ORGANIC PHASE FABRICATION OF CORE-SHELL MATERIALS FOR THE ENCAPSULATION OF BIOMOLECULES BAI JIANHAO (B.Eng. (Hons.), NUS) A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY DIVISION OF BIOENGINEERING NATIONAL UNIVERSITY OF SINGAPORE 2010 ACKNOWLEDGEMENTS I would like to thank the Division of Bioengineering and National University of Singapore for provision of scholarship and research grant that has given me the opportunity to pursue this PhD course in NUS. I would also like to commend on the excellent facilities and common sharing equipments provided by the department and which have definitely allowed me to conduct my research effectively and with convenience. I am extremely appreciative of the guidance and help my supervisor, Dr. Trau, Dieter Wilhelm, have offered to me during the many hurdles I have encountered in my entire course of work. The countless constructive suggestions and directions he has given me were instrumental in the timely completion of my PhD work. I would like to thank Dr. Martin Mak Wing Cheung for supervising me as well during his working stint in the NanoBioanalytics Laboratory. He was patient in mentoring me during my initial research years and I have learned many research related skills from him. The other members of the NanoBioanalytics Laboratory are also acknowledged for the assistance and support they have provided me. I thank Prof. Colin Sheppard and members of the Optical Imaging Laboratory for the use of their optical imaging systems. I am also grateful to Ms Cheng Jinting for her support and the many research experiences she has shared with me. Most importantly, I thank my father, Pah Tuck Weng, and mother, Ng Ou, for their constant support and immense care given to me since birth. Of course, all these would not have been possible if not for God’s Love and Blessing! i PUBLICATIONS, CONFERENCES & AWARDS Publications 1) W. C. Mak*; J. Bai*; X. Y. Chang; D. Trau, Matrix-Assisted Colloidosome ReversePhase Layer-By-Layer: Encapsulating Biomolecules in Hydrogel Microcapsules with Extremely High Efficiency and Retention Stability Langmuir, 2009, 25, 769-775. *Authors contributed equally 2) J. Bai; S. Beyer; W. C. Mak; D. Trau, Fabrication of Inflated LbL Microcapsules with a “Bead-in-a-Capsule” Architecture Soft Matter, 2009, 5, 4152 - 4160. 3) J. Bai; S. Beyer; W. C. Mak; R. Rajagopalan; D. Trau, Inwards Buildup of Concentric Polymer Layers: A Method for Biomolecule Encapsulation and Microcapsule Encoding Angew. Chem. Int. Ed., 2010, 49, 5189 - 5193. International Conferences 1) W. C. Mak; C. Y. Soh; K. Y. Cheung; J. Bai; D. Trau, Layer-By-Layer Surface Engineered Hydrogel Microcapsules-The Encapsulation Efficiency and Temperature Stability for Biochemical Processing Application, World Congress on Bioengineering 2007, Thailand. ii 2) J. Bai; W. C. Mak; X. Y. Chang; D. Trau, Organic Phase Coating of Polymers onto Agarose Microcapsules for Encapsulation of Biomolecules with High Efficiency, 13th International Conference on Biomedical Engineering, Singapore. Oral Presentation. In book series, IFMBE Proceedings, Volume 23, pg 821-824. 3) J. Bai; S. Beyer; D. Trau, Reverse-Phase Layer-by-Layer Assembly of Polymers: A Strategic Tool for New Applications, World Congress on Bioengineering 2009, Hong Kong. Poster Presentation. 4) J. Bai; D. Trau, A Novel Polymer-Hydrogel Complex Formation for Encapsulating Low Molecular Weight Macromolecules, Encoding Hydrogel Microcapsules and Release Applications, Formula VI 2010, Stockholm. Oral Presentation. Student Workshops 1) J. Bai; D. Trau, Strategies for Organic Phase Encapsulation of Biomolecules within Agarose Microbeads Through the Use of Polymers, 3rd East Asian Pacific Student Workshop on Nano-Biomedical Engineering 2009. Oral Presentation. Awards 1) Outstanding Paper Award, 13th International Conference on Biomedical Engineering iii SUMMARY Encapsulation of biomolecules within microcapsules has seen tremendous progress in the biomedical field, such as in bioanalysis, bioreactor and drug delivery applications, and is a growing interest amongst researchers in the recent years. The popularity of microcapsules stems from its minute nature that allows for efficient exchange of materials between the microcapsules and its environment. Core-shell materials, a sub-class of microcapsules, are commonly employed for the encapsulation of biomolecules due to the mechanical stability these microcapsules can provide. The “matrix-assisted Layer-byLayer (LbL) encapsulation” technique, performed in an aqueous dispersant, is an example of hydrogel core-shell materials fabricated for the encapsulation of biomolecules. However, biomolecule encapsulation problems (e.g. low encapsulation efficiency, poor control on encapsulated biomolecules quantity or poor retention stability) are associated with using current aqueous phase encapsulation techniques. The use of an organic phase for fabrication of core-shell materials and encapsulation of biomolecules is rarely demonstrated. Therefore, this PhD work involves the novel organic phase fabrication of core-shell materials and encapsulation of biomolecule with high encapsulation efficiency and retention stability. Desired biomolecules are first pre-loaded into agarose microbeads fabricated via a waterin-oil emulsion technique. Using an emulsion technique allows all pre-mixed biomolecules within the hydrogel solution to be pre-loaded into the the resultant hydrogel microbeads. Following, these agarose microbeads are stabilized in anyhydrous 1-butanol by depositing amino-polystyrene microparticles along the periphery and surface of each iv microbead. Termed as ‘matrix-assisted colloidosomes (MACs)’, the surfaces of these microparticles stabilized agarose microbeads were next deposited with polymers (nonionized polyallylamine (niPA) and poly(acrylic acid) (niPAA) ) dissolved in 1-butanol, via the Reverse-Phase LbL technique, for the fabrication of polymeric shells around each MAC core template. It was demonstrated that a high encapsulation efficiency of biomolecules could be obtained through the organic phase fabrication MAC RP-LbL coreshell materials; and with almost 100% retention stability after days incubation in an aqueous dispersant. In addition, encapsulated glucose oxidase (GOx) and horseradish peroxidise (HRP) could retain their bioactivity in these MAC RP-LbL core-shell materials. Asides from microparticles, ADOGEN® 464 (a cationic surfactant) was also used to stabilize these agarose microbeads in 1-butanol. High retention stability of dextran (> 500,000 Da) was observed but poor retention stability of dextran (< 155,000 Da) was observed for agarose (core) RP-LbL (shell) microcapsules fabricated using ADOGEN® 464 and the RP-LbL technique. This highlights that the use of agarose (core) RP-LbL (shell) microcapsules fabricated using ADOGEN® 464 and the RP-LbL technique is more suitable for encapsulating higher MW biomolecules with high retention stability. Remarkably, incubation of only niPA with agarose microbeads in 1-butanol (as solvent and dispersant respectively) results in a thick uniform polymeric layer forming in the peripheral matrix around each core microbead. This novel polymeric shell fabrication technique is driven by diffusion and is termed as the “inwards deposition of concentric niPA layers” technique. Upon stabilization of these layers into shells, with a cross-linker, these core-shell materials could be stably dispersed in an aqueous phase and were demonstrated to be capable of encapsulating and retaining pre-loaded low MW FITCv dextran (4,000 Da). The retention efficiency was determined to be ~95% after a days incubation period in an aqueous dispersant. Separate incubation of niPA or niPA conjugated with a dye (FITC or TRITC), inclusive of washing steps, results in the fabrication of shells consisting of distinct coloured striated layers. Permutation of the color sequence allows for encoding purposes. It was also demonstrated that the thickness could be tuned, through manipulation of niPA volume or incubation time, and would therefore allow for an agarose core-shell microcapsule encoding system with at least levels of encoding. Lastly, encapsulated GOx and HRP were demonstrated to have retained their bioactivity in these unique encoded core-shell materials and further highlight the potential of utilizing the “inwards deposition of concentric niPA layers” technique for potential multiplexing applications. vi TABLE OF CONTENTS PAGE NUMBER ACKNOWLEDGEMENTS . I PUBLICATIONS, CONFERENCES & AWARDS . II SUMMARY IV LIST OF SCHEMES . XI LIST OF FIGURES XII ABBREVIATIONS XVIII CHAPTER – INTRODUCTION . CHAPTER – LITERATURE REVIEW . 2.1 INTRODUCTION . 2.2 TECHNIQUES FOR THE FABRICATION OF MICROCAPSULES . 2.2.1 Self-Assembled Phopholipid Bilayers (Liposomes) . 2.2.2 Solvent Evaporation 2.2.3 Interfacial Assembly of Microparticles . 2.2.4 Interfacial Polymerization . 2.2.5 Layer-by-Layer (LbL) Technique 10 2.3 ENCAPSULATION OF BIOMOLECULES WITHIN LBL MICROCAPSULES . 16 2.3.1 Encapsulation in Core-Shell LbL Microcapsules 16 2.3.2 Encapsulation in Hollow LbL Microcapsules . 20 2.4 APPLICATIONS OF BIOMOLECULES LOADED LBL MICROCAPSULES . 23 2.4.1 Biosensors . 23 2.4.2 Bioreactors 24 2.4.3 Drug Releasing/Therapeutics 25 CHAPTER – OBJECTIVE & SPECIFIC AIMS . 27 3.1 OBJECTIVE . 27 3.2 SPECIFIC AIMS . 27 vii 3.2.1 Specific Aim – Fabrication of Hydrogel Microbeads as a Core Template and Suitable Vessel of Biomolecules. 27 3.2.2 Specific Aim – Selection of Organic Solvent and Polymers for Organic Phase Fabrication of Core-Shell Materials 28 3.2.3 Specific Aim – Organic Phase Fabrication of Hydrogel Core-Shell Materials . 28 3.2.4 Specific Aim – Characterization of Fabricated Core-Shell Materials . 29 3.2.5 Specific Aim – Encapsulation of Macromolecular Biomolecules within Fabricated Core-Shell Materials 29 CHAPTER – FABRICATION OF HYDROGEL MICROBEADS & SELECTION OF ORGANIC SOLVENT AND POLYMERS FOR FABRICATION OF POLYMERIC SHELLS 30 4.1 INTRODUCTION . 30 4.2 MATERIALS AND METHODS . 32 4.3 RESULTS AND DISCUSSION . 33 4.3.1 Selection and Fabrication of Hydrogel Microbeads as Core Templates 33 4.3.2 Selection of Suitable Organic Solvent and Polymers for Organic Phase Fabrication of Core-Shell Materials 35 4.3.3 Stability of Agarose Microbeads in 1-Butanol 37 4.4 CONCLUSION 37 CHAPTER – ENCAPSULATION OF BIOMOLECULES WITHIN MICROPARTICLES STABILIZED AGAROSE MICROBEADS VIA THE REVERSE-PHASE LAYER-BY-LAYER TECHNIQUE 39 5.1 INTRODUCTION . 39 5.2 MATERIALS & METHODS . 41 5.3 RESULTS & DISCUSSION . 47 5.3.1 Morphology and Stability of “Matrix-Assisted” Colloidosomes (MACs) in 1-Butanol 47 5.3.2 Importance of an Organic Phase to Prevent Leaching of Pre-Loaded Biomolecules from MACs . 49 5.3.3 Demonstration of Organic Phase Fabrication of Non-Ionized Polyelectrolyte (niPolyelectrolyte) Multilayer Shell onto PS Microparticles via the RP-LbL Technique 51 viii Chapter – Conclusion & Future Works significantly to the field of colloidal science and engineering and microencapsulation of biomolecules. For example, the unique inwards deposition phenomenon observed could lead to further developments of other novel core-shell materials and the encoding system can allow for the development of complex and multiplexed microcapsule bio-analytical systems. 