Biomineralization
Biomineralization,,
biomimetic
biomimetic and
and non
non-classical
classical crystallization
crystallization
Ways
Waysto
tounderstand
understandthe
thesynthesis
synthesisand
and
formation
formationmechanisms
mechanismsof
ofcomplex
complexmaterials
materials
Helmut Cölfen
Max-Planck-Institute of Colloids & Interfaces, Colloid Chemistry,
Research Campus Golm, D-14424 Potsdam
Coelfen@mpikg-golm.mpg.de
CaCO
CaCO33 Biominerals
Biominerals
Coccolith (Calcite)
Nacre (Aragonite)
Herdmania momus
(Vaterite)
Characteristics of Biominerals:
Complex forms generated from inorganic systems
with simple structure
Characteristics of inorganic crystals:
Simple geometrical form but often complicated
crystal structure
Default: Rhombohedral Calcite
Important
Important biominerals
biominerals
Mineral
Formula
Calciumcarbonate
Calcite
CaCO3

Algae / Exoskeleton
Birds / Eggshell
Fishes / Gravity sensor
Mussels / Exoskeleton
Sea urchins / Spikes
Aragonite
Vaterite
Calciumphosphate
Hydroxyapatite
Octa-Calciumphosphate
Function


Silica
Iron oxide
Magnetite
Ca10(PO4)6(OH)2 Vertebrates / Skeleton, teeth
Ca8H2(PO4)6
Vertebrates / Precursor for
bone formation
SiO2
Algae / Exoskeleton
Fe3O4
Salmon, Tuna & bacteria /
Magnetic field sensor
Chitons / teeth

Evolution
Evolution of
of aa mussel
mussel shell
shell
SEM of broken mussel shell
TEM thin cut of
developing mussel
shell
SEM of broken mussel shell
after protein removal
SEM of developing
mussel shell
SEM of broken mussel
shell after CaCO3 removal
SEM mature mussel shell
Biominerals are often formed in confined reaction environments
Epitactical
Epitactical match
match between
between crystal
crystal and
and matrix
matrix
Nacre:
Periodicity in
protein β-sheets
and β-chitin fibers
in close geometrical match to
lattice spacing of
aragonite 001
face.
„Soft Epitaxy“
Bone
Bone formation
formation
Sub-nm
regime
- X - Y - GLY - X - Y - GLY - X - Y - GLY - X - Y -
Amino acid sequence
(primary structure)
left handed α-helix
(secondary structure)
0,286 nm
2,86 nm
right handed triple helix
(tertiary structure)
Rod-like
tropocollagen
nm regime
300 nm
Stained collagen fibril
as seen in TEM
300 nm
Characteristics:
40 nm 27 nm
Collagen fibril
(quatery structure)
μm regime
67 nm
Mineralization of oriented plate-like apatite
crystals starting in the hole regions of the
collagen fibrils. In (A), plate-shaped crystals
are arranged in linear coplanar arrays forming
grooves through the fibrils. In (B) these grooves
are shown to be separated by four layers of
triple helical collagen molecules. Figure not
drawn to scale.
mm - m regime
Sequence of progressive bone development. Osteoblasts (OB) oriented with their backs toward the capillary
vasculature (C), secrete osteoid (O) away from the vasculature, causing the formation of a bony strut (B), and
eventually forming a second layer of bone (B2). The stacked layer (SC), which provides osteoprogenitor cells
for the process, continues to expand in the direction of bone growth.
Mineralization
occurs in organic
matrix
Superstructure
formation over
several length
scales
Here over 9
magnitudes in
length !
