Compositions and Properties of Titanium and Zinc Incorporated PBG - - PowerPoint PPT Presentation
Compositions and Properties of Titanium and Zinc Incorporated PBG Glass Code Glass Composition (mol%) Calcium Sodium Phosphorous Titanium Zinc Oxide Oxide Pentoxide Dioxide Oxide
Glass Composition (mol%) Glass Code Calcium Sodium Phosphorous Titanium Zinc Oxide Oxide Pentoxide Dioxide Oxide Ca30Na20P50T0Z0 Ca30Na15P50Z5 Ca30Na15P50T5 Ca29Na15P50T5Zn1 Ca28Na15P50T5Zn3 Ca27Na15P50T5Zn5 30 20 50 0 0 30 15 50 0 5 30 15 50 5 0 29 15 50 5 1 27 15 50 5 3 25 15 50 5 5
Compositions and Properties of Titanium and Zinc Incorporated PBG
1 and 7 Days HOS Viability
1 Day HOS Attachment
7 Days HOS Attachment
0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1.1 1.2 1.3 1.4 1.5 1.6 T0Z0 Z5 Thermanox T5 T5Z1 T5Z3 T5Z5
Glass Extract Relative Growth
Day 1 Day 4 Day 7
Effect of Glass Extract on Cell Proliferation
0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1.1 1.2 1.3 1.4 1.5 1.6 T Z Z 5 T h e r m a n
T 5 T 5 Z 1 T 5 Z 3 T 5 Z 5
Glass Code Relative Growth
Day 1 Day 3 Day 5 Day 7 Day 10 Day 14 Day 21
Effect of Composition on Cell Proliferation
Ca16Na39P45 Ca16Na38P45Ga1 Ca16Na36P45Ga3 Ca16Na34P45Ga5 16 39 45 16 38 45 1 16 36 45 3 16 34 45 5 Calcium Sodium Phosphorous Oxide Oxide Pentoxide Ga Glass composition (mol%) Glass code
♦Novel gallium-doped phosphate based glasses(PBGs) were prepared using conventional melt quenching method ♦Both density and glass transition temperature (Tg) was found to increase as the Ga content increased in the glass system ♦Degradation rate of glasses in water found to decrease with the increase of Ga content. pH was found to maintain at neutral for 3 and 5 mol% Ga doped PBGs but showed initial increase for 0 and 1 mol% Ga doped PBGs ♦Calcium and sodium ion release from the glasses were analysed using IC ♦Gallium and phosphorous release from the glasses were analysed using ICP-MS ♦Antimicrobial activity of these glasses against E. coli, P. aeruginosa and S. aureus were tested by disc diffusion assay ♦The antibacterial effect was found to decrease as the Ga content increased in the glass system, with
Ca16Na38P45Ga1 showing maximum bactericidal effect against E. coli, P. aeruginosa and S. aureus
Density and glass transition temperatures as a function of gallium oxide content (mol%)
2.5 2.55 2.6 2.65 2.7 2.75 2.8 0.5 1 1.5 2 2.5 3 3.5 4 4.5 5 Gallium Oxide Content (mol%) Density (g.cm-3) 335 340 345 350 355 360 365 370 375 Glass Transition Temperature ( 0C) Density Tg
Dissolution and pH analysis of the 0, 1, 3 and 5 mol% Ga-doped PBGs.