119 Chapter – Conclusion & Future Works 8.2 Future Works In this PhD work, only poly(allylamine) and poly(acrylic acid) had been adopted for the organic phase fabrication of core-shell material. Use of other polymers such as polyvinylpyrrolidone, polyethyleneimine and poly(styrene sulfonic acid) dissolved in 1-butanol may be explored as well for the organic phase fabrication of the polymeric shells around the agarose core templates. The encapsulation efficiency and retention stability of macromolecular biomolecules within the different core-shell materials fabricated from different permutations of these polymers can be compared as well. Agarose is a polysaccharide and temperature sensitive hydrogel that was used for the fabrication of the microcapsules. Other hydrogels with different properties may be explored for the organic phase fabrication of the hydrogel core-shell materials. These include alginic acid, gelatine, polyacrylamide, hyaluronic acid and carboxymethylcellulose. The “inwards deposition of concentric niPA layers” technique is a novel one that can be used for the fabrication of agarose core-shell materials with thick polymeric shells. This can be easily performed in a few hours and with good reproducibility. However, the exact deposition of niPA within the agarose matrices and diffusion kinetics of niPA into the agarose matrices are not yet well understood. To understand the deposition mechanism, various polymers of different properties (electrostatic charges, hydrophobicity, hydrophilicity and molecular weight) can be 120 Chapter – Conclusion & Future Works incubated with the agarose core templates and studied. Such polymers include polyethyleneimine, poly-L-lysine, poly(acrylic acid) and poly(styrene sulfonic acid). Diffusion kinetics can be easily understood by measuring the time it takes for niPA to completely deposit within the matrices of agarose template of different diameters. Computer modeling of the diffusion kinetics with the hypothesized deposition mechanism and parameters may be performed for further understanding and for comparison between theoretical and experimental results. In Chapter 7, it was briefly demonstrated that release of encapsulated materials can be achieved via the use of DSP (amino bi-functional cross-linker with a cleavable central disulfide group). Other cross-linkers (as mentioned in Chapter 7) along with their release profiles and with respect to different thickness of the niPA shell used can be studied for possible biomedical applications such as DNA delivery. Conjugated polymers have seen immense advances in the field of bioanalysis. Yet, most analytical works described are performed in solution or on a solid platform. It would be interesting if this unique group of polymers can be encapsulated within core-shell materials for biosensing or bioanalytical purposes. To improve the uniformity in size distribution of the agarose microcapsules, the use of Shirasu Porous Glass membranes (Zhou et al., 2007, 2009) or microfluidics can be adopted to ensure that the core templates formed are uniformly sized. Automation of the core-shell material fabrication process may even be possible if microfluidics is used. 121 References References Antipov, A.A., Sukhorukov, G.B., Donath, E. and Möhwald, H. Sustained Release Properties of Polyelectrolyte Multilayer Capsules. J. Phys. Chem. B 105, pp. 2281-2284, 2001. Antipov, A.A., Sukhorukov, G.B., Leporatti, S., Radtchenko, T.L., Donath, E. and Möhwald, H. Polyelectrolyte Multilayer Capsule Permeability Control. Colloid Surf. Physicochem. Eng. Aspects 198, pp. 535-541, 2002. Antipov, A.A., Shchukin, D., Fedutik, Y., Zanaveskina, I., Klechkovskaya, V., Sukhorukov, G.B. and Möhwald, H. Urease-Catalyzed Carbonate Precipitation inside the Restricted Volume of Polyelectrolyte Capsules. Macromol. Rapid Commun. 24, pp. 274277, 2003a. Antipov, A.A., Shchukin, D., Fedutik, Y., Petrov, A.I., Sukhorukov, G.B. and Möhwald, H. Carbonate Microparticles for Hollow Polyelectrolyte Capsules Fabrication. Colloid Surf. Physicochem. Eng. Aspects 224, pp. 175-183, 2003b. Atkin, R., Davie, P., Hardy, J. and Vincent, B. Preparation of Aqueous Core/Polymer Shell Microcapsules by Internal Phase Separation. Macromolecules 37, pp. 7979-7985, 2004. Bai, J., Beyer, S., Mak, W.C. and Trau, D. Fabrication of Inflated LbL Microcapsules with a ‘Bead-in-a-Capsule’ Morphology. Soft Matter 5, pp. 4152-4160, 2009. Bai, J., Beyer, S., Mak, W.C., Rajagopalan, R. and Trau, D. Inwards Buildup of Concentric Polymer Layers: A Method for Biomolecule Encapsulation and Microcapsule Encoding. . Angew. Chem. Int. Ed. 49, pp. 5189-5193, 2010. Balabushevich, N.G., Tiourina, O.P., Volodkin, D.V., Larionova, N.I. and Sukhorukov, G.B. Loading the Multilayer Dextran Sulfate/Protamine Microsized Capsules with Peroxidase. Biomacromolecules 4, pp. 1191-1197, 2003. Beyer, S., Mak, W.C. and Trau, D. Reverse-Phase LbL-encapsulation of Highly Water Soluble Materials by Layer-by-Layer Polyelectrolyte Self-assemble. Langmuir 23, pp. 8827-8832, 2007. Binks, B.P. Particles as Surfactants-Similarities and Differences. Curr. Opin. Colloid Interface Sci. 7, pp. 21-41, 2002. Bon, S.A.F., Cauvin, S. and Colver, P.J. Colloidosomes as Micron-sized Polymerisation Vessels to Create Supracolloidal Interpenetrating Polymer Network Reinforced Capsules. Soft Matter, 3, pp. 194-199, 2007. 122 Borodina, T.N., Grigoriev, D.O., Andreeva, D.V., Möhwald, H. and Shchukin, D.G. Polyelectrolyte Multilayered Nanofilms as a Novel Approach for the Protection of Hydrogen Storage Materials. ACS Appl. Mater. Interfaces 1, pp. 996-1001, 2009. Borodina, T.N., Markvicheva, E., Kunizhev, S., Möhwald, H., Sukhorukov, G.B. and Kreft, O. Controlled Release of DNA from Self-Degrading Microcapsules. Macromol. Rapid Commun. 28, pp. 1894-1899, 2007. Brown, J.Q., Srivastava, R. and McShane, M.J. Encapsulation of Glucose Oxidase and an Oxygen-Quenched Fluorophore in Polyelectrolyte-Coated Calcium Alginate Microspheres as Optical Glucose Sensor Systems. Biosens. Bioelectron. 