From almost atomic to
macroscopic scale
Tooth
Tooth enamel
enamel
Enamel
Growth
direction
•
•
•
Tomes process
Crystallization and assembly of fibers
into rods on the μm scale
Ordered layer structure of rods provides
mechanical strength
Permanent further crystallization and
density increase up to 95 wt.-% mineral
Crystallization often occurs highly directed
Magnetotactic
Magnetotactic bacteria
bacteria
Nucleation on inside
wall of vesicle
Linear organization for
magnetic compass
10 nm
Phase
Phasetransitions
transitionsand
andmatrix
matrixassisted
assistednucleation
nucleation
Adapted from S.Mann IMPRS lectures, Golm 2003
Sequence of kinetic inhibitions and phase
transformations, resulting in high selectivity
degree in crystal structure and composition
Direct mechanisms to increase S:
Ion pumping + redox
Ion complexation/decomplexation
Enzymatic regulation
Indirect mechanisms to increase S:
Ion transport
Water extrusion
Proton pumping
General
General biomineralization
biomineralization features
features
•
•
•
•
•
•
•
•
Uniform particle size
Well defined structure and composition
High levels of spatial organization
Complex morphologies
Controlled aggregation and texture
Preferential crystallographic orientation
Higher order assembly
Hierarchical structures
Adapted from S.Mann IMPRS lectures, Golm 2003
Known
Known mechanisms
mechanisms of
of Biomineralization
Biomineralization
•
•
•
Stabilization
Morphology control by selective adsorption
Control of the crystal modification by
„Soft Epitaxy“
•
•
•
•
•
Static templates
Confined reaction environments
Adaptive construction and synergistic effects
Structural reconstruction
Higher order assembly
Polymer structures as static templates
PS-PAA Gel + Fe3O4
Control
Fe2+- loaded
Magnetic gel
Biomineral
Biomimetics
Mollusc tooth
Homogeneous 38 nm
Zoom
1 μm
Fe3O4 15 - 35 nm
in Polysaccharide Protein Gel
Templated 17 nm
M. Breulmann, H.P. Hentze
Double hydrophilic block copolymers
Molecular tool:
Handle
Head
= hydrophilic, interacting with mineral
= hydrophilic, non interacting with mineral
Amphiphilic behaviour induced in
presence of mineralic surfaces
allows
- Size/shape control
- Stabilization
Advantage
Advantageof
ofDHBC
DHBCdesign
designfor
forCaCO
CaCO33
PMAA
PE block too long, PE block and stab.
stab. block too short block good ratio
PEO-b-PMAA
PE block too short, PE block far too short,
stab. block too long stab. block too long
Double hydrophilic block copolymers
Precursor Polymers
Functionalities
OH
COOH
PEG-b-PEI
(Linear and branched)
PEG-b-PMAA
PEG-b-PB
Modular Synthesis
M = 3000 – 10000 g/mol
SO3H
PO3H2
PO4H2
CH3SCN
NR3, HNR2, H2NR
Hydrophobic
M. Sedlak, M. Breulmann, J. Rudloff, P. Kasparova, S. Wohlrab, T.X. Wang
BaSO
BaSO44 Morphogenesis
Morphogenesis at
at pH
pH 55
2μm
0.5μm
PEG-b-PEI-SO3H
PEG-b-PMAA-Asp
0.5μm
2μm
No additive
2μm
PEG-b-PEI-COOH
L.M. Qi
PEG-b-PMAA-PO3H2
CdS
-b-PEIbranched
CdS ++ PEO
PEO-b-PEI
branched
L.M. Qi
•
•
•
•
Particle size adjustable 2 - 4 nm
Monodisperse stable particles
No photooxidation
Branched PEI more effective than
linear or dendrimer
2.0
1.6
Absorbance
1.4
PL Intensity (a.u.)
0.05 g/l
0.10 g/l
0.25 g/l
0.50 g/l
1.00 g/l
1.8
1.2
1.0
0.8
0.6
0.05 g/l
0.10 g/l
0.25 g/l
0.50 g/l
1.00 g/l
0.4
0.2
0.0
250
300
350
400
450
Wavelength (nm)
500
550
400
450
500
550
Wavelength (nm)
600
650
Selective
Selective Adsorption
Adsorption
Gold crystallized in presence of PEG-b-1,4,7,10,13,16-Hexaazacycloocatadecan
(Hexacyclen) EI macrocycle
Interference patterns
due to deformations in
sub-Angström range
Very thin platelets
200 nm
Very thin platelets with
exposed 111 surface,
Well developed plasmon
band in UV/Vis
Au (111) surface and
adsorbed hexacyclen
molecule in vacuum
S.H. Yu
Surfaces as static template
CaCO3 +
O
m
n
OPO(OH)2
PEG(m)-b-PHEE(n)-(PG rad%)
J. Rudloff
CaCO 3 (s) + CO 2 (g) + H 2 O (l)
50 μm
50 μm
Ca 2+ (aq) + 2 HCO 3 -(aq)
50 μm
Structural
Structural reconstruction
reconstruction
pH = 5
BaSO4 with
PEG-b-PMAA-PO3H2
Parallel cut to to fiber axis Perpendicular cut to fiber axis
Diameter 20 - 30 nm
Fiber axis is [210]
Bundles of
single
crystalline
fibers
L.M. Qi
Structural
Structural reconstruction
reconstruction
pH = 5
•
•
BaSO4 fibers obtained in the presence of PEG-b-PMAA-PO3H2
on carbon films with an aging time of 5d.
