Surface post-modification of polylactide is combined to CuAAc click chemistry or thiol-yne click photochemistry to yield antibiofilm surfaces. INTRODUCTION As a direct result of the life expectancy increase, implants are increasingly used for the restoration of human anatomy and functions. However, this is accompanied with the development of biomaterialassociated septic failures1. To limit these risks, the modulation of the antibacterial surface properties of prosthetic materials appears therefore as a convenient and efficient strategy. In this frame, postpolymerization modification approaches, especially click chemistry, have attracted much attention in the last decade2. In this communication, we wish to report on the recent post-modification strategies developed by our group to yield polylactide antibacterial surfaces. EXPERIMENTAL METHODS PLA surfaces were activated via anionic chemical modification to anchor alkyne moieties, according to a reaction previously described by our group3. This clickable PLA intermediate was then engaged in two distinct strategies. In a first approach, well-controlled quaternized PDMAEMA chains (5-10 kDa) with an azide chain-end were synthesized and covalently grafted to the PLA clickable surfaces by CuAAc 1,3- dipolar cycloaddition. In a second approach, cationic derivatives of α,β-poly(N-2-hydroxyethyl)-aspartamide (PHEA) were functionalized with lipoic acid (LA). Photoactivated thiol-yne reaction was done under UV by reacting PHEA-LA at the surface of the clickable PLA in the presence of TCEP. All polymers have been characterized by NMR and SEC analyses. After careful washes, surfaces were analysed by AFM and XPS. Antibacterial activity was tested against four bacterial strains. Adherence of bacteria and biofilm formation were tested. Cytocompatibility was evaluated with L929 fibroblasts. All data are expressed as means ± SD and correspond to measurements in triplicate. RESULTS AND DISCUSSION Thanks to the use of optimized and mild activation conditions, alkyne functionalized PLA surfaces were obtained without degradation as already reported elsewehere6. CuAAc cycloaddition of QPDMAEMA chains was evaluated by XPS and showed an overall 10% coverage of the PLA surface by QPDMAEMA. No residual copper was detected. Activity was strong against all bacterial strains, including E. Coli and S. Aureus with a clear dependence of the antibacterial activity over QPDMAEMA molecular weight and alkylating agent (Figure 1). Best results were obtained for Mn = 10 000 g/mol and heptyl group with adherence reduction factors > 99.999% (ASTM E 2149–01) and strong bactericidal activity. Fig. 1. Antibacterial activity of antibacterial PLA surface (red bars) against four bacterial strains. PLA plates are shown as control (brown bars).4 With short reaction times and no metals used, UV photoactivated thiol-yne grafting of PHEA-LA was studied as a green alternative to CuAAc. Advantageously, the heterogeneous surface reaction took place in aqueous media. Various solvent mixtures were tested and best results were obtained in slightly acidic water/ethanol (1:1) medium, in the presence of TCEP and with a 15 min irradiation time per PLA plate side. Under these conditions, XPS confirmed the covalent grafting of the cationic PHEA-LA derivative. As for CuAAc, antibacterial and antibiofilm activities were tested with again reduction factors > 99.999% and up to 80% biofilm decrease for all four bacterial strains. Interestingly, whatever the methodology used all surfaces showed a good cytocompatibility towards L929 fibroblasts cells with respect to TCPS control. CONCLUSION Antibacterial PLA surface modification was obtained in an efficient two steps approach by combining anionic and click chemistries. For the first time green thiol-yne strategy was applied to PLA surfaces, which paves the way to further developments in the biomedical field. REFERENCES 1. Campoccia D. et al. Biomaterials 34:8018-8029, 2013 2. Theato P. & Klok H-A. Eds. Wiley-CH, 2013 3. El Habnouni S. et al. Adv. Funct. Mater. 21:3321-3330, 2011 4. El Habnouni S. et al. Acta Biomater. 9 :7709-7718, 2013 ACKNOWLEDGMENTS Authors thank the French MESR for PhD fundings.