(a) Weight loss vs. time
y = 0.0417x R2 = 0.9792 y = 0.0236x R2 = 0.9866 y = 0.0073x R2 = 0.9851 y = 0.0037x R2 = 0.995 1 2 3 4 5 6 24 48 72 96 120 144 Time(hrs) Weight loss (mg.mm
2)
Ga0 Ga1 Ga3 Ga5 6 7 8 9 20 40 60 80 100 120 Time(hrs) pH Ga0 Ga1 Ga3 Ga5
pH vs.time
Cumulative ion release profiles of the 0, 1, 3 and 5 mol% Ga-doped PBGs
(a) calcium ion release vs. time (b) sodium ion release vs. time (c) phosphorous ion release vs. time and (d) gallium ion release vs. time
y = 2.7145x R2 = 0.936 y = 1.3007x R2 = 0.9531 y = 0.1765x R2 = 0.1549 y = 0.2764x R2 = 0.9477
50 100 150 200 250 24 48 72 96 120 144 Time(hrs) Na ion release (ppm) 1 3 5
y = 0.4049x R2 = 0.9843 y = 0.0983x R2 = 0.9167 y = 0.063x R2 = 0.8143 y = 0.7058x R2 = 0.9555
10 20 30 40 50 60 24 48 72 96 120 144 Time(hrs) Ca ion release (ppm) 1 3 5
y = 50.905x R2 = 0.9489 y = 35.016x R2 = 0.9624 y = 6.0693x R2 = 0.9217 y = 1.2474x R2 = 0.9974
500 1000 1500 2000 2500 3000 3500 4000 4500 5000 24 48 72 96 120 144 Time(hrs) P ion release (ppm) 1 3 5
y = 0.3935x R2 = 0.8336 y = 0.2334x R2 = 0.9313 y = 0.1143x R2 = 0.9973 y = 0.0024x R2 = 0.9798
10 20 30 40 50 60 24 48 72 96 120 144 Time(hrs) Ga ion release (ppm) 1 3 5
(a) (b) (c) (d)
Disc diffusion assay conducted on 0, 1, 3 and 5 mol% Ga-doped PBGs against Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus (MRSA) and Clostridium difficile
5 10 15 20 25 30 1 3 5 Ga (mol%) Diameter (mm) E.coli P.aeruginosa S.aureus MRSA C.difficle
The effect of 0, 1 and 3 mol% Ga-doped PBGs
Pseudomonas aeruginosa.
7 7.25 7.5 7.75 8 8.25 4 12 Time(hrs) Log CFU/mm2 control C16Ga0 C16Ga1 C16Ga3
Ca30Na20P50 Ca30Na17P50Ag3 Ca30Na15P50Ag5 30 20 50 30 17 50 3 30 15 50 5 Calcium Sodium Phosphorous Oxide Oxide Pentoxide Ag Glass composition (mol%) Glass code
♦P. aeruginosa causes a range of biofilm-associated infections such as Cystic fibrosis ♦Phosphate Based Glasses doped with silver were used in the CDFF to study their effect on these biofilms ♦Biofilms growth were analyzed using scanning electron microscope (SEM) ♦The effect of silver on the biofilms were tested by counting P. aeruginosa colony-forming units (CFUs) at different time ♦ ‘Live and dead’ cell distribution in the biofilm was determined by confocal laser scanning microscope (CLSM).
Constant Depth Film Fermentor (CDFF)
4 5 6 7 8 25 50 75 100 125 150 Time (hours) Log (CFU.mm
HA Ag- 5 mol% Ag
Log10 CFU/mm2 of P. aeruginosa in biofilms formed on hydroxyapatite discs (HA), Ca30Na20P50 disc (Ag-), and Ca30Na15P50Ag5 discs (5 mol% Ag).
Crystal structure of hydroxyapatite
a b c
Ca10(PO4)6(OH)2
P21/b Ca P O H Ca(1) Ca(2) HO- columns PO4
3-
O1 O1 O2 O1 O2 O2 O3 O3 O3 OH O3’ O3 O3’ O1 O2 O3
Ca(1) site Ca(2) site
1- Mg-HA
Synthesis and characterisation of Mg-substituted HA
Ca(NO3)2.6H2O + Mg(NO3)2.4H2O + NH4H2PO4
1/ pH ~ 10.0, 100˚C, 5h 2/ centrifugation / washing 3/ 100˚C drying under vacuum, 12h
“Ca10-xMgx(PO4)6(OH)2”
x = 0, 0.25, 0.5, 0.75, 1.0, 1.25, 1.5, 2.0
Elemental analyses, SEM, XRD, TG, DTA, IR, Raman, SSA measurements High-temperature XRD Catalytic studies Solid State NMR (31P, 1H, 43Ca, 25Mg )
O1 O1 O2 O1 O2 O2 O3 O3 O3
Ca(1) site
OH O3’ O3 O3’ O1 O2 O3
Ca(2) site
Mg2+ ? 2- Mg-HA
700 750 800 850 900 950 1000 1050 1100 2.5 5 7.5 10 12.5 15 17.5 20 22.5
x(Mg) (%) T (˚C) DTA XRD
Temperature of formation of whitlockite depends on Mg-content Indication of the site of incorporation of Mg ?