21, pp. 212-216, 2005. Cayre, O.J., Noble, P.F. and Paunov, V.N. Fabrication of Novel Colloidosome Microcapsules with Gelled Aqueous Core. J. Mater. Chem. 14, pp. 3351-3355, 2004. Caruso, F. Hollow Capsule Processing through Colloidal Templating and Self-Assembly. Chem. Eur. J. 6, pp. 413-419, 2000. Caruso, F. Nanoengineering of Particle Surfaces. Adv. Mater. 13, pp. 11-22, 2001. Caruso, F., Yang, W., Trau, D. and Renneberg, R. Microencapsulation of Uncharged Low Molecular Weight Organic Materials by Polyelectrolyte Multilayer Self-Assembly. Langmuir 16, pp. 8932-8936, 2000a. Caruso, F., Trau, D., Möhwald, H. and Renneberg, R. Enzyme Encapsulation in Layer-byLayer Engineered Polymer Multilayer Capsules. Langmuir 16, pp. 1485-1488, 2000b. Chaize, B., Colletier, J., Winterhalter, M. and Fournier, D. Encapsulation of Enzyme in Liposomes: High Encapsulation Efficiency and Control of Substrate Permeability. Artif. Cells, Blood Substitutes, Biotechnol. 32, pp. 67-75, 2004. Chinnayelka, S. and McShane, M.J. Microcapsule Biosensors Using Competitive Binding Resonance Energy Transfer Assays based on Apoenzymes. Anal. Chem. 77, pp. 55015511, 2005. Cortez, C., Tomaskovic-Crook, E., Johnston, A.P.R., Radt, B., Cody, S.H., Scott, A.M., Nice, E.C., Heath, J.K. and Caruso, F. Targeting and Uptake of Multilayered Particles to Colorectal Cancer Cells. Adv. Mater. 18, pp. 1998-2003, 2006. Dai, Z., Heilig, A., Zastrow, H., Donath, E. and Möhwald, H. Novel Formulations of Vitamins and Insulin by Nanoengineering of Polyelectrolyte Multilayers around Microcrystals. Chem. Eur. J. 10, pp. 6369-6374, 2004. De Geest, B.G., Vandenbroucke, R.E., Guenther, A.M., Sukhorukov, G.B., Hennink, W.E., Sanders, N.N., Demeester, J. and De Smedt, S.C. Intracellularly Degradable Polyelectrolyte Microcapsules. Adv. Mater. 18, pp. 1005-1009, 2006a. 123 De Geest, B.G., Jonas, A.M., Demeester, J. and De Smedt, S.C. Glucose-Responsive Polyelectrolyte Capsules. Langmuir 22, pp. 5070-5074, 2006b. Decher, G. Fuzzy Nanoassemblies: Toward Layered Polymeric Multicomposites. Science 277, pp. 1232-1237, 1997. Decher, G., Hong, J.D. and Schmitt, J. Buildup of Ultrathin Multilayer Films by a Selfassembly Process: III. Consecutively Alternating Adsorption of Anionic and Cationic Polyelectrolytes on Charged Surfaces. Thin Solid Films 210/211, pp. 831-835, 1992. Decher, G., Lvov, Y. and Schmitt, J. Proof of Multilayer Structural Organization in SelfAssembled Polycation-Polyanion Molecular Films. Thin Solid Films 244, pp. 772-777, 1994. Déjugnat, C., Haložan, D. and Sukhorukov, G.B. Defined Picogram Dose Inclusion and Release of Macromolecules using Polyelectrolyte Microcapsules. Macromol. Rapid Commun. 26, pp. 961-967, 2005. Deng, C., Dong, W., Adalsteinsson, T., Ferri, J.K., Sukhorukov, G.B. and Möhwald, H. Solvent-filled Matrix Polyelectrolyte Capsules: Preparation, Structure and Dynamics. Soft Matter 3, pp.1293-1299, 2007. Donath, E., Sukhorukov, G.B., Caruso, F., Davis, S.A. and Möhwald, H. Novel Hollow Polymer Shells by Colloid-Templated Assembly of Polyelectrolytes. Angew. Chem. Int. Ed. 37, pp. 2201-2205, 1998. Dong, W.F., Liu, S., Wan, L., Mao, G., Kurth, D.G. and Möhwald, H. Controlled Permeability in Polyelectrolyte Films via Solvent Treatment. Chem. Mater. 17, pp. 49924999, 2005. Doumèche, B., Heinemann, M., Büchs, J., Hartmeier, W. and Ansorge-Schumacher, M.D. Enzymatic Catalysis in Gel-Stabilized Two-Phase Systems: Improvement of the Solvent Phase. J. Mol. Catal. B: Enzym. 18, pp. 19-27, 2002. Dowding, P.J., Atkin, R., Vincent, B. and Bouillot, P. Oil Core-Polymer Shell Microcapsules Prepared by Internal Phase Separation from Emulsion Droplets. I. Characterization and Release Rates for Microcapsules with Polystyrene Shells. Langmuir 20, pp. 11374-11379, 2004. Duan, L., He, Q., Yan, X., Cui, Y., Wang, K. and Li, J. Hemoglobin Protein Hollow Shells Fabricated Through Covalent Layer-by-Layer Technique. Biochem. Biophys. Res. Commun. 354, pp. 357-362, 2007a. Duan, L., He, Q., Wang, K., Yan, X., Cui, Y., Möhwald, H. and Li, J. Adenosine Triphosphate Biosynthesis Catalyzed by F0F1 ATP Synthase Assembled in Polymer Microcapsules. Angew. Chem. Int. Ed. 46, pp. 6996-7000, 2007b. 124 Eriksen, J. and Foote, C.S. Electron-Transfer Fluorescence Quenching and Exciplexes of Cyano-substituted Anthracenes. J. Phys. Chem. 82, pp. 2659-2662, 1978. Feng, Z., Wang, Z., Gao, C. and Shen, J. Direct Covalent Assembly to Fabricate Microcapsules with Ultrathin Walls and High Mechanical Strength. Adv. Mater. 19, pp. 3687-3691, 2007. Fischlechner, M., Zschömig, O., Hofmann, J. and Donath, E. Engineering Virus Functionalities on Colloidal Polyelectrolyte Lipid Composites. Angew. Chem. Int. Ed. 44, pp. 2892-2895, 2005. Fleer, G.J., Cohen Stuart, M.A., Scheutjens, J.M.H.M., Cosgrove, T. and Vincent, B. in Polymers at Interfaces (1st edn), Chapman & Hall, London, (ch.2.), pp. 27-42, 1993. Gao, C., Donath, E., Möhwald, H. and Shen, J. Spontaneous Deposition of Water-Soluble Substances into Microcapsules: Phenomenon, Mechanism and Application. Angew. Chem., Int. Ed. 41, pp. 3789-3793, 2002a. Gao, C., Liu, X., Shen, J. and Möhwald, H. Spontaneous Deposition of Horseradish Peroxidase into Polyelectrolyte Multilayer Capsules to Improve its Activity and Stability. Chem. Commun. 17, pp.1928-1929, 2002b. Gaponik, N., Radtchenko, I.L., Sukhorukov, G.B., Weller, H. and Rogach, A.L. Toward Encoding Combinatorial Libraries: Charge-Driven Microencapsulation of Semiconductor Nanocrystals Luminescing in the Visible and Near IR. Adv. Mater. 14, pp. 879-882, 2002. Georgieva, R., Moya, S., Hin, M., Mitlöhner, R., Donath, E., Kiesewetter, H., Möhwald, H. and Bäumler, H. Permeation of Macromolecules into Polyelectrolyte Microcapsules. Biomacromolecules 3, pp. 517-524, 2002. Ghan, R., Shutava, T., Patel, A., John, V.T. and Lvov, Y. Enzyme-Catalyzed Polymerization of Phenols within Polyelectrolyte Microcapsules. Macromolecules 37, pp. 4519-4524, 2004. Gittins, D.I. and Caruso, F. Multilayered Polymer Nanocapsules Derived from Gold Nanoparticle Templates. Adv. Mater. 