Heterogeneous nucleation on carbon films
Perfectly flat surface of growth edge due to
surface minimization
L.M. Qi
Crystallographically oriented nanoparticle attachment
Single crystalline anatase TiO2
particles hydrothermally
synthesized in 0.001 M HCl
Topotactic particle attachment
at high energy surfaces (112)
a) Single primary crystal,
b) four primary crystals
forming a single crystal via
oriented attachment,
c) Single crystalline anatase
particle
R.L. Penn, J.F. Banfield; Geochim. Cosmochim. Acta. 63 (1999) 1549 - 1557
1)
2)
3)
L.M. Qi
Nucleation of
amorphous
nanoparticles,
polymer
stabilization
5)
4)
-
Crystallization of
amorphous
particles, directed
particle fusion
6)
7)
„ Perfect Single Crystals Fiber Bundles: Elongation along [210]
]
10
[2
500nm
<001>
400
410
420
220
210
200
4-10 4 -20
2-10
2-20
0-20
020
-220
-210 -200
-2-10 -2-20
-4 -20
-400 -4-10
XY
projection
S.H. Yu
Structural
Structural reconstruction
reconstruction
10 μm
BaSO4, 2 mM,
polyacrylate ( Mn = 5100)
0.11mM, pH = 5.5, 25
°C
1 μm
Cuttlefish bone β-Chitin +
aragonite
100 μm
2 μm
S.H. Yu
Higher order assembly
30 min
60 min
100 nm
100 nm
90 min
120 min
[PEG-b-PMAA] = 1 g/l,
[CaCO3] = 8 mM,
pH = 10
•
Chains of
aggregated
amorphous
nanoparticles
•
Dumbbell
shaped and
spherical
aggregates
•
Nanoparticle
crystallization
Primary crystals 40 nm
10 μm
10 μm
Zoom
L.M. Qi
Light microscopy on CaCO3 with PEO-b-PMAA
5.6
8
8-15 min
19-30 min
45-90 min
3.8
7
6
3.2
Images not
drawn to scale
5
l/r
2.7
4
1.8
pH 6.5
3
1.4
2
1
0
2
4
6
8
10
12
r/µm
A. Reinecke, H.G. Döbereiner
14
16
Higher order assembly
Aggregate structures are also observed if the crystals are
dissoved with HCl
A. Reinecke, H.G. Döbereiner
Higher
Higher order
order assembly
assembly
pH
11
L.M. Qi
10
9
[Polymer] / [CaCO3]
••
Supersaturation
-mineral interaction
pH) and
Supersaturationand
andstrength
strengthof
ofpolymer
polymer-mineral
interaction((pH)
and
[Polymer]
[Polymer]//[CaCO
[CaCO3]]play
playimportant
importantrole
rolefor
formorphogenesis
morphogenesis
3
Higher order assembly
CaCO3 +
PEG-b-PEDTA-C17H35
CaCO3 in oyster marrow matrix
P. Harting 1873
Composed of nanoparticles
M. Sedlak
Higher
Higher order
order assembly
assembly
L.M. Qi
pH 11
pH 7
pH 9
pH 5
pH 3
BaSO4 with PEG-b-PMAA additive; pH variation
BaCO3 Morphogenesis
BaCO3 + PEG-b-PMAA (1g/L), Ba2+ = 10 mM,
gas-diffusion reaction, 2 days
Different growth stages from rods (1),
growth at the ends (2), via dumbbells (3) to
spheres (4).
2 μm
Apparently no directed
nanoparticle assembly but
dendritic radial outgrowth.
Default
50 μm
3
3
1
4
3
1
2
3 μm
2
4
1 μm
S.H. Yu
Chirality introduction by a racemic polymer
S.H. Yu, K. Tauer
1 g L-1, pH = 4, [BaCl2] = 10 mM, on
glass slip, gas diffusion reaction
14 d
14 d
Amorphous
precursor after
5h
PEG-b-[(2-[4-Dihydroxy phosphoryl)-2-oxabutyl) acrylate
ethyl ester,
Mw = 120000 g/mol, PEO = 2000 g/mol
BaCO3 orthorhombic crystal structure
011 & 022 broadened
Restricted growth
3000
111
311
022
004
020
211
013
202
200
011
002
201
1000
Vacuum equilibrium
morphology
122
104
220
213
With
polymer
2000
102
Intensity (a.u.)