High-temperature XRD characterisation
Mg seems to have been incorporated in Ca(1) site
O1 O1 O2 O1 O2 O2 O3 O3 O3
Ca(1) site
OH O3’ O3 O3’ O1 O2 O3
Ca(2) site
3- Mg-HA
C CH3 C H3C OH CH methylbutynol (MBOH) H3C C H3C O acetone HC CH acetylene
alkaline surface +
Mg-HA
Catalytic properties of Mg-substituted HA
In Mg-HA, the reactivity of the OH group is expected to change if Mg enters the Ca(2) site No clear correlation between the catalytic activity and the % of Mg in the lattice Reaction catalysed by alkaline surfaces (presence of OH groups), HA have been shown to catalyse this reaction
OH O3’ O3 O3’ O1 O2 O3
Confirmation that Mg has been incorporated in Ca(1) site
4- Mg-HA
43Ca solid-state NMR of Ca10(PO4)6(OH)2 Isotope Spin Natural abundance(%) Quadrupole moment Q Relative receptivity
43Ca
7/2 0.135
4.19*10-5
fwhm = 963 Hz fwhm = 1042 Hz
5- Mg-HA Quadrupolar interaction main cause of line-broadening
8.45 T 4 kHz, D1 = 1s RAPT-1pulse NS = 180 000, LB = 80 14.1 T 4 kHz, D1 = 1s RAPT-1pulse NS = 81 000, LB = 80 18.75 T 4 kHz, D1 = 0.1s RAPT-1pulse NS = 1197824, LB = 80
ppm
50 100 150 200
O1 O1 O2 O1 O2 O2 O3 O3 O3 O1 O1 O2 O1 O2 O2 O3 O3 O3
OH O3’ O3 O3’ O1 O2 O3
Ca(2) Ca(1)
Shorter Ca-O average distance Smaller coordination number 6 Calcium atoms Longer Ca-O average distance Larger coordination number 4 Calcium atoms
43Ca solid-state NMR of HA
6- Mg-HA
δiso = 10.6 ppm Cq = 2.7 kHz ηq = 0.65 δiso = -3.4 ppm Cq = 2.5 kHz ηq = 0.35
(ppm)
50 100 150 200 (ppm)
50 100 150 200
18.75 T 14.1 T
43Ca isotopic labelling of non-substituted HA planned
to confirm the proposed distinction between both sites.
43Ca solid-state NMR of
Ca10-xMgx(PO4)6(OH)2 (x ~ 1.0)
Difficult to conclude on the site of incorporation of Mg with 43Ca NMR at this stage go to higher fields. HA Mg-HA 7- Mg-HA
Change in lineshape: Decrease at the higher frequencies 14.1 T 4 kHz RAPT-1pulse
8- Bone
Hierarchal structure of bone
Whole bone Fibril arrays Mineralized collagen fibrils HA crystals Several levels
3 main components:
(carbonated HA)
(collagen)
Non collageneous proteins
Weiner, Annu. Rev. Mater. Sci. 1998, 28, 271
9- Bone
Structural studies on bones
SEM (+ TEM) IR Solid state NMR
1H NMR: Signals of water and
31P NMR: Mineral phase (mainly apatite) 13C NMR: Organic phase (mainly collagen)
EDXS Presence of C, O, N, Ca, P mainly Ca/P ratio bigger than in HA
ppm
10 20 30 40
10- Bone
Towards 43Ca NMR studies of bones
Calcium in bones Model compounds
Mineral phase:
2- anions
Mineral phase:
Organic environment:
Organic environment:
Can 43Ca solid-state NMR give insight into the different environments of Ca2+ in bones ?
11- Bone
43Ca NMR: bone vs hydroxyapatite
2 2 2 2 2
csd 2 1 2 q 2 2 1 2 2 csd q 1
W B B W B B ) (LW W W ) (LW ⎟ ⎠ ⎞ ⎜ ⎝ ⎛ + ⎟ ⎠ ⎞ ⎜ ⎝ ⎛ = + =
Stronger chemical-shift dispersion in bone Wq = 806 Hz, Wcsd = 718 Hz Wq = 944 Hz, Wcsd = 458 Hz Multiple-field analysis Second order quadrupolar broadening (Wq) vs Chemical Shift dispersion (Wcsd)
Bone Apatite
12- Bone
43Ca NMR studies of bones and related compounds
Warwick University 14.1 T 4 kHz RAPT-1pulse
Perspectives
Bone Mg-HA
43Ca NMR
Ca-phosphoproteins: find a model compound Ca-phosphates: more compounds to analyze (collaboration with C. Gervais, UPMC, Paris) Bone: try higher magnetic field
Complementary measurements:
XRD Ba-HA, Aldrich-HA Rietveld studies on Mg-HA and Ba-HA ? TEM + EDXS Catalysis: Results to confirm on another Ba-HA
Further studies to perform Write first paper on synthesis and characterisations (other than NMR)
(ppm)
31P MAS NMR of (CaO)0.4(Na2O)0.1(P2O5)0.5
→ XRD shows the presence of NaCa(PO3)3 and Ca(PO3)3
31P Refocused INADEQUATE
Utilises the J-coupling through-bond interaction → Identifies phosphorous sites directly linked by P-O-P bonds Dipolar coupling removed by MAS*
(ppm)
Ca(PO3)2 NaCa(PO3)3 ???
