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Engineering of Amphiphilic Block Copolymers for Drug and Gene Delivery
... On the other hand, the oil-in-water emulsion is used to encapsulate lipophilic substances. In this case, the continuous phase is established by hydrophilic solutions [50], whereas surfactants with high hydrophilic-lipophilic balance are used to stabilize such emulsions [51]. Besides the appropriateness of surfactant, other factors that need to be optimized are electro-spinnable continuous phase polymer and properties of the droplet phase. ...
A chronic wound in diabetic patients is a major public health concern with socioeconomic and clinical manifestations. The underlying medical condition of diabetic patients deteriorates the wound through physiological, metabolic, molecular, and cellular pathologies. Consequently, a wound enters a vicious pathological inflammatory cycle. Many therapeutic approaches are in practice to manage diabetic wounds hence ensuring the regeneration process. Polymer-based biomaterials have come up with high therapeutic promises. Many efforts have been devoted, over the years, to build an effective wound healing material using polymers. The electrospinning technique, although not new, has turned out to be one of the most effective strategies in building wound healing biomaterials due to the special structural advantages of electrospun nanofibers over the other formulations. In this review, careful integration of all electrospinning approaches has been presented which will not only give an insight into the current updates but also be helpful in the development of new therapeutic material considering pathophysiological conditions of a diabetic wound.
... In addition, when formed or prepared from aqueous solutions, such ultrafine particles often display various forms of physical instability (i.e., aggregation, recrystallization and/or Ostwald ripening) owing to their high surface-to-volume ratio and consequential high surface energy [5]. Circumvention of such stability problems with nanoformulations would require the use of stabilizers such as polymers [6,7], electrolytes [8], lipids [9,10] and surfactants [11,12]. ...
The influence of critical operating parameters on the Flash Nanoprecipitation (FNP) and resulting material properties of curcumin (CUR) nanoparticles has been evaluated using a confined impinging jets-with-dilution mixer (CIJ-D-M). It has been shown that the mixing rate, molecular weight of polymeric stabilizer (i.e., polyethylene glycol-b-poly (DL-lactide) di-block copolymer; PEG-PLA) and drug-to-copolymer mass ratio all exert a significant impact on the particle size and stability of the generated nanosuspensions. The attainable mean particle size and span of the nanoparticles through optimization of these process parameters were approximately 70 nm and 0.85 respectively. However, the optimized nanosuspension was only stable for about two hours after preparation. Co-formulation with polyvinylpyrrolidone (PVP) substantially extended the product lifespan to 5 days at ambient conditions and two weeks at 4˚C. Results from zeta potential measurement and X-ray photoelectron spectroscopy (XPS) suggested that the enhanced stability is probably due to the formation of an additional protective barrier by PVP around the particle surface, thereby suppressing the dissociation of PEG-PLA from the particles and preventing CUR leakage from inside. Long-term storage stability (>1 year) could be achieved by lyophilization of the optimized nanosuspension with Kleptose (hydroxypropyl-β-cyclodextrin), which was shown to be the only effective lyoprotectant among all the ones tested for the CUR nanoparticles. At an optimal concentration of Kleptose (1.25% w/v), the redispersibility (Sf/Si; ratio of the final and initial particle sizes) and encapsulation efficiency of lyophilized CUR nanoparticles were about 1.22 and 94%, respectively.
Copyright © 2015. Published by Elsevier B.V.
Recent years have witnessed increasing interest in nano-sized drug delivery systems in the field of medicine as they can improve the pharmacokinetic and biodistribution characteristics of drugs, thereby reducing side effects and improving patient compliance. Since the first approval of a liposome containing doxorubicin in 1995, many other nano-sized platforms, such as polymer–drug conjugates, dendrimers, or inorganic nanoparticles, among others, have entered clinical trials or are already available. This book chapter offers a general overview of the different aspects that should be considered for the rational design of nanosystems for drug delivery, including examples of organic and inorganic nanosystems that can be used for this purpose. The possible translation from this basic research to the healthcare market is also discussed.
Several non-permanent polycations possessing substantial buffering capacity below physiological pH, such as lipopolyamines and polyethylenimines, are efficient transfection agents per se, i.e. without the addition of lysosomotropic bases, or cell targeting, or membrane disruption
agents. These vectors have been shown to deliver genes as well as oligonucleotides both in vitro and in vivo. Our hypothesis is that their efficiency relies on extensive endosome swelling and rupture that provides an escape mechanism for the polycation/DNA particles.
