C HARACTERIZATION OF S OLID L IPID N ANOPARTICLES Cécile Allais #, F. Artzner #, T. Narayanan §, T. Gulik-Krzywicki ‡, G. Keller # and M. Ollivon # # Equipe de Physico-Chimie des Systèmes Polyphasés, UMR CNRS 8612, Faculté de Pharmacie, 5 Rue Jean-Baptiste Clément Chatenay-Malabry Cedex, France § E.S.R.F., Grenoble, France ‡ Centre de Génétique Moléculaire, CNRS, Gif-sur-Yvette, France

Pharmaceutical Point of View: VECTORISATION of POORLY WATER-SOLUBLE DRUGS Active compounds:  Hydrophobicity  Crystallinity Characteristics :  Nanometric Size: ~100nm  Lipidic Matrix: non toxic  Solid Matrix: slow release System : - model of hydrophobic molecule = CHOLESTEROL - matrix lipid = COCOA BUTTER : mixture of triglycerides

? Physical Characteristics: - SHAPE ? - HOMOGENEITY ? - POLYMORPHISM ? Techniques Optic Microscopy X-ray Diffraction Electronic Microcopy S OLID L IPID N ANOPARTICLES 100< d < 300 nm Low concentration of lipids: < 1% (w/w)

Cocoa Butter/Cholesterol Nanoparticles O PTIC M ICROSCOPY IN D ARK F IELD Light scattering h

O PTIC M ICROSCOPY IN D ARK F IELD Nanoparticles just After Preparation at Room Temperature Nanoparticles Stocked 1 Month at 4°C Non Spherical Particle Spherical Particle h

100 nm Presence of Flat and Hemispherical Sides E LECTRONIC M ICROSCOPY (T. Gulik-Krzywicki) 100 nm

C OCOA B UTTER P OLYMORPHISM IN N ANOPARTICLES (SAXS on ID2 Beamline, E.S.R.F., T. Narayanan) Cocoa Butter Nanoparticles q (Å -1 ) Intensity (a.u.) 54.3 Å 48.6 Å Å 16.2 Å After quenching at 8°C I + II/III q (Å -1 ) Intensity (a.u.) 48.2 Å 44.2 Å 16.1 Å 14.7 Å After making at 10°C II/III + IV X-ray patterns at10°C [Lipids] eau <1% (w/w) Cocoa Butter Polymorphism in Bulk: I < II < III < IV < V < VI [Loisel et al., 1998] 2L3L

P OLYMORPHISM C ONTROL ? (SAXS on ID2 Beamline, E.R.S.F., T. Narayanan) CB/Cholesterol 50/50 (w/w) Nanoparticles II/III + IV + CHOLESTEROL q (Å -1 ) Intensity (a.u.) 47.8 Å 16.1 Å 44 Å 14.5 Å 34.5 Å 17.1 Å Making at 10°C II/III + IV + CHOLESTEROL q (Å -1 ) Intensity (a.u.) 47.8 Å44.2 Å 34.5 Å 17.1 Å 16.1 Å 14.7 Å Tempering at 30°C X-ray patterns at10°C [Lipids] eau <1% (w/w)

P OLYMORPHISM C ONTROL ? (SAXS on ID2 Beamline, E.S.R.F., T. Narayanan) CB/Cholesterol 50/50 (w/w) Nanoparticles q (Å -1 ) -10°C -12°C -14°C -16°C -18°C -20°C -22°C -24°C -26°C -28°C -30°C -32°C -34°C -36°C -38°C -40°C T melting Nanoparticles< T melting Bulk Only 2L structures: I < II < III < IV < V < VI 2L3L

C ONCLUSION microsegregation  Non Homogeneous Particles:microsegregation  Polymorphism Study:low concentration: 1% (w/w) Particle Model Dark-FieldMicroscopyElectronicMicroscopy X-ray Diffraction  MultiTechniques Approach:-size -shape 2L

X- RAY D IFFRACTION (T. Narayanan, E.S.R.F., Grenoble) Mobile sample carrier with a capillary tube Vacuum tube containing detector Incident X-ray beam ID 2 Beamline