23Na MAS NMR
Isotropic chemical shift = (4.5 ± 0.5) ppm Quadrupolar coupling constant = (2.10 ± 0.05) MHz Asymmetry parameter = 0.95 ± 0.02
23Na Multiple Quantum MAS
Refocuses 2nd order quadrupolar interaction for the central transition 1st order quadrupolar interaction is removed by MAS → Obtain a spectrum free of quadrupolar broadening in the second dimension → Overlapping lines can be resolved unambiguously
23Na 3QMAS of (ZnO)0.1(CaO)0.3(Na2O)0.1(P2O5)0.5
31P refocused INADEQUATE allows identification of phosphate chains 23Na MQMAS allows different sodium-containing phases to be observed
Short paper on 31P INADEQUATE results What is the effect of adding the ZnO?
63Cu NMR
Supervisor: Dr. Julian Jones Sponsor: NovaThera Ltd.
Department of Material Imperial College London
TEM TEM SEM AFM TEM SEM SEM SAXS WAXS RAMAN NMR Orcel et al. and Himmel et al. FIB
Heat Treatment Effects on Surface Area
0.00 20.00 40.00 60.00 80.00 100.00 120.00 140.00 160.00 180.00 200.00 Before Stabilisation After Stabilisation After 2hr Sintering Surface Area m2/g
Heat Treatment Effects on Modal Pore Size
0.00 2.00 4.00 6.00 8.00 10.00 12.00 14.00 16.00 18.00 20.00 Before Stabilisation After Stabilisation After 2hr Sintering M
Nitrogen Adsorption Porosity Analysis
Heat Treatment Effects on Pore Volume
0.0000 0.1000 0.2000 0.3000 0.4000 0.5000 0.6000 0.7000 0.8000 Before Stabilisation After Stabilisation After 2hr Sintering Pore Volume cc/g
Nitrate Decomposition
Calcium Nitrate Decomposition
Heat Treatment Effects on Surface Area
0.00 100.00 200.00 300.00 400.00 500.00 600.00 700.00 800.00 900.00 Before Stabilisation After Stabilisation After 2hr Sintering Surface Area m2/g 70S30C 100S
Heat Treatment Effects on Total Pore Volume
0.0000 0.1000 0.2000 0.3000 0.4000 0.5000 0.6000 0.7000 0.8000 Before Stabilisation After Stabilisation After 2hr Sintering Total Pore Volume cc/g 70S30C 100S
Silica Ca(NO3)2
Calcium Nitrate Crystalline before Stabilisation
XRD Pattern of 70S30C without Stabilisation
200 400 600 800 1000 1200 1400 1600 1800 2000 5 15 25 35 45 55 65 75 85 95 2θ /° In te n sity
Crystalline Lattice Pattern before Stabilisation under TEM (scale bar 5 nm)
No Striking Peaks
Nominal composition
Glass code P2O5 (mol%) CaO (mol%) Na2O (mol%) Fe2O3 (mol%) S_Fe_1 50 30 19 1 S_Fe_2 50 30 18 2 S_Fe_3 50 30 17 3 S_Fe_4 50 30 16 4 S_Fe_5 50 30 15 5
Fe K-edge XANES 7100 7120 7140 7160 7180 7200 7220
1
FeO Fe3O4 Fe2O3 S_Fe_1 S_Fe_2 S_Fe_3 S_Fe_4 S_Fe_5 Fe2 (SO4)3
Fe pre-edge feature 7108 7112 7116 7120
0.00 0.03
FeO Fe2O3 S_Fe_2 S_Fe_5 Fe2 (SO4)3
Fe coordination information
7112.5 7113.0 7113.5 7114.0 7114.5 0.0 0.1 0.2 0.3 0.4
: Fe(III) in Td coordination : Fe(III) in C3V coordination : Fe(III) in Oh coordination : Fe(III) in Oh coordination measured in this study : iron-phosphate glasses samples
Fe EXAFS
5 6 7 8 9 20 40 60 80
k/Å
k
3χ(k)
2 4 6 8 10 30 60 90 120
r /Å FTk
3χ(k)
From top to bottom: S_Fe_1, S_Fe_2, S_Fe_3, S_Fe_4, S_Fe_5 glasses: k3 weighted EXAFS (left) and Fourier transform (right). Slide lines: experimental data; dashed lines: theoretical simulation. Averaging Fe-O distance: 1.95 Å Averaging Fe…P separation:3.19 Å
Write paper based on this work Beam time next week on Gallium phosphate glasses Work on incorporating calcium chelating agent into phosphate-silicate network (samples are ready for characterization)
Fe is in Fe(III) oxidation state Fe is located in a distorted octahedral coordinated oxygen cage Fe is incorporated in the phosphate network
Neutron Diffraction
1 2 3 4 5 2 4 6 8
T(r) / barns Å
r / Å
D M Pickup, I Ahmed, P Guerry, JC Knowles, M E Smith and R J Newport, The structure of phosphate glass biomaterials from neutron diffraction and 31P nuclear magnetic resonance data. Journal of Physics: Condensed Matter, in press.