12, pp. 1947-1949, 2000. Gittins, D.I. and Caruso, F. Tailoring the Polyelectrolyte Coating of Metal Nanoparticles. J. Phys. Chem. B. 105, pp. 6846-6852, 2001. Glinel, K., Sukhorukov, G.B., Möhwald, H., Khrenov, V. and Tauer, K. Thermosensitive Hollow Capsules Based on Thermosensitive Polyelectrolytes. Macromol. Chem. Phys. 204, pp. 1784-1790, 2003. Grellmann, K.H., Watkins, A.R. and Weller, A. Electron Transfer Mechanism of Fluorescence Quenching in Polar Solvents. II. Tetracyanoethylene and Tetracyanobenzene as Quenchers. J. Phys. Chem. 76, pp. 3132-3137, 1972. 125 Hoffman, A.S. Hydrogels for Biomedical Applications. Adv. Drug Delivery Rev. 43, pp. 3-12, 2002. Hsu, M.F., Nikolaides, M.G., Dinsmore, A.D., Bausch, A.R., Gordon, V.D., Chen, X., Hutchison, J.W. and Weitz, D.A. Self-assembled Shells Composed of Colloidal Particles: Fabrication and Characterization. Langmuir 21, pp. 2963-2970, 2005. Ibarz, G., Dähne, L., Donath, E. and Möhwald, H. Resealing of Polyelectrolyte Capsules after Core Removal. Macromol. Rapid Commun. 23, pp. 474-478, 2002. Ioan, C. E., Aberle, T. and W. Burchard. Structure Properties of Dextran. 2. Dilute Solution. Macromolecules 33, pp. 5730-5739, 2000. Itoh, Y., Matsusaki, M., Kida, T. and Akashi, M. Enzyme-Responsive Release of Encapsulated Proteins from Biodegradable Hollow Capsules. Biomacromolecules 7, pp. 2715-2718, 2006. Jaber, J.A. and Schlenoff, J.B. Recent Developments in the Properties and Applications of Polyelectrolyte Multilayers. Curr. Opin. Colloid Interface Sci. 11, pp. 324-329, 2006. Johnston, A.P.R., Read, E.S. and Caruso, F. DNA Multilayer Films on Planar and Colloidal Supports: Sequential Assembly of Like-Charged Polyelectrolytes. Nano Lett. 5, pp. 953-956, 2005. Joki, T., Machluf, M., Atala, A., Zhu, J., Seyfried, N.T., Dunn, I.F., Abe, T., Carroll, R.S. and Black, P.M. Continuous Release of Endostatin From Microencapsulated Engineered Cells for Tumor Therapy. Nat. Biotechnol. 19, pp. 35-39, 2001. Kabanov, V.A., Skobeleva, V.B., Rogacheva, V.B. and Zezin, A.B. Sorption of Proteins by Slightly Cross-Linked Polyelectrolyte Hydrogels: Kinetics and Mechanism. J. Phys. Chem. B 108, pp. 1485-1490, 2004. Kabanov, V.A., Zezin, A.B., Rogacheva, V.B. and Prevish, V.A. Active Transport of Linear Polyions in Oppositely Charged Swollen Polyelectrolyte Networks. Makromol. Chem. 190, pp. 2211-2216, 1989. Kamineni, V.K., Lvov, Y.M. and Dobbins, T.A. Layer-by-Layer Nanoassembly of Polyelectrolytes Using Formamide as the Working Medium. Langmuir 23, pp. 7423-7427, 2007. Kim, J.W., Jung, M.O., Kim, Y.J., Ryu, J.H., Kim, J., Chang, I.S., Lee, O.S. and Suh, K.D. Stabilization of Enzyme by Exclusive Volume Effect in Hydrophobically Controlled Polymer Microcapsules. Macromol. Rapid Commun. 26, pp. 1258-1261, 2005. Kim, J., Fernández-Nieves, A., Dan, N., Utada, A.S., Marquez, M. and Weitz, D.A. Colloidal Assembly Route for Responsive Colloidosomes with Tunable Permeability. Nano Lett. 7, pp. 2876-2880, 2007. 126 Köhler, K. and Sukhorukov, G.B. Heat Treatment of Polyelectrolyte Multilayer Capsules: A Versatile Method for Encapsulation. Adv. Funct. Mater. 17, pp. 2053-2061, 2007. Kozlovskaya, V., Ok, S., Sousa, A., Libera, M. and Sukhishvili, S.A. Hydrogen-Bonded Polymer Capsules Formed by Layer-by-Layer Self-Assembly. Macromolecules 36, pp. 8590-8592, 2003. Kozlovskaya, V. and Sukhishvili, S.A. pH-Controlled Permeability of Layered HydrogenBonded Polymer Capsules. Macromolecules 39, pp. 5569-5572, 2006. Kreft, O., Georgieva, R., Bäumler, H., Steup, M., Müller-Röber, B., Sukhorukov, G.B. and Möhwald, H. Red Blood Cell Templated Polyelectrolyte Capsules: A Novel Vehicle for the Stable Encapsulation of DNA and Proteins. Macromol. Rapid Commun. 27, pp. 435-440, 2006. Kreft, O., Javier, A.M., Sukhorukov, G.B. and Parak, W.J. Polymer Microcapsules as Mobile pH-Sensors. J. Mater. Chem. 17, pp. 4471-4476, 2007a. Kreft, O., Prevot, M., Möhwald, H. and Sukhorukov, G.B. Shell-in-Shell Microcapsules: A Novel Tool for Integrated, Spatially Confined Enzymatic Reactions. Angew. Chem. Int. Ed. 46, pp. 5605-5608, 2007b. Labropoulos K.C., Niesz, D.E., Danforth, S.C. and Kevrekidis, P.G. Dynamic Rheology of Agar Gels: Theory and Experiments. Part I. Development of a Rheological Model. Carbohydr. Polym. 50, pp. 393-406, 2002. Ladam, G., Schaad, P., Voegel, J.C., Schaaf, P., Decher, G. and Cuisinier, F. In Situ Determination of the Structural Properties of Initially Deposited Polyelectrolyte Multilayers. Langmuir 16, pp. 1249-1255, 2000. Laib, S. and Routh, A. F. Fabrication of Colloidosomes at Low Temperature for the Encapsulation of Thermally Sensitive Compounds. J. Colloid Interface Sci. 317, pp. 121129, 2008. Lavergne, F., Cot, D. and Ganachaud, F. Polymer Microcapsules with “Foamed” Membranes. Langmuir 23, pp. 6744-6753, 2007. Lee, H., Jeong, Y. and Park T.G. Shell Cross-Linked Hyaluronic Acid/Polylysine Layerby-Layer Polyelectrolyte Microcapsules Prepared by Removal of Reducible Hyaluronic Acid Microgel Cores. Biomacromolecules 8, pp. 3705-3711, 2007. Levy, T., Déjugnat, C. and Sukhorukov, G.B. Polymer Microcapsules with CarbohydrateSensitive Properties. Adv. Funct. Mater. 18, pp. 1586-1594, 2008. Lin, Y., Skaff, H., Emrick, T., Dinsmore, A.D. and Russell, T.P. Nanoparticles Assembly and Transport at Liquid-Liquid Interfaces. Science 299, pp. 226-229, 2003. 127 Loxley, A. and Vincent, B. Preparation of Poly(methylmethacrylate) Microcapsules with Liquid Cores. J. Colloid Interface Sci. 208, pp. 49-62, 1998. Lvov, Y., Antipov, A.A., Mamedov, A., Möhwald, H. and Sukhorukov, G.B. Urease Encapsulation in Nanoorganized Microshells. Nano Lett. 1, pp. 125-128, 2001. Mak, W.C., Cheung, K.Y. and Trau, D. Diffusion Controlled and Temperature Stable Microcapsule Reaction Compartments for High-Throughput Microcapsule-PCR. Adv. Funct. Mater. 