Nano-sized hollow polymeric particles are the focus of very intense research due to their potential application in catalysis, separation, diagnostics, drug delivery, and biomolecular-release systems. [1-3] However, only a limited number of polymers can be used as constituents of nanoparticles designed for in vivo applications.[4, 5] The material of the nanoparticle should be degradable to non-toxic low-molecular mass products that can be easily excreted from the organism. Typical examples are polyesters such as polylactide and polycaprolactone. [4, 5]
The fate of polymeric nanoparticles in biological media, especially during in vivo applications, is strongly determined by the biophysicochemical characteristics of their surfaces.[2, 6, 7] As with most surfaces, upon contact with biological media, interfacial properties are rapidly changed by coating with proteins.[8-10] Subsequent colloidal instability [9, 10] of the nanoparticles (e.g. particle aggregation, flocculation, precipitation, etc.) or adsorption of undesirable proteins impair the designed particle functions and initiate serious unfavourable biological responses. Modifications aiming at increasing nanoparticle stability and prolonging their circulation time in blood are of high importance for such systems.[11] Several authors have studied the interaction of polymeric nanoparticles with model protein solutions.[12-15] However, fouling properties of nanocarriers in complex biological media have not been sufficiently addressed yet.
The urgent need for nanoparticles stable in biological fluids turned us towards the development of a new approach for protecting polymeric particles from fouling. The lessons learnt in Chapters 4 to 9 indicate that if particles were to be prepared, coatings based on highly efficient antifouling brushes must be used. In order to achieve this poly(MeOEGMA) brushes were grafted directly from the surface of polycaprolactone nanocapsules (NC) via surface initiated ATRP.
12.1 Synthetic strategy
Figure C12-1 depicts the synthetic strategies for the preparation of antifouling polymeric nanocapsules. Polymeric nanocapsules (NC) of α-carboxy, ω-hydroxy poly(ε-caprolactone) (PCL) containing Mygliol 812® in the core were prepared by interfacial polymer deposition following solvent displacement. The prepared particles had a diameter of 170 nm determined from the particle size distribution measured by quasi elastic light scattering (QELS) and ζ-potential of -23.3 ± 0.5 mV. The negative value of ζ-potential was
12-2 caused previou exploite Subsequ controll shows t brush th
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Figure C12-2. Right side: Kinetics of the surface-initiated ATRP of MeOEGMA brush layers on poly(PCL) nanocapsules in water (triangles) and PBS (circles). The layer thickness was determined by QELS. Left side: TEM images of NC@MeOEGMA prepared by ATRP in PBS for 90 minutes and dried (A and B). A dark initial NC of a diameter about 170 nm is surrounded by a grey corona of the poly(MeOEGMA) coating (B).
12.2 Fouling in biological media
The fouling and stability of the nanoparticles prepared was studied by monitoring the changes in the size distribution of uncoated NC and NC coated with a 60 nm MeOEGMA shell (hydrodynamic diameter 290 nm) induced by the adsorption of HSA (5 mg·mL-1), foetal bovine serum (FBS, 10%), or undiluted human blood plasma (BP) were observed by QELS.
After contact with HSA, FBS or BP the size distribution of NC@MeOEGMA with a mean diameter value of 290 nm changed only marginally (Figure C12-3 a) while the size distribution of uncoated NC with a mean diameter of 170 nm was shifted to markedly larger diameters reaching mean values of about 470 nm in HSA and 320 nm in FBS or BP (Figure C12-3 b). Considering the molecular dimensions of proteins, the large increase in size of uncoated NC (up to 200 nm) could not be due to the formation of a thick protein corona around individual particles but due to the NC clustering induced by protein adsorption at the surface of uncoated PLC shell. A similar effect was previously reported for gold nanoparticles in contact with HSA.[19] The negligible change in size of NC@MeOEGMA indicates that not only were stable but also that a only a negligible fouling might have occurred as in the case of poly(MeOEGMA) brushes grown from planar surfaces.
CHAPTER 12.NANOCARRIERS STABLE IN BIOLOGICAL FLUIDS
12-4
Figure C12-3. Interaction of NC@MeOEGMA (a) and uncoated NC (b) with solution of human serum albumin (HSA), fetal bovine serum (FBS), and undiluted human blood plasma (BP). Diameter distribution of NC@MeOEGMA (a) and uncoated NCs (b) was measured by QELS in PBS (black) and after incubation with HSA solution (5 mg.mL-1, grey dotted), FBS (10% black dotted), or undiluted BP (grey) for 15 minutes. The black-dotted curve for FBS superimposes the grey one for BP in the figure (b).