Benzylamine, hexylamine and aniline‐initiated polymerizations of D , L ‐phenylalanine and L ‐phenylalanine N ‐carboxyanhydride ( D , L ‐Phe‐NCA and L ‐Phe‐NCA) were performed in various solvents. The isolated polypeptides were characterized by MALDI‐TOF mass spectroscopy. Exclusively linear polypeptides having one amide and one amino endgroup were found, when the polymerizations were conducted in a closed reaction vessel with dioxane or sulfolane as reaction media. Traces of water competed with aniline as initiator, when the reaction vessel was closed with a drying tube. In N , N ‐dimethylformamide (DMF) formyl endgroups were formed at polymerization temperatures of 60 °C. In DMF and N ‐methylpyrrolidone, cyclic oligo‐ and polypeptides were formed by a solvent‐induced polymerization initiated by zwitterions, which may compete with the primary amine‐initiated polymerizations.
MALDI‐TOF mass spectrum of a poly( D , L ‐phenylalanine) polymerized in NMP without addition of an initiator.
magnified image MALDI‐TOF mass spectrum of a poly( D , L ‐phenylalanine) polymerized in NMP without addition of an initiator.
The reaction of β-alanine N-carboxyanhydride (β-Ala NCA) (1) with various primary and secondary amines was investigated at room temperature. Three reaction products were found: poly(β-alanine), β-alaninamides, and N′-substituted β-ureidopropionic acids. The mole ratio of these reaction products depends on the reaction conditions, above all on the NCA/amine ratio, and on the nucleophilicity/basicity ratio of the amines. All amines with pKa-values > 8,0, including the amino end-groups of the growing chains, cause the formation of β-isocyanatocarboxylate ions by deprotonation of the β-NCAs, and thus, the formation of β-ureidocarboxylic acids. If the chain ends are involved, this reaction sequence is a termination step, and it is this termination step which is responsible, for that high polymerization degrees (> 60) are not accessible by a primary or secondary amine-initiated polymerization of β-NCAs. The β-ureidocarboxylic acid end-groups are detectable in the 13C NMR spectra of the poly(β-amide)s.
Recently, polypeptide hybrid polymers, particularly poly(ethylene glycol) (PEG)-polypeptide
block copolymers, have been attracting significant interest for polymeric therapeutics, such as drug
and gene delivery systems, utilizing their most relevant feature, that is the formation of micelles
with adistinguished core-shell architecture. Of particular interest in the polypeptides is
that avariety of functional groups, such as carboxyl groups and amino groups, are available
as aside chain, and that they have propensities of low toxicity and biodegradability. The segregated
polypeptide core of the micelle embedded in the hydrophilic palisade serves as areservoir for
avariety of drugs as well as of genes with diverse characteristics. The micelles have been
developed with various functions, such as biocompatibility, stimuli- and environment-sensitivity,
and targetability, aimed at their clinical use. Smart micelles have emerged as promising carriers
that enhance the effect of drugs and genes with minimal side effects. In this review, recent advances
in drug and gene delivery by polypeptide hybrid micelles, mostly accomplished in our group, are comprehensively
described. Focus is placed on the design of PEG-polypeptide hybrid block copolymers, starting from
the development of the drug-loading micelle systems to current efforts to establish agene delivery
system with apolyion complex (PIC) micelle, one of the most attractive topics in nanomedicine.
This article reviews recent developments in the polymerization of ar-amino acid-N-carboxyanhydrides (NCAs) to form polypeptides. Traditional methods used to polymerize these monomers are described, and limitations in the utility of these systems for the preparation of polypeptides with controlled molecular weights and narrow molecular weight distributions are discussed. The development of transition-metal-based initiators, which activate the monomers to form covalent active species, permits the formation of polypeptides via the living polymerization of NCAs. In these systems, polymer molecular weights are controlled by monomer-to-initiator stoichiometry, polydispersities are low, and block copolypeptides can be prepared. The scope and limitations of these initiators and their key features and mode of operation are described in detail in this highlight. (C) 2000 John Wiley & Sons, Inc.