TOWARD POTENT ANTIBIOFILM DEGRADABLE MEDICAL DEVICES: GENERIC METHODOLOGIES FOR THE SURFACE MODIfiCATION OF POLYLACTIDE
Sardo C;Cavallaro G;
2014
Abstract
Surface post-modification of polylactide is combined to CuAAc click chemistry or thiol-yne click photochemistry to yield antibiofilm surfaces. INTRODUCTION As a direct result of the life expectancy increase, implants are increasingly used for the restoration of human anatomy and functions. However, this is accompanied with the development of biomaterialassociated septic failures1. To limit these risks, the modulation of the antibacterial surface properties of prosthetic materials appears therefore as a convenient and efficient strategy. In this frame, postpolymerization modification approaches, especially click chemistry, have attracted much attention in the last decade2. In this communication, we wish to report on the recent post-modification strategies developed by our group to yield polylactide antibacterial surfaces. EXPERIMENTAL METHODS PLA surfaces were activated via anionic chemical modification to anchor alkyne moieties, according to a reaction previously described by our group3. This clickable PLA intermediate was then engaged in two distinct strategies. In a first approach, well-controlled quaternized PDMAEMA chains (5-10 kDa) with an azide chain-end were synthesized and covalently grafted to the PLA clickable surfaces by CuAAc 1,3- dipolar cycloaddition. In a second approach, cationic derivatives of α,β-poly(N-2-hydroxyethyl)-aspartamide (PHEA) were functionalized with lipoic acid (LA). Photoactivated thiol-yne reaction was done under UV by reacting PHEA-LA at the surface of the clickable PLA in the presence of TCEP. All polymers have been characterized by NMR and SEC analyses. After careful washes, surfaces were analysed by AFM and XPS. Antibacterial activity was tested against four bacterial strains. Adherence of bacteria and biofilm formation were tested. Cytocompatibility was evaluated with L929 fibroblasts. All data are expressed as means ± SD and correspond to measurements in triplicate. RESULTS AND DISCUSSION Thanks to the use of optimized and mild activation conditions, alkyne functionalized PLA surfaces were obtained without degradation as already reported elsewehere6. CuAAc cycloaddition of QPDMAEMA chains was evaluated by XPS and showed an overall 10% coverage of the PLA surface by QPDMAEMA. No residual copper was detected. Activity was strong against all bacterial strains, including E. Coli and S. Aureus with a clear dependence of the antibacterial activity over QPDMAEMA molecular weight and alkylating agent (Figure 1). Best results were obtained for Mn = 10 000 g/mol and heptyl group with adherence reduction factors > 99.999% (ASTM E 2149–01) and strong bactericidal activity. Fig. 1. Antibacterial activity of antibacterial PLA surface (red bars) against four bacterial strains. PLA plates are shown as control (brown bars).4 With short reaction times and no metals used, UV photoactivated thiol-yne grafting of PHEA-LA was studied as a green alternative to CuAAc. Advantageously, the heterogeneous surface reaction took place in aqueous media. Various solvent mixtures were tested and best results were obtained in slightly acidic water/ethanol (1:1) medium, in the presence of TCEP and with a 15 min irradiation time per PLA plate side. Under these conditions, XPS confirmed the covalent grafting of the cationic PHEA-LA derivative. As for CuAAc, antibacterial and antibiofilm activities were tested with again reduction factors > 99.999% and up to 80% biofilm decrease for all four bacterial strains. Interestingly, whatever the methodology used all surfaces showed a good cytocompatibility towards L929 fibroblasts cells with respect to TCPS control. CONCLUSION Antibacterial PLA surface modification was obtained in an efficient two steps approach by combining anionic and click chemistries. For the first time green thiol-yne strategy was applied to PLA surfaces, which paves the way to further developments in the biomedical field. REFERENCES 1. Campoccia D. et al. Biomaterials 34:8018-8029, 2013 2. Theato P. & Klok H-A. Eds. Wiley-CH, 2013 3. El Habnouni S. et al. Adv. Funct. Mater. 21:3321-3330, 2011 4. El Habnouni S. et al. Acta Biomater. 9 :7709-7718, 2013 ACKNOWLEDGMENTS Authors thank the French MESR for PhD fundings.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
