et biodégradation des matériaux biorésorbables Jean CUDANE Centre de Recherches sur les Biopolymères Artificiels UMR CNRS 5473 Faculté de Pharmacie, Montpellier, France jcoudane@univ-montp1.fr Sophia Antipolis 3 Mars 26 Plan Introduction-mécanisme-évaluation enzymatique 1
Poly (α-hydroxy acides) Biomatériaux polymères de synthèse dégradables PLA/PLGA (biomédical) C CH CH 3 n C CH 2 n PCL (environnemental) C (C H 2 ) 5 n introduction General Degradation Mechanism of PLA-based Devices 1-Penetration of H 2 2-Hydrolysis of polymeric chains Water uptake : Surface mechanism enzymatic Core mechanism Hydrolytic Massive objects Decrease of molar masses Size-dependent formation of low molecular masses compounds Variation in composition (copolymers) Modification of surface aspect Differentiation between surface and core 3-Elimination of degradation compounds a-weight loss b-acid lactic titration 2
introduction Degradation percentage vs random chain scission Molar masses Mn= 72 DPn = 1 DPn= 5 Mn= 72 DPn =1 DPn= 5 Mn= 72 DPn =1 DPn= 5 1/1 Mn/2 1/1 1/1 No lag time SEC-Half degradation Weight loss Soluble oligomers, Mn<72, DPn<1 At least 1/1 Lag time WL-degradation Lactic acid formation At least 5/1 DP=1 LA-degradation Lag time introduction Degradation percentage vs depolymerization Molar masses Mn= 72 DPn= 1 1 chain breakings (1/1) Mn 71 DPn= 99 Mn constant no significant SEC-degradation Weight loss WL-degradation Immediate weight loss No lag time Lactic acid formation LA-degradation Immediate lactate formation No lag time 3
introduction Random Chain Scission Depolymerization Molar masses Mn decrease No lag time Mn constant Weight loss delayed weight loss Lag time Immediate weight loss Good evaluation of degradation No lag time Lactic acid delayed lactate formation Lag time Immediate lactate formation Good evaluation of degradation No lag time Plan Introduction-mécanismes enzymatique 4
PLA 37.5 GA 25 37.5% LLact 37.5% DLact 25% Gly PLA x PLAxGAy First step of degradation: water uptake PBS 37 C water uptake x= % unités L-lactique y= % unités glycolique plates (2-3 mm) Time (days) 3 Li et al 199 Distilled water 37 C Stereochemistry Hydrophobicity crystallinity PLA 75 GA 25 75% LLact % DLact 25% Gly water uptake Time (weeks) 1 second step of degradation: hydrolysis of polymeric backbone PBS, 37 C plates (2-3 mm) Li et al 199 modulus (Gpa) PLA 37.5 GA 25 Continuous decrease of molar masses 1 Time (days) core SEC modulus (Gpa) 5 Time (weeks) surface PLA 75 GA Time min 25 Core-surface effect 5
Third step of degradation: weight loss PBS 37 C plates (2-3 mm) Li et al 199 Weight loss % PLA 37.5 GA 25 lag time Time (days) Delayed loss of matter Weight loss % PLA 75 GA 25 Time (weeks) Random chain scission General degradation mechanism of PLA-based devices Soluble oligomers Soluble oligomers Time in weeks 15 Water absorption and degradation without loss of matter Beginning of the loss of matter Role of Polydispersity Same results in vivo Central degradation and formation of an outer membrane Formation of hollow structure Weight loss 6
interne- cas du PLA From massive objects to micro/nanoparticles 7
water uptake (%) Influence of size on degradation From plates to microparticles plates films Molecular weight % of initial Mw plates Grizzi et al, 1996 µspheres + others PLA 5 Weight loss (%) plates films L-lactic acid (mm) plates beads µspheres films Plates 2-3mm µspheres 125-25 µm Beads.5-1mm Films 6µm Differentiation core-surface Influence of size on degradation Grizzi et al, 1996 From plates to microparticles Molecular weights (SEC) plates films microspheres beads Degradation time maximum 3 weeks 8