Organotin compounds were found to lead to polymerization of N-carboxy anhydrides. The polymerization was studied in detail using γ-benzyl N-carboxyl-t-glutamate anhydride (BGA). Compounds such as tributyltin methoxide, bis(tributyltin)oxide, and N-tributyltin imidazole polymerized BGA while others like dibutyltin dichloride, which are Lewis acids, failed. Polymerization of BGA in dioxane at various monomer to dibutyltin dimethoxide ratios showed a first order reaction to monomer. The plot of In M0/M1 vs time showed two stage kinetics, the second one being faster. The pseudo first order rate constants were smaller than those for primary amine initiated polymerizations and much smaller than that for polymerization initiated by sodium methoxide. The molecular weights were independent of the monomer to initiator ratio both in dioxane and in DMF. In the reaction of an equimolar amount of tributyltin methoxide with NCA, the methyl ester of the amino acid was formed.
This article reviews recent developments in the polymerization of α-amino acid- N-carboxyanhydrides (NCAs) to form polypeptides. Traditional methods used to polymerize these monomers are described, and limitations in the utility of these systems for the preparation of polypeptides with controlled molecular weights and narrow molecular weight distributions are discussed. The development of transition-metal-based initiators, which activate the monomers to form covalent active species, permits the formation of polypeptides via the living polymerization of NCAs. In these systems, polymer molecular weights are controlled by monomer-to-initiator stoichiometry, polydispersities are low, and block copolypeptides can be prepared. The scope and limitations of these initiators and their key features and mode of operation are described in detail in this highlight. © 2000 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 38: 3011–3018, 2000
A novel structural triblock copolymer of poly(γ-benzyl-l-glutamic acid)-b-poly(ethylene oxide)-b-poly(ε-caprolactone) (PBLG-PEO-PCL) was synthesized by a new approach in the following three steps: (1) sequential anionic ring opening polymerization (ROP) of ethylene oxide and ε-caprolactone with an acetonitrile/potassium naphthalene initiator system to obtain a diblock copolymer CN-PEO-PCL with a cyano end-group; (2) conversion of the CN end-group into NH2 end-group by hydrogenation to obtain NH2-PEO-PCL; (3) ROP of γ-benzyl-l-glutamate-N-carboxyanhydrides (Bz-l-GluNCA) with NH2-PEO-PCL as macroinitiator to obtain the target triblock copolymer. The structures from CN-PEO precursor to the triblock copolymers were confirmed by FT-IR and 1H NMR spectroscopy, and their molecular weights were measured by gel permeation chromatography. The monomer of Bz-l-GluNCA can react almost quantitatively with the amino end-groups of NH2-PEO-PCL macroinitiator by ROP.
Directed toward pharmacological applications of lactose-carrying polystyrene, (poly (, PVLA), its body distribution, clearance from blood and specific binding to receptors have been investigated using radiolabelled PVLA. 125I-Labelled PVLA was prepared via copolymerization of (VLA) with 10 mol% of 4-(2-propenyl)phenyl acetate followed by radiolabelling of the latter component. When 125I-labelled PVLA was injected into rats through their tail veins, the radio-activity was distributed highly to liver, less to thyroid gland, cecum-large intestine, urine, feces and blood, and much less to lung, heart, kidney, spleen, pancreas, small intestine and urinary bladder. Its concentration to liver was visible by whole-body autoradiography. It was clarified that about 97% of PVLA was distributed to parenchymal liver cells and only 3% to nonparenchymal liver cells. The radioactivity in blood was decreased with time according to a biexponential curve. A two open compartment model is proposed on the basis of the pharmacokinetic analysis of the equation, which elucidated that PVLA migrated rapidly from blood to parenchymal liver cells. Specific binding between 125I-labelled PVLA and asialoglycoprotein receptors on parenchymal liver cells was demonstrated by its inhibition with asialofetuin. The bond dissociation constant estimated by Scatchard analysis was Kd= 1.4 × 10−9 M. The binding was as strong as those of several naturally occurring asialoglycoproteins. These properties of PVLA, as liver-specific targeting materials using galactose ligands as recognition signals to asialoglycoprotein receptors, are discussed with the conformational structures of PVLA which can carry drugs in their hydrophobic regions.
