Contribution (Oral/Keynote)
Towards new orientations in the research and development of biomimetic apatite nanosystems
Christophe Drouet1,Ahmed Al-Kattan1, Sabrina Rollin-Martinet1,2, David Grossin1, Eric Champion2,
Fabrice Rossignol2, Chantal Damia2, Pascal Dufour1, Jeannette Dexpert-Ghys3, Veronique Santran4,
Christèle Combes1, Christian Rey1
1CIRIMAT
Carnot Institute, University of Toulouse, France
University of Limoges, France
3CEMES, University of Toulouse, France
4ICELLTIS, Verniolle, France
2SPCTS,
christophe.drouet@ensiacet.fr
INTRODUCTION
The development of bio-inspired systems is an increasingly appealing field of research, especially for
setting-up advanced biomaterials exhibiting physico-chemical characteristics and/or biological
properties as close as possible to those of natural systems. A typical example is that of biomimetic
nanocrystalline calcium phosphate apatites (nanoAp) that mimic bone mineral [1]. In contrast to
stoichiometric hydroxyapatite (HA), such biomimetic nanoAp were shown to exhibit, like biological
apatites, a non-apatitic phospho-calcic hydrated layer on the surface of the nanocrystals which confers
to these materials an increased bioactivity by facilitating ionic surface exchanges (with ions from
surrounding fluids) or adsorption processes. In this context, these last years have witnessed several
cutting-edge progresses in the field of biomimetic apatite bio-oriented applications. These recent
evolutions originated in particular from i) a more precise physico-chemical characterization of
nanocrystalline apatitic materials and ii) from the development of non-conventional processing
approaches. In this contribution we illustrate two examples of such recent advances of nanoAp-based
systems, that may widen their already-existing domains of applications. The first topic is relative to the
possibility to consolidate biomimetic apatite compounds by Spark Plasma Sintering at “low”
temperatures (typically ≤150°C) while retaining the hydrated and nanocrystalline characters of the initial
powders, thus enabling one to prepare advanced bioceramics still composed of biomimetic apatite
nanocrystals in view of enhanced bone repair. In contrast, the second example deals with the
preparation and evaluation of colloidal 30-to-100nm nanoparticles associating lanthanide-doped apatite
nanocrystals and a phospholipid moiety in view intracellular medical imaging.
EXPERIMENTAL
Preparation of samples for SPS consolidation in view of advanced bone tissue engineering:
In view of SPS consolidation, nanocrystalline apatite powders were precipitated at RT and pH 7.2 from
solutions of calcium nitrate (0.3M) and ammonium hydrogenphosphate (0.6M). The precipitates were
left to mature between 20 min and 3 weeks), and then filtered, washed and freeze-dried. Spark Plasma
Sintering (SPS) consolidation was then undergone. The non-conventional heating source (electrical
current crossing a conductive matrix containing the powder to consolidate) enables fast heating and
cooling rates aiming at limiting crystal growth and dehydration phenomena. SPS was processed at
150°C, under a mechanical pressure of 100MPa, in 1 atm argon, for 20 min.
Preparation of biomimetic apatite-based colloidal nanoparticles for medical imaging:
The preparation of apatite-based colloids, eventually doped with lanthanide luminescent elements, was
carried out by coprecipitation from calcium (± lanthanide) nitrate(s), ammonium hydrogenphosphate and
in the presence of a biocompatible phospholipid moiety: 2-amino-ethylphosphate denoted “AEP”. The
molar ratio between AEP/(Ca + lanthanide) ratio was tested in the range 0.2-1. The (Ca + lanthanide)/P
Contribution (Oral/Keynote)
ratio was fixed to 3. Lanthanide doping varied from 0 to 2 at.% relative to Ca. A maturation stage at
100°C for 16 h. was realized for the obtainment of fluid colloids, purified by dialysis. The cytotoxicity of
the colloidal nanoparticles was tested by MTT tests on human stem cells from the adipose tissue
(AMSC) and on ZR-75-1 breast cancer cells. Cell internalization was checked using ICP-AES titrations.
Several complementary techniques (XRD, FTIR/Raman, MET, DLS, luminescence…) were used for
characterizing the samples prepared in this study, either as powders, consolidated ceramics or colloids.
RESULTS AND DISCUSSION
The analysis of SPS-consolidated biomimetic-apatite bioceramics showed that the hydrated and
nanocrystalline characters of the initial powders were beneficially retained thanks to the use of the SPS
non-conventional sintering technique enabling fast heating and cooling rates. Also, we proved that the
non-apatitic hydrated layer initially present on the surface of biomimetic apatite nanocrystals was
essentially conserved after SPS treatment at 150°C and 100MPa, thus enabling one to prepare
potentially highly-bioactive and resorbable bioceramics, in contrast to systems obtained from hightemperature sintering of stoichiometric hydroxyapatite.
In another application field, we showed that luminescent lanthanide (typically Eu or Tb)-doped apatite
colloids synthesized with AEP as biocompatible stabilizing agent could be prepared by soft chemistry,
that they exhibited a low cytotoxicity, and that the nanoparticles features (size (Fig. 1), chemistry…)
were favorable for an internalization for example by ZR-75-1 breast cancer cells, thus enabling to
foresee promising perspectives in the diagnosis of cancers by way of medical imaging.
CONCLUSION
The possibility to tailor the chemical composition and (non)stoichiometry of the nanocrystalline
biomimetic apatites, and eventually the nature and amount of grafted molecules on the surface of the
nanocrystals, enables one to foresee new orientations, in particular for the setup of a new generation of
highly-bioactive biomimetic ceramics for advanced bone regeneration… or for the preparation of
nanosystems in medicine, for example for medical imaging.
References
[1] C. REY, C. COMBES, C. DROUET, M. J. GLIMCHER, Osteoporosis Internaitonal, 20 (2009) 1013.
Acknowledgement
The Authors acknowledge the ANR Agency for providing financial support to the “NanoBiocer” project.
Figure
AEP
Eu3+
Eu-doped apatite nanoparticle
Colloid
Luminescent 30-nm biomimetic apatite-based colloidal nanoparticles for medical imaging