18, pp. 2930-2937, 2008a. Mak, W.C., Cheung, K.Y. and Trau, D. Influence of Different Polyelectrolytes on Layerby-Layer Microcapsule Properties: Encapsulation Efficiency and Colloidal and Temperature Stability. Chem. Mater. 20, pp. 5475-5484, 2008b. Mak, W.C., Bai, J.H., Chang, X. Y. and Trau, D. Matrix-Assisted Colloidosome Reverse Layer-by-Layer Encapsulating Biomolecules in Hydrogel Microcapsules with Extremely High Efficiency and Retention Stability. Langmuir. 25, pp. 769-775, 2009. Murua, A., Castro, M., Orive, G., Hernández, R.M. and Pedraz, J.L. In Vitro Characterization and In Vivo Functionality of Erythropoietin-Secreting Cells Immobilized in Alginate-Poly-L-Lysine-Alginate Microcapsules. Biomacromolecules 8, pp. 3302-3307, 2007. Nii, T. and Ishii, F. Encapsulation Efficiency of Water-Soluble and Insoluble Drugs in Liposomes Prepared by the Microencapsulation Vesicle Method. Int. J. Pharm. 298, pp. 198-205, 2005. Noble, P.F., Cayre, O.J., Alargova, R.G., Velev, O.D. and Paunov, V.N. Fabrication of “Hairy” Colloidosomes with Shells of Polymeric Microrods. J. Am. Chem. Soc. 126, pp. 8092-8093, 2004. Ochs, C.J., Such, G.K., Städler, B. and Caruso, F. Low-Fouling, Biofunctionalized ,and Biodegradable Click Capsules. Biomacromolecules 9, pp. 3389-3396, 2008. Oh, K.T., Bronich, T.K., Kabanov, V.A. and Kabanov, A.V. Block Polyelectrolyte Networks from Poly(acrylic acid) and Poly(ethylene oxide): Sorption and Release of Cytochrome C. Biomacromolecules 8, pp. 490-497, 2007. Olson, F., Hunt, C.A., Szoka, F.C., Vail, W.J. and Papahadjopoulos, D. Preparation of Liposomes of Defined Size Distribution by Extrusion Through Polycarbonate Membranes. Biochim. Biophys. Acta 557, pp. 9-23, 1979. Park, M., Deng, S. and Advincula, R.C. Sustained Release Control via Photo-CrossLinking of Polyelectrolyte Layer-by-Layer Hollow Capsules. Langmuir 21, pp. 52725277, 2005. 128 Petrov, A.I., Volodkin, D.V. and Sukhorukov, G.B. Protein-Calcium Carbonate Coprecipitation: A Tool for Protein Encapsulation. Biotechnol. Prog. 21, pp. 918-625, 2005. Peyratout, C.S. and Dähne, L. Tailor-Made Polyelectrolyte Microcapsules: From Multilayers to Smart Containers. Angew. Chem. Int. Ed. 43, pp. 3762-3783, 2004. Prakash, S. and Chang, T. M. S. Microencapsulated Genetically Engineered Live E. coli DH5 Cells Administered Orally to Maintain Normal Plasma Urea Level in Uremic Rats. Nat. Med. 2, pp. 883-887, 1996. Qiu, X., Leporatti, S., Donath, E. and Möhwald, H. Studies on the Drug Release Properties of Polysaccharide Multilayers Encapsulated Ibuprofen Microparticles. Langmuir 17, pp. 5375-5380, 2001. Quinn, J.F., Johnston, A.P.R., Such, G.K., Zelikin, A.N. and Caruso, F. Next Generation, Sequentially Assembled Ultrathin Films: Beyond Electrostatics. Chem. Soc. Rev. 36, pp. 707-718, 2007. Radtchenko, I.L, Sukhorukov, G.B., Leporatti, S., Khomutov, G.B., Donath, E. and Möhwald, H. Assembly of Alternated Multivalent Ion/Polyelectrolyte Layers on Colloidal Particles. Stability of the Multilayers and Encapsulation of Macromolecules into Polyelectrolyte Capsules. J. Colloid Interface Sci. 230, pp. 272-280, 2000. Ramarao, C., Ley, S.V., Smith, S.C., Shirley, I.M. and DeAlmeida, N. Encapsulation of Palladium in Polyurea Microcapsules. Chem. Commun. 10, pp. 1132-1133, 2002. Reis, C.P., Neufeld, R.J., Vilela, S., Ribeiro, A.J. and Veiga, F. Review and Current Status of Emulsion/Dispersion Technology using an Internal Gelation Process for the Design of Alginate Particles. J. Microencapsul. 23, pp. 245-257, 2006. Rijiravanich, P., Somasundrum, M. and Surareungchai, W. Femtomolar Electrochemical Detection of DNA Hybridization Using Hollow Polyelectrolyte Shells Bearing Silver Nanoparticles. Anal. Chem., 80, pp. 3904-3909, 2008. Rogacheva, V.B., Prevysh, V.A., Zezin, A.B. and Kabanov, V.A. Interpolymer Reactions between Network and Linear Polyelectrolytes. Polym. Sci. U.S.S.R. 30, pp. 2262-2270. 1988. Rosanberg, R.T. and Dan, N.R. Controlling Surface Porosity and Release from Hydrogels Using a Colloidal Particle Coating. J. Colloid Interface Sci. 349, pp. 498-504, 2010. Rose, R., Zelikin, A.N., Johnston, A.P.R., Sexton, A., Chong, S., Cortez, C., Mulholland, W., Caruso, F. and Kent, S.J. Binding, Internalization, and Antigen Presentation of Vaccine-Loaded Nanoengineered Capsules in Blood. Adv. Mater. 20, pp. 4698-4703, 2008. 129 Rusling, J.F., Hvastkovs, E.G., Hull, D.O. and Schenkman, J.B. Biochemical Applications of Ultrathin Films of Enzymes, Polyions and DNA. Chem. Comm. 2, pp. 141-154, 2008. Serizawa, T., Kamimura, S., Kawanishi, N. and Akashi, M. Layer-by-Layer Assembly of Poly(vinyl alcohol) and Hydrophobic Polymers Based on Their Physical Adsorption on Surfaces. Langmuir 18, pp. 8381-8385, 2002. Shchukin, D.G. and Sukhorukov G.B. Nanoparticle Synthesis in Engineered Organic Nanoscale Reactors. Adv. Mater. 16, pp. 671-682, 2004. Shenoy, D.B., Antipov, A.A., Sukhorukov, G.B. and Möhwald, H. Layer-by-Layer Engineering of Biocompatible, Decomposable Core-Shell Structures. Biomacromolecules 4, pp. 265-272, 2003. Shutava, T.G., Kommireddy, D.S. and Lvov, Y.M. Layer-by-Layer Enzyme/Polyelectrolyte Films as a Functional Protective Barrier in Oxidizing Media. J. Am. Chem. Soc. 128, pp. 9926-9934, 2006. Skirtach, A.G., Antipov, A.A., Shchukin, D.G. and Sukhorukov, G.B. Remote Activation of Capsules Containing Ag Nanoparticles and IR Dye by Laser Light. Langmuir 20, pp. 6988-6992, 2004. Skirtach, A.G., Dejugnat, C., Braun, D., Susha, A.S., Rogach, A.L., Parak, W.J., Möhwald, H. and Sukhorukov, G.B. The Role of Metal Nanoparticles in Remote Release of Encapsulated Materials. Nano Lett. 5, pp. 1371-1377, 2005. Skirtach, A.G., Javier, A.M., Kreft, O., Köhler, K., Alberola, A.P., Möhwald, H., Parak, W.J. and Sukhorukov G.B. Laser-Induced Release of Encapsulated Materials inside Living Cells. Angew. Chem. Int. Ed. 45, pp. 4612-4617, 2006. Srivastava, R., Brown, J.Q., Zhu, H. and McShane, M.J. Stable Encapsulation of Active Enzyme by Application of Multilayer Nanofilm Coatings to Alginate Microspheres. Macromol. Biosci. 