The aim of this study was to develop micelle-forming poly(ethylene oxide)-block-poly(ε-caprolactone) (PEO-b-PCL)-based block copolymers bearing functional side groups on the PCL block. Substituted monomer, i.e, α-benzyl carboxylate-ε-caprolactone, was synthesized by anionic activation of ε-caprolactone and further treatment with benzyl chloroformate. Successful substitution of benzyl carboxylate on ε-caprolactone monomer was evidenced by 1H NMR and mass spectroscopy. Ring-opening polymerization of α-benzyl carboxylate-ε-caprolactone with methoxy PEO (5000 g mol-1) as initiator and stannous octoate as catalyst was used to prepare PEO-b-poly(α-benzyl carboxylate ε-caprolactone) (PEO-b-PBCL). Further catalytic debenzylation of PEO-b-PBCL produced PEO-b-poly(α-carboxyl-ε-caprolactone) (PEO-b-PCCL). Ring-opening polymerization of a ε-caprolactone/α-benzyl carboxylate-ε-caprolactone mixture and further reduction of the product were also used to prepare block copolymers with various degrees of benzyl carboxylate or carboxyl group substitution. The calculated molecular weights determined by 1H NMR and gel permeation chromatography (GPC) for block copolymers were in good agreement with the theoretical values. The polydispersity of PEO-b-PBCL and PEO-b-PCCL block copolymers was 1.74 and 1.52, respectively. PEO-b-PBCL and PEO-b-PCCL block copolymers assembled to spherical micelles having average diameters of 62 and 20 nm based on dynamic light scattering (DLS) measurement, respectively. PEO-b-PBCL formed micelles at extremely low concentrations (cmc of 9.8 × 10-2 μM). The presence of carboxylic group on the PCCL block raised the cmc of PEO-b-PCCL to 1220 × 10-2 μM. For block copolymers with PCL-co-PCCL core structures, a decrease in cmc as well as an increase in size was observed as the level of PCL to PCCL was raised. Novel PEO-b-poly(ester) block copolymers with aromatic and reactive side groups on the polyester block have tremendous potential in the design of optimized carriers for the delivery of various therapeutic agents, as they can assemble to biodegradable nanoscopic micelles with chemically tailorable core structures.
A novel type of catalyst, tri-n-butylphosphine-metal carboxylate (metal: Ni, Co, Cr, Cd, or Mg) catalyst system, was found for the polymerization of γ-benzyl L-glutamate N-carboxyanhydride. Molecular weights of polymers obtained with this type of catalyst depend on the nature of metal component of the catalysts and follow the order of Ni > Co > Cd > Mg > Cr. The polymers obtained with the nickel acetate-tri-n-butylphosphine catalyst system in higher than 90% yield have molecular weights larger than 2 × 105 and relatively sharp molecular weight distributions: i.e., M̄w/M̄n = 1.4-2.1.
There is a wide-spread consensus that characteristics of drug vehicles determine the applicability of the site-specific delivery of drugs. This article focused on the promising features of block copolymer micelles as drug vehicles mimicking the natural carrier-systems with supramolecular structures (i.e. viruses and lipoproteins). Considerable discussions are made on the physicochemical characteristics of polymeric micelles in aqueous milieu with shedding light on earlier works done in the field. Advantageous features of polymeric micelles as drug vehicles are summarized as: (1) formation of environmentally-separated microcontainer of drugs through supramolecular assemblage, (2) installation of anchoring moiety on the surface, (3) duration in the biological compartment and (4) programmable chronological stability. Then, our recent work concerning polymeric micelles with anti-tumor activity is presented to demonstrate these advantageous features of polymeric micelles. Worth noticing is that higher anti-tumor activity was achieved by adriamycin-conjugated micelles compared with parental adriamycin, indicating that a considerable improvement in cancer chemotherapy is feasible by the use of appropriate vehicle systems.
The progress in both peptide and polymer chemistry has led to the preparation of “hybrids” to yield compounds with properties not achievable with the separate components. In this review we aim at describing the current synthetic methodologies for the preparation of peptide containing block copolymers, either being completely peptidic or as hybrids with synthetic polymer blocks. The different techniques to prepare these polymers are ordered by level of control over peptide sequence and hence their molecular structure. First, recent developments in NCA polymerization will be discussed, which enable the construction of well-defined high molecular weight peptide based block copolymers, albeit with no absolute control over amino acid composition. Solution phase peptide synthesis allows the preparation of small peptide sequences, which can be incorporated into the side chain or the main chain of hybrid polymer architectures. Application of solid phase peptide synthesis and the newly developed peptide ligation methods has resulted in an extension of the length of the peptide fragments that can be conveniently incorporated into hybrid polymer structures. Finally absolute control over amino acid sequence, combined with the ability to create high molecular weight species is accomplished with the application of protein engineering to the field of polymer science.