X-ray Diffraction
Correlation R / Å (±0.2) N (±0.5) P-O 1.6 4 Ca-O 2.3 5 O-O 2.5 4.5 Ca-O 3 1 P-P 3 2 O-O 3.5 8.5 P-Ca 3.6 5 Ca-Ca 3.7 2 5 10 15 20
0.0 0.2 0.4 10 20 30 40
0.0 0.1
X-ray Diffraction RMC Q / Å
i(Q) / atoms barn
Q / Å
Neutron Diffraction RMC
Neutron Diffraction
1 2 3 4 5 0.0 0.5 1.0 1.5 2.0
T(r) / barns Å
r / Å
1 2 3 4 5 2 4 6 8
T(r) / barns Å
r / Å
X-ray Diffraction
Pickup, D.M., et al., The structure of phosphate glass biomaterials from neutron diffraction and 31P nuclear magnetic resonance data. Journal of Physics: Condensed Matter, in press.
Differentiated two Ca-O bond lengths, a shorter distance of ~2.34 Å has a coordination of ~5.4 and a longer distance of ~2.80 Å has a coordination of ~1.4. The total Ca-O coordination is ~6.8. This coordination relates to the crystal structure of a capped trigonal prism. Onb coordination with Ca is ~1.7, therefore for every Onb that is only bonded to one Ca there are eight that are bonded to two Ca . From the correlation distance an O-Ca-O bond angle between 72° and 89° is calculated which indicates a distorted octahedral structure – close to the capped trigonal prism indicated.
Kent ● Warwick ● Imperial ● UCL
Zr K-edge EXAFS of Zr phosphate glass
1 2 3 4 5 6 7 8 9 10 10 20 30 40 50 60 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
2 4 6 8 10 12 14
FT[chi(k)] Experimental Theory r (ang) k
3 chi(k)
Experimental Theory k (ang-1)
xLi2O:2ZrO2:40P2O5
x=56 x=58 x=56 x=58
Correlation R (Å) N DWF ± 0.02 ± 20% ± 0.005 Sample 56 Zr-O 2.06 5.6 0.010 Zr-P 3.64 1.7 0.010 Zr-O 4.63 2.1 0.005 Sample 58 Zr-O 2.06 5.9 0.011 Zr-P 3.64 2.3 0.008 Zr-O 4.63 2.5 0.006
Zr-O, -P agreement
Correlation R (Å) N DWF Zr-O 2.06 5.6 0.011 Zr-P 3.64 1.7 0.010
Zr(P2O7) Zr-O average distance is 2.03 Å with a coordination number of 6. Zr-P average distance is 3.49 Å with a coordination number of 6. LiZr2(PO4)3 Zr-O average distance is 2.06 & 2.07 Å with a coordination numbers of 6. Zr-P average distance is 3.46 & 3.47 Å with a coordination numbers of 6.
Zr-BO, -NBO distances
Zr P NBO BO NBO BO NBO 1.5 2.1 1.6 4.4
109°
4.3 Numbers shown in BLUE are found by ND of phosphate based glasses. The number in RED is given by the first peak in the Zr EXAFS. Those in GREEN and ORANGE are calculated and agree well with the second Zr-O coordination sphere.
Correlation R (Å) N DWF Zr-O 4.06 5.7 0.033 Zr-O 4.64 4.0 0.012