5, pp. 717-727, 2005. Such, G.K., Quinn, J.F., Quinn, A., Tjipto, E. and Caruso, F. Assembly of Ultrathin Polymer Multilayer Films by Click Chemistry. J. Am. Chem. Soc. 128, pp. 9318-9319, 2006. Such, G.K., Tjipto, E., Postma, A., Johnston, A.P.R. and Caruso, F. Ultrathin, Responsive Polymer Click Capsules. Nano Lett. 7, pp. 1706-1710, 2007. Sukhorukov, G.B., Donath, E., Lichtenfeld, H., Knippel, E., Knippel, M., Budde, A. and Möhwald, H. Layer-by-Layer Self Assembly of Polyelectrolytes on Colloidal Particles. Colloids Surf., A 137, pp. 253-266, 1998. Sukhorukov G.B., Brumen, M., Donath, E. and Möhwald, H. Hollow Polyelectrolyte Shells: Exclusion of Polymers and Donnan Equilibrium. J. Phys. Chem. B 103, pp. 64346440, 1999. 130 Sukhorukov, G.B., Donath, E., Moya, S., Susha, S., Voigt, A.S., Hartmann, J. and Möhwald, H. Microencapsulation by Means of Step-Wise Adsorption of Polyelectrolytes. J. Microencapsulation 17, pp. 177-185, 2000. Sun, Q. and Deng, Y. In Situ Synthesis of Temperature-Sensitive Hollow Microsphere via Interfacial Polymerization. J. Am. Chem. Soc. 127, pp. 8274-8275, 2005. Tang, Z., Wang, Y., Podsiadlo, P. and Kotov, N.A. Biomedical Applications of Layer-byLayer Assembly: From Biomimetics to Tissue Engineering. Adv. Mater. 18, pp. 32033224, 2006. Tong, W., Gao, C. and Möhwald, H. Single Polyelectrolyte Microcapsules Fabricated by Gluteraldehyde-Mediated Covalent Layer-by-Layer Assembly. Macromol. Rapid Commun. 27, pp. 2078-2083, 2006. Torini, L., Argillier, J.F. and Zydowicz, N. Interfacial Polycondensation Encapsulation in Miniemulsion. Macromolecule 38, pp. 3225-3236, 2005. Trau, D., Yang, W.J., Seydack, M., Caruso, F., Yu, N.T. and Renneberg, R. Nanoencapsulated Microcrystalline Particles for Superamplified Biochemical Assays. Anal. Chem., 74, pp. 5480-5486, 2002. Trau, D. and Renneberg, R. Encapsulation of Glucose Oxidase Microparticles within a Nanoscale Layer-by-Layer Film: Immobilization and Biosensor Applications. Biosens. Bioelectron. 18, pp. 1491-1499, 2003. Velev, O.D., Furusawa, K. and Nagayama, K. Assembly of Latex Particles by Using Emulsion Droplets as Templates. 1. Microstructured Hollow Spheres. Langmuir 12, pp. 2374-2384, 1996. Volodkin, D.V., Petrov, A.I., Prevot, M. and Sukhorukov, G.B. Matrix Polyelectrolyte Microcapsules: New System for Macromolecule Encapsulation. Langmuir 20, pp. 33983406, 2004a. Volodkin, D.V., Larionova, N.I. and Sukhorukov, G.B. Protein Encapsulation via Porous CaCO3 Microparticles Templating. Biomacromolecules 5, pp. 1962-1972, 2004b. Wang, Y. and Caruso, F. Enzyme Encapsulation in Nanoporous Silica Spheres. Chem. Commun. 13, pp. 1528-1529, 2004. Wang, D., Duan, H., and Möhwald, H. The Water/Oil Interface: the Emerging Horizon for Self-Assembly of Nanoparticles. Soft Matter 1, pp. 412-416, 2005. Wang, K., He, Q., Yan, X., Cui, Y., Qi, W., Duan, L. and Li, J. Encapsulated Photosensitive Drugs by Biodegradable Microcapsules to Incapacitate Cancer Cells. J. Chem. Mater. 17, pp. 4018-4021, 2007. 131 Wang, Z., Feng, Z. and Gao, C. Stepwise Assembly of the Same Polyelectrolytes Using Host-Guest Interaction to Obtain Microcapsules with Multiresponsive Properties. Chem. Mater. 20, pp. 4194-4199, 2008. Yang, S., Zhang, Y., Yuan, G., Zhang, X. And Xu, J. Porous and Nonporous Nanocapsules by H-Bonding Self-Assembly. Macromolecules 37, pp. 10059-10062, 2004. Ye, S., Wang, C., Liu, X., Tong, Z., Ren, B. and Zeng, F. New Loading Process and Release Properties of Insulin from Polysaccharide Microcapsules Fabricated Through Layer-by-Layer Assembly. J. Controlled Release 112, pp. 79-87, 2006. Yow, H.N. and Routh, A.F. Formation of Liquid Core-Polymer Shell Microcapsules. Soft Matter 2, pp. 940-949, 2006. Yow, H.N. and Routh, A.F. Release Profiles of Encapsulated Actives from Colloidosomes Sintered for Various Durations. Langmuir 25, pp. 159-166, 2009. Yu, A., Wang, Y., Barlow, E. and Caruso, F. Mesoporous Silica Particles as Templates for Preparing Enzyme-Loaded Biocompatible Microcapsules. Adv. Mater. 17, pp. 1737-1741, 2005. Yu, L., Gao, Y., Yue, X., Liu, S. and Dai, Z. Novel Hollow Microcapsules Based on IronHeparin Complex Multilayers. Langmuir 24, pp. 13723-13729, 2008. Zebli, B., Susha, A.S., Sukhorukov, G.B., Rogach, A.L. and Parak, W.J. Magnetic Targeting and Cellular Uptake of Polymer Microcapsules Simultaneously Functionalized with Magnetic and Luminescent Nanocrystals. Langmuir 21, pp. 4262-4265, 2005. Zelikin, A.N., Quinn, J.F. and Caruso, F. Disulfide Cross-Linked Polymer Capsules: En Route to Biodeconstructible Systems. Biomacromolecules 7, pp. 27-30, 2006a. Zelikin, A.N., Li, Q. and Caruso, F. Degradable Polyelectrolyte Capsules Filled with Oligonucleotide Sequence. Angew. Chem. Int. Ed. 45, pp. 7743-7745, 2006b. Zelikin, A.N., Becker, A.L., Johnston, A.P.R., Wark, K.L., Turatti, F. and Caruso, F. A General Approach for DNA Encapsulation in Degradable Polymer Microcapsules. ACS Nano 1, pp. 63-69, 2007. Zezin, A.B., Rogacheva, V.B. and Kabanov, V.A. Interpolymer Complexes Formed by Polyelectrolyte Gels. J. Intell. Mater. Syst. Struct. 5, pp. 144-146, 1994. Zhang, Y., Yang, S., Guan, Y., Cao, W. and Xu, J. Fabrication of Stable Hollow Capsules by Covalent Layer-by-Layer Self Assembly. Macromolecules 36, pp. 4238-4240, 2003. Zhang, H., Tumarkin, E., Peerani, R., Nie, Z., Sullan, R.M.A, Walker, G.C. and Kumacheva, E. Microfluidic Production of Biopolymer Microcapsules with Controlled Morphology. J. Am. Chem. Soc. 128, pp. 12205-12210, 2006. 