Amphiphilic poly(ethylene glycol)-b-polylactide (PEG/PLA) copolymers with an aldehyde group at one end and a methacryloyl group at the other chain end were synthesized by anionic polymerization. The efficiencies of the functionalization at both ends were almost quantitative. The amphiphilic block copolymers formed micelles in aqueous media. Acetal groups on the micelle surface were quantitatively converted to aldehyde groups by an acid treatment. The end methacryloyl group located in the core of the micelle was polymerized effectively to form core−shell-type nanoparticles having reactive aldehyde groups on the surface. The size of the reactive nanoparticle was 20−30 nm which was constant with temperatures up to 60 °C. The stability of the micelle was also confirmed by a sodium dodecyl sulfate (SDS) treatment. When SDS was added to the nanosphere solution to 20 mg/mL, the particle was not collapsed. The particle was stable enough even in organic solvents. This functionalized micelle having high stability is not only expected to have wide utilities in biomedical applications (including drug delivery, diagnosis, and surface modification through the coupling of bioactive substances) but also to be of great interest as a novel supramolecular architecture.
Dispersion polymerization of ε-caprolactone initiated with diethylaluminum alkoxide, carried out in the 1,4-dioxane:heptane (1:9 v/v) mixture at room temperature in the presence of poly(dodecyl acrylate)-g-poly(ε-caprolactone) used as the surface active agent, proceeds in two stages. In the first stage the growth of the poly(ε-caprolactone) chains is initiated in solution and, when the molecular weight of growing macromolecules comes close to 1000, the primary particles are nucleated and all propagating chains become incorporated into growing microspheres. In the second stage the polymerization process consists of propagation taking place inside microspheres into which monomer molecules diffuse from solution. During the first stage the apparent propagation rate constant is low and does not exceed 1 × 10-2 L·mol-1·s-1. In the second stage, due to the high local concentration of growing species confined in polymer particles, the apparent propagation rate constant becomes much higher. For monomer concentrations in the region from 3.9 × 10-1 mol·L-1 to 4.3 × 10-1 mol·L-1 and for initiator concentrations ranging from 3.4 × 10-3 mol·L-1 to 2.6 × 10-2 mol·L-1 the apparent propagation rate constant ( ) varied from 4.79 × 10-1 L·mol-1·s-1 to 6.5 × 10-2 L·mol-1·s-1.
The anionic polymerization of ∈-caprolactone in tetrahydrofuran with potassium tert-butoxide results in a living ring-chain equilibrium system. The product distribution is essentially determined by the entropy term, the lower cyclics being favored over the linear chains at higher dilution. Thus the equilibrium state is readily changed by dilution or concentration of the living system. The molar cyclization equilibrium constant decreased in proportion to the -2.5 power of the ring size, in accord with the Jacobson-Stockmayer theory, with the exception of the trimer which equilibrated at an appreciably lower concentration than expected. Terminating the reaction before establishment of the equilibrium provides direct evidence that the cyclic oligomers are produced by back-biting degradation from the initially formed linear polymers.
Courses of polymerization of N-carboxyanhydrides of γ-benzyl L- and DL-glutamate and of L- and DL-alanine with nickel dl-2-methylbutyrate-tri-n-butylphosphine catalyst system and with n-hexylamine were followed by the ir spectroscopic method. These kinetic results revealed clearly the significant difference in behavior observed between nickel catalyst and amine catalyst and suggest that the former has a stereoregulating power higher than the latter or nickel acetate catalyst system. This interpretation was supported by DL copolymerization of γ-benzyl glutamate N-carboxyanhydride and the main-chain configuration of its polymer.
The development of biodegradable cationic polymers for use in nonviral gene delivery was discussed. This high molecular weight multiblock copolymer (MBC) consists of repeating units of low molecular weight poly(ethylene glycol) conjugated to low molecular weight cationic(L-lysine). These MBC protected pDNA from endonuclease digestion for at least two hours. The pK of the conjugated imidazoles was found to be 4.75 which would facilitate buffering at low pH environments of the late endosome/lysosome.
