thi Nam, Nguyen Thi Thom, Nguyen Thu Phuong, Dinh Thi Mai Thanh. Electrodeposition and

characterization of Mg2+

, Sr2+

, F-, Na

+ co-doped hydroxyapatite coatings on

316L stailess steel. Processdings of 6th Asian Symposium on Advanced

Materials, 740 - 746, 2017.

6. Vo Thi Hanh, Le Thi Duyen, Pham Thi Nam and Dinh Thi Mai Thanh. Study on the electrochemical behavior of NaHAp/316L stainless steel

materials in solution simulated body fluid. Vietnam Journal of Chemistry

55(5E1,2), 114-119, 2017.

7. Vo Thi Hanh, Pham Thi Nam, Nguyen Thu Phuong, Nguyen Thi Thom, Le Thi Phuong Thao, Dinh Thi Mai Thanh. Electrodeposition and

characterization of magnesium hydroxyapatite coatings on 316L stailess

steel. Vietnam Journal of Chemistry, 55(5), 657-662, 2017.

8. Vo Thi Hanh, Pham Thi Nam, Nguyen Thu Phuong, Dinh Thi Mai Thanh. Electrodeposition of co-doped hydroxyapatite coatings on 316L stailess

steel. Vietnam Journal of Science and technology, 56 (01), 94-101, 2018.

9. Vo Thi Hanh, Le Thi Duyen, Pham Thi Nam and Dinh Thi Mai Thanh. The

influence of NaNO3 and H2O2 to electrodeposition process of sodium doped

hydroxyapatite on 316L stailess steel substrates, HNUE Journal of Science

accepted 6/2017 (DOI: 10.18173/2354-1059.2017-0011).

Documents

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