132 Zhang, X., Chen, H. and Zhang, H. Layer-by-Layer Assembly: From Conventional to Unconventional Methods. Chem. Comm. 14, pp. 1395-1405, 2007. Zhigaltsev, I.V., Maurer, N., Akhong, Q., Leone, R., Leng, E., Wang, J., Semple, S.C. and Cullis, P.R. Liposome-Encapsulated Vincristine, Vinblastine and Vinorelbine: A Comparative Study of Drug Loading and Retention. J. Controlled Release 104, pp. 103111, 2005. Zhou, Q.Z., Wang, L.Y., Ma, G.H. and Su, Z.G. Preparation of Uniform-sized agarose beads by microporous membrane emulsification technique. J. Colloid Interface Sci. 311, pp. 118-127, 2007. Zhou, Q.Z., Ma, G.H. and Su, Z.G. Effect of membrane parameters on the size and uniformity in preparing agarose beads by premix membrane emulsification. J. Membr. Sci. 326, pp. 694-700, 2009. Zhu, H. and McShane, M.J. Macromolecule Encapsulation in Diazoresin-Based Hollow Polyelectrolyte Microcapsules. Langmuir 21, pp. 424-430, 2005. 133 [...]... phase for the fabrication of hydrogel core- shell materials could achieve high encapsulation efficiency of biomolecules In addition, using a hydrophobic solvent for fabrication of hydrogel core- shell materials is an uncharted area and where different polymer interactions or phenomenon can be explored, thus resulting in fabrication of novel core- shell materials The objective of this work is therefore to... technique for the fabrication of core- shell and hollow microcapsules for biomedical application and will be reviewed in the following section 12 Chapter 2 – Literature Review 2.2.5.1 Core- Shell LbL Microcapsules The fabrication of core- shell materials via the LbL technique is a procedure that is technically simple and easy to perform The material desired as the core is usually utilized as the template for the. .. the ability to contain a high percentage of water that provides a favorable environment for the encapsulated biomolecules Loss of biomolecules during the phase of LbL polymer deposition and “semi-permeable membrane” fabrication is unfortunately too significant and results in low encapsulation efficiency of the biomolecules (Mak et al., 2008a) 2 Chapter 1 - Introduction The use of an organic phase for. .. availability of these templates with near uniform dimensions and in the region of nano – micrometers for fabrication of core- shell materials 2.2.5.2 Hollow LbL Microcapsules Fabrication of hollow LbL microcapsules requires the exact procedures for the fabrication of core- shell LbL microcapsules with the inclusion of an additional template sacrificial step that removes the template and thus resulting in hollow... for the Advancement of Science) 10 Figure 2.3 Schematic illustration of the Reverse -Phase LbL (RP-LbL) technique for encapsulation of biomolecules in an organic solvent i & ii) Deprotonation of cationic and protonation of the anionic polyelectrolyte and dissolution in an organic solvent iii) Preparation of a suspension of highly water soluble biomolecules in an organic solvent iv) Deposition of the. .. interaction between the encapsulated materials (allowing for rapid intercalating reactions) and the incorporation of magnetic nanoparticles within the architecture of the microcapsules for specific location targeting (Zebli et al., 2005) In this chapter, different techniques for the fabrication of microcapsules will be discussed Following, the fabrication of hollow or core- shell microcapsules via the Layer-by-Layer... Mechanism into the Matrices of Agarose Microbeads 93 7.3.4 Organic Phase Fabrication of Core- Shell Materials via the Inwards Diffusion and Deposition of niPA Layers into the Matrices of Agarose Microbeads 99 7.3.5 Spatial Distribution, Retention Efficiency and Release of Encapsulated Dextran from Core- Shell Materials Fabricated via Inwards Deposition of niPA 102 7.3.6 Encoding of Agarose Microbeads... 2001) Alternatively, the template could be microcrystals of the biomolecules where the deposition of the polymers is done in special conditions to prevent dissolution of the microcrystals (Beyer et al., 2007; Trau and Renneberg, 2003) Direct deposition of the polymers would encapsulate the biomolecules and thereby forms the microcapsules However, removal of the template or transferring the encapsulated... the surface of the template to form the first polymer layer Washing is performed to remove excess PSS before coating of a positively charge polyelectrolyte, poly (allylamine hydrochloride) (PAH), to form the second polymer layer Washing is again performed before the cycle is repeated until the number of desired layers is obtained (Decher, 1997, Reproduced by permission of The American Association for. .. American Association for the Advancement of Science) Currently, the most popular technology for fabrication of hollow or core- shell microcapsules is via the Layer-by-Layer technology (Decher, 1997) This technology was first introduced in 1991 and is relatively simple as it relied on the interaction of oppositely charged polymers for the formation of the microcapsules The process behind the LbL 10 Chapter . ORGANIC PHASE FABRICATION OF CORE- SHELL MATERIALS FOR THE ENCAPSULATION OF BIOMOLECULES BAI JIANHAO (B.Eng. (Hons.), NUS) A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF. materials and encapsulation of biomolecules is rarely demonstrated. Therefore, this PhD work involves the novel organic phase fabrication of core- shell materials and encapsulation of biomolecule. Vessel of Biomolecules. 27 3.2.2 Specific Aim 2 – Selection of Organic Solvent and Polymers for Organic Phase Fabrication of Core- Shell Materials 28 3.2.3 Specific Aim 3 – Organic Phase Fabrication