Stable and monodispersive polyion complex micelles were prepared in an aqueous milieu through electrostatic interaction between a pair of oppositely-charged block copolymers with poly(ethylene glycol) segments: poly(ethylene glycol)-poly(L-lysine) block copolymer (PEG-P(Lys)) and poly(ethylene glycol)-poly(alpha,beta-aspartic acid) block copolymer (PEG-P(Asp)). It was confirmed from photon correlation spectroscopy (dynamic light scattering) that the scaled average characteristic line width (Gamma/K-2) was independent of the magnitude of the scattering vector (K-2), and the diffusion coefficient (D-T) kept constant regardless of the concentration, indicating that the polyion complex micelles were spherical particles without any secondary aggregates. Further, polydispersity indexes (mu(2)/Gamma(2)) were always less than 0.1 in the range of the measured concentration (1-10 mg/mL). The hydrodynamic radius at infinite dilution of polyion complex micelles was then determined to be 15.2 nm by using the Stokes-Einstein equation. The unimodal size distribution with d(w)/d(n) of 1.07 was confirmed from the correlation function profile by the histogram analysis. The size of polyion complex micelles was unchanged even after a 1-month storing, suggesting that the polyion complex micelles are in thermodynamic equilibrium. Viscosity measurement as well as laser-Doppler electrophoresis provided evidence of the stoichiometry of the polyion complex micelles formation. These polyion complex micelles have potential utility as vehicles for charged compounds, i.e., proteins and nucleic acids, in the field of drug delivery.
It is well-known that thermal treatment of aspartic acid produces poly(anhydroaspartic acid) (polysuccinimide), which can then be hydrolyzed to sodium polyaspartate. Both polysuccinimide and sodium polyaspartate have been analyzed by a variety of one- and two-dimensional NMR techniques. H-1 NMR indicates that sodium polyaspartate invariably contains; a 3:1 ratio of beta:alpha linkages, under a variety of synthesis and hydrolysis conditions. C-13 NMR and H-1/C-13 HMBC data show that,the sequencing is random. Residual succinimide levels in sodium polyaspartate are detectable down to about 1% by 1H NMR. COSY data of polysuccinimide and H-1/N-15 HMQC-TOCSY of polysuccinimide synthesized from 100% N-15 monomer indicate that phosphoric acid catalysis produces linear polymers, while uncatalyzed materials are branched.
Block copolymer micelles (polymer−metal complex micelles) containing platinum(II) complexes in the core were prepared through the complexation of cis-dichlorodiammineplatinum(II) (cisplatin, CDDP) with poly(ethylene glycol)−poly(α,β-aspartic acid) block copolymer (PEG−P(Asp)) in an aqueous medium. Dynamic light scattering measurements revealed that CDDP-complexed micelles had diameters of approximately 20 nm with considerably narrow distribution. The critical substitution molar ratio of CDDP to Asp residues in PEG−P(Asp) (CDDP/Asp) to form a stable micelle structure was determined to be 0.5. The CDDP-complexed micelle, which was stable in distilled water at room temperature, started to dissociate in approximately 10 h of the induction period accompanying the sustained release of platinum(II) complex from the micelle in physiological saline (0.15 M NaCl solution) at 37 °C. The release rate of CDDP was inversely correlated with the chain length of P(Asp) segments in the block copolymer. This property of sustained CDDP release with modulated micelle dissociation leads to the potential utility of the CDDP-complexed micelle as a tumor-directed carrier system.
In aqueous systems, polymeric micelles based on AB block copolymers of poly(ethylene oxide) (PEO) and poly(beta-benzyl L-aspartate) (PBLA) were investigated. First, AB block copolymers were synthesized using amino-terminated PEO to initiate the polymerization of beta-benzyl L-aspartate N-carboxy anhydride (BLA-NCA). The composition and molecular weights of the block copolymers were established using H-1 NMR. Micellar solutions of PEO-PBLA block copolymer were characterized by static and dynamic light scattering. Photophysical means were used to study the polymeric micelles. From changes in the fluorescence intensity and shifts in the excitation spectrum of pyrene upon micellization, critical micelle concentrations (cmc) of PEO-PBLA block copolymers were obtained. The vibrational structure of pyrene monomer fluorescence was altered in PEO-PBLA micellar solutions consistent with low polarity within the PBLA core. In PEO-PBLA micellar solutions, 1,3-(1,1'-dipyrenyl)propane intramolecular excimer emission, relative to monomer emission, was very weak; this indicates very low mobility of PBLA segments within the micellar core. Further evidence for the limited motion of the PBLA segments in the core was obtained by H-1 NMR. This limited motion of the PBLA segments in the micellar core is in contrast to low molecular weight surfactants which commonly show a higher degree of motion within their cores.