12.3 Toward bioapplications
As part of our efforts to turn systems from proof of concepts to real applications, the compatibility of each synthetic strategy with the final intended application must be examined. Even though the previously shown system led to antifouling nanocapsules with superior stability in real biological media, biomedical application requires very stringent conditions that are not compatible with any remaining traces of copper catalyst.
In order to tackle this problem a surface initiated controlled polymerisation ensuring high grafting density and a minimum amount of copper catalyst was necessary. The best and probably only candidate to meet these challenges is the novel ARGET ATRP which combines the advantages of ATRP while using only ppb of copper and tolerant to oxygen traces.
Nanospheres (NS) of polycaprolactone (Figure C12-4 black) were prepared and decorated with ATRP initiator (Figure C12-4 blue) utilising the same procedure described above. ARGET ATRP of MeOEGMA was carried out with only 400 parts per billion of CuCl2 complexed by tris(2-pyridylmethyl)amine. Citric acid was used as innocuous
12-5
reducing agent. Figure C12-4 shows the increase of hydrodynamic radius from 60 to ca.
350 nm evidencing the successful polymerisation.
Figure C12-4. ARGET ATRP of MeOEGMA from polymeric nanospheres. Distribution of hydrodynamic radii of nanospheres (black), NS decorated with initiator after dialysis purification (blue) and NS@MeOEGMA prepared by ARGET ATRP at room temperature using citric acid as reducing agent.
12.4 References
[1] H. Hillaireau, P. Couvreur, Cell. Mol. Life Sci. 2009, 66, 2873.
[2] I. Lynch, K. A. Dawson, Nano Today 2008, 3, 40.
[3] C. Vauthier, K. Bouchemal, Pharm. Res. 2009, 26, 1025.
[4] J. S. Chawla, M. M. Amiji, Int. J. Pharm. 2002, 249, 127.
[5] R. Gref, P. Couvreur, G. Barratt, E. Mysiakine, Biomaterials 2003, 24, 4529.
[6] A. E. Nel, L. Madler, D. Velegol, T. Xia, E. M. V. Hoek, P. Somasundaran, F.
Klaessig, V. Castranova, M. Thompson, Nature Materials 2009, 8, 543.
[7] I. Lynch, A. Salvati, K. A. Dawson, Nature Nanotechnology 2009, 4, 546.
[8] C. Rodriguez-Emmenegger, E. Brynda, T. Riedel, Z. Sedlakova, M. Houska, A. B.
Alles, Langmuir 2009, 25, 6328.
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[9] I. Lynch, T. Cedervall, M. Lundqvist, C. Cabaleiro-Lago, S. Linse, K. A. Dawson, Adv.
Colloid Interface Sci. 2007, 134-135, 167.
[10] J. J. Park, M. C. Weiger, S. H. De Paoli Lacerda, D. Pristinski, M. L. Becker, J. F.
Douglas, D. Raghavan, A. Karim, Langmuir 2010, 26, 4822.
[11] V. P. Torchilin, Adv. Drug Del. Rev. 2006, 58, 1532.
[12] C. J. Ochs, G. K. Such, B. Städler, F. Caruso, Biomacromolecules 2008, 9, 3389.
[13] C. R. Kinnane, G. K. Such, G. Antequera-García, Y. Yan, S. J. Dodds, L. M. Liz-Marzan, F. Caruso, Biomacromolecules 2009, 10, 2839.
[14] M. A. Dobrovolskaia, P. Aggarwal, J. B. Hall, S. E. McNeil, Mol. Pharm. 2008, 5, 487.
[15] C.-K. Huang, C.-L. Lo, H.-H. Chen, G.-H. Hsiue, Adv. Funct. Mater. 2007, 17, 2291.
[16] C. E. Mora-Huertas, H. Fessi, A. Elaissari, Int. J. Pharm. 2010, 385, 113.
[17] Q. Zhang, E. E. Remsen, K. L. Wooley, J. Am. Chem. Soc. 2000, 122, 3642.
[18] C. Wu, H. Y. Guo, J. W. Hu, Acta Chim. Sinica 2009, 67, 1621.
[19] S. H. D. P. Lacerda, J. J. Park, C. Meuse, D. Pristinski, M. L. Becker, A. Karim, J. F.
Douglas, ACS Nano 2010, 4, 365.
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