4000
Default
0
011
15
20
25
30
35
40
45
50
2θ (degree)
020
111
110 favourable for
polymer interaction
S.H. Yu, K. Tauer
Generation of unidirectional aggregation
Particles aggregate and
fuse predominately at 011
Aggregation /
growth direction
Favourable
for polymer
adsorption
Unfavourable
for polymer
adsorption
Polymer is sterically
demanding
S.H. Yu, K. Tauer
Possible
Possible
Impossible
Polymer adsorption on the timescale of aggregation
Chirality introduction by a racemic polymer
Overlay of
directed
aggregation
and turn
Alteration
between 020
and 011
aggregation
creates turn
S.H. Yu, K. Tauer
Control experiments on mechanism
Increase lateral growth
Decrease lateral growth
Particle attachment prevails
over polymer adsorption.
Polymer adsorption prevails
over particle attachment.
Polymer 1 g L-1, pH = 5.6, Less particle
charge as IEP = 10.0 – 10.5
Polymer 2 g L-1, pH = 4,
[BaCl2] = 10 mM
S.H. Yu, K. Tauer
Organic Mesocrystals, the D,L-Alanin example
Default
41 % functionalization
PEG-PEI
O
Supersaturated solution
at 65 °C cooled to 20 °C
n
cPolymer = 1 g/l
HO
PEG-b-PEI-s-But-COOH
200 μm
100 μm
S. Wohlrab
Organic Mesocrystals, the D,L-Alanin example
2 μm
2 μm
Crystalline appearance but:
•
•
•
Rough and even surfaces
Nonplanar surfaces
Cracks indicate swollen structure
S. Wohlrab
Partial polymer dissolution
after boiling in MeOH
indicates hybrid structure
D,L Alanin + 1 g/l PEG-b-PEI-s-But-COOH
Organic Mesocrystals, the D,L-Alanin example
Overview
Rough face
100 μm
2 μm
Fracture structure
2 μm
Single crystal
Powder
Mesocrystal
X-ray single crystal analysis
D,L Alanin + 10 g/l
PEG-b-PEI-s-But-COOH
S. Wohlrab
Different modes of cyrstal growth
Nucleation
clusters
Crystal growth
Primary
nanoparticles
> 3 nm
Temporary
Stabilization
Mesoscale
assembly
Mesocrystal
Amplification Mesoscale
assembly
Single crystal Iso-oriented
crystal
Organic Mesocrystals, the D,L-Alanin example
D,L Alanin + 1 g/l PEG-b-PEI-s-But-COOH + NaCl
0N
0.1 N
1.0 N
2.0 N
3.0 N
4.0 N
5.0 N
saturated
Salt addition dramatically changes the mesocrystal morphology
• Counterplay between electrostatic & vdW forces
• Polymers attach to different surfaces
• D,L Alanin molecule is already a dipole !
S. Wohlrab
Conclusions
Conclusions
•
Bio-inspired mineralization can transfer biomineralization
principles towards the synthesis of advanced organic-inorganic
hybrid materials at ambient temperature in water.
•
Self-assembled superstructures can be generated by tuneable
interacting organic block copolymer additives. But: Self
assembly only possible up to the micron scale (Need for
macroscopic templates)
•
Up to now, synergistic effects as often present in
biomineralization processes cannot yet be applied in bioinspired mineralization. Bio-inspired mineralization has still the
character of a model system.
•
Structure formation mechanisms are still often unknown due to
demanding analytics.
•
Principles can be extended to organic crystals exploiting new
variables like chirality or inherent molecular dipoles.
Bio-inspired mineralization offers a large playground for future materials
Conclusions
Conclusions
•
Bio- and biomimetic mineralization offer many indications
for non-classical crystallization routes e.g. a crystal is not
always built up from ions or molecules but by precursor
particles via mesoscopic transformations
•
Mesocrystals can be isolated for organic crystals with low
lattice energy, for the inorganic counterparts, the lattice
energy is too high and forces crystallite fusion with
resulting defect structures
•
Often, amorphous precursors are observed and
mesoscopic transformation generally plays an important
role in the final alignment of the crystallites.
•
Coding of crystal surfaces by additives can alter the
mesocrystal structure and thus the structure of the final
single crystal (additive inclusions)
Recommended
Recommended reading
reading
•
S. Mann, Biomineralization, Oxford University Press,
Oxford, 2001
•
S. Mann, J. Webb, R. J. P. Williams, Biomineralization:
Chemical and biochemical perspectives, VCH Weinheim,
1989
•
S. Weiner, CRC Critical Reviews in Biochemistry, 1986,
20, 365 - 408
•
H. Cölfen, S. Mann, Angew. Chem. Int. Edit. 2003, 42,
2350-2365
•
S.H. Yu, H. Cölfen, J. Mater. Chem. 2004 in press