Water uptake (%) 4 3 2 Molar Masses 1 1 2 Time (days) 1 8 6 4 2 1 2 3 Time (days) Chemical modification of PCL H (CH 2) 4 C C (CH 2) 4 C H 1-x H Water uptake (%) CH PCL 4 3 2 1 PCLCH1 Weight loss (%) 25 2 15 1 5 C PCLCH1 Mw PCL Mw x Gimenez et al, 2 Phosphate buffer (ph 7.4), 37 C, plates of polymers ( Ø = 1cm, t= 2mm) Substitution ratio = 11%, 4, 8, 12, 16, Time (hours) 5 1 15 2 Time (days) PCLCH1 PCL Surface Aspect (ESEM) Magnification: x 15 PCL t 98 PCLCH1 t 98 9
H (CH 2) 4 C C H 1-x (CH 2) 4 C CH H C x x=11% 1-Apparent water uptake : (m f -m i )/m i PCLCH 11 : 3% in 12h PCL :.7% in 28 weeks 2-Molecular Weights PCLCH 11 : after 24 weeks Mn / 2.5, Mw / 7 PI PCL : after 28 weeks Mn / 1.1 and Mw / 1.3 3-Weight loss PCLCH 11 : 22% in 24 weeks PCL : 1.6% in 28 weeks Preferential Degradation of acidic units : PCLCH 11 : After 24 weeks : 3.5% CH groups on the chain Cas des copolymères PLA et PGA : dégradation essentiellement PCL peu de dégradation Li et al, 25 Copolymères statistiques PCL-PGA H (CH 2 ) 4 C H C H C C 1-x H x Prise d eau (%) Perte de masse (%) Temps (semaines) Temps (semaines) Cop1 à cop7: taux de PCL 1
Cas des copolymères Copolymères blocs PEG 2 -PLA 3 Stefani et al, 26 LA/E 1 9 8 7 6 5 P5 P6 P61 P67 Effet cœur/surface 4 P61UF 3 2 hydrosoluble 1 1 2 3 4 5 6 7 8 9 Préparation «anionique» PLA PEG LA/E 9 8 7 6 5 4 3 2 1 2 4 6 8 1 12 Préparation «octanoate d étain» Après purification PSn5 PSn5P PSn5UF PSn5 PSn5P PSn5UF Conclusions on degradation of PLA-based polymers Degradation rate can be modulated according to some parameters: Structural parameters Stereoisomery of PLA X X > 92 semi-crystalline : slow degradation rate X 5 amorphous : higher degradation rate Size parameters ther parameters Copolymers GA : increase hydrophylicity higher degradation rate PE (blocs): variation of proportions Massive objects : differentiation core surface Micro and nanospheres : no differentiation core-surface Degradation rate function of S/V ratio. Molecular masses, Polydispersity Reaction medium : ph, Temperature, presence of enzyme Purity of polymers PLA, PLAGA : days to months 11
Plan Introduction enzymatique enzymatique degradation medium: enzymes Landry et al 1996 PLA 5 nanoparticles 1 nm, Albumin coating ph = 1.2, Pepsin ph = 7.5, Pancreatic lipase (Mn = 38) 8 8 8 Percentage of PLA converted in lactate Percentage of PLA converted in lactate Weight loss % 5 Incubation time (min) 5 Incubation time (min) 12
enzymatique degradation medium: enzymes Landry et al 1996 PLA 5 nanoparticles 1 5 1 6 Molar Mass (log scale) No variation in PLA 5 Molar Masses Surface erosion enzymatique Tritiated PCL : Determination of biodegradation kinetics Ponsart et al, 21 H T (CH 2 ) 4 C C (CH 2 ) 4 C C H 1-x H x x <<1% Environmental Conditions: PCL film, activated sludge, aqueous medium, 37 C % of initial radioactivity 1 8 6 4 2 1 2 3 4 5 6 7 biomass 4 3 2 1 1 2 3 4 5 Time (days) After 72 days : 75-85% of the initial radioactivity were found in the incubation medium under the form of tritiated water and the rest on the form of biomass 13
enzymatique PCL Methylation: influence de la structure sur la biodegradation H CH 3 (CH 2) 4 C C (CH 2) 4 C C H 1-x H x (x=15%) [Ponsart et al., 23] (ph 7.4, 37 C) Biodégradation (ph 7.4, 37 C, pseudomonas cepacia lipase) PCLCH 3 PCL PCLCH 3 < PCL Pas de variation significative (pas de dégradation) Diminution de la vitesse de biodégradation weight loss (%) 6 4 2, 1, 2, 3, 4, 5, time (days) conclusions Paramètres structuraux Stéréoisomérie, cristallinité, hydrophilie Masses molaires, polymolécularité Copolyméres : variation de la composition, variation de la vitesse de dégradation Taille des objets bjets massifs: differentiation coeur-surface Micro and nanospheres : pas de differentiation coeur-surface Vitesse de dégradation fonction du rapport S/V. Autres paramètres Milieu réactionnel: ph, Temperature, presence of enzyme 14
remerciements S. Li, H. Garreau, I. Grizzi S. Ponsart, S. Gimenez, A. des Rieux 15