The melt polycondensation reaction of the prepolymer prepared from N-(benzyloxycarbonyl)-L-aspartic acid anhydride (N-CBz-L-aspartic acid anhydride) and low molecular weight poly(ethylene glycol) (PEG) using titanium isopropoxide (TIP) as a catalyst produced the new biodegradable poly(L-aspartic acid-co-PEG). This new copolymer had pendant amine functional groups along the polymer backbone chain. The optimal reaction conditions for the preparation of the prepolymer were obtained by using a 0.12 mol % of p-toluenesulfonic acid with PEG 200 for 48 h. The weight-average molecular weight of the prepolymer increased from 1,290 to 31,700 upon melt polycondensation for 6 h at 130°C under vacuum using 0.5 wt % TIP as a catalyst. The synthesized monomer, prepolymer, and copolymer were characterized by FTIR, 1H- and 13C-NMR, and UV spectrophotometers. Thermal properties of the prepolymer and the protected copolymer were measured by DSC. The glass transition temperature (Tg) of the prepolymer shifted to a significantly higher temperature with increasing molecular weight via melt polycondensation reaction, and no melting temperature was observed. The in vitro hydrolytic degradation of these poly(L-aspartic acid-co-PEG) was measured in terms of molecular weight loss at different times and pHs at 37°C. This pH-dependent molecular weight loss was due to a simple hydrolysis of the backbone ester linkages and was characterized by more rapid rates of hydrolysis at an alkaline pH. These new biodegradable poly(L-aspartic acid-co-PEG)s may have potential applications in the biomedical field. © 1998 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 36: 2949–2959, 1998
The n ‐hexylamine‐initiated polymerization of N ε ‐trifluoroacetyl‐ L ‐lysine N ‐carboxyanhydride in N , N ‐dimethyformamide was studied by nonaqueous capillary electrophoresis. A polypeptide with a broad molecular weight distribution was obtained and side reactions were clearly identified for polymerization at room temperature. The possibility of living polymerization at 0 °C was demonstrated.
Synthesis of living polypeptides by primary amine initiated polymerization of NCA at low temperatures.
image Synthesis of living polypeptides by primary amine initiated polymerization of NCA at low temperatures.
Studies have shown that the dose-limiting toxicity of amphotericin B (AmB), a key drug for systemic mycoses, depends on its self-aggregation state. In a step toward understanding the various factors in blood mediating the toxicity of AmB, we have investigated the effect of serum albumin, the most abundant plasma protein, on the aggregation state of AmB using absorption spectroscopy. The critical aggregation concentration (CAC) of AmB, which coincides with its concentration at the onset of toxicity (hemolysis), was 1.1 μM, but rose in proportion to the level of serum albumin (1.0 to 4.0% w/v). The CAC of AmB was 8.0 μM at 4.0% w/v serum albumin, which is considerably higher than peak therapeutic levels of AmB in plasma (i.e., 2.0 μM). Serum albumin (4.0% w/v) lowered the degree of aggregation of AmB (size of aggregates) above the CAC and increased its solubility. The results suggest that serum albumin attenuates the toxicity of AmB at a membrane level by affecting its aggregation state. In this way, serum albumin in blood may balance deleterious effects of AmB mediated by serum low-density lipoproteins. ©2000 Wiley-Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 89: 1589–1593, 2000
This article reviews recent developments in the polymerization of α-amino acid-N-carboxyanhydrides (NCAs) to form polypeptides. Traditional methods used to polymerize these monomers are described, and limitations in the utility of these systems for the preparation of polypeptides with controlled molecular weights and narrow molecular weight distributions are discussed. The development of transition-metal-based initiators, which activate the monomers to form covalent active species, permits the formation of polypeptides via the living polymerization of NCAs. In these systems, polymer molecular weights are controlled by monomer-to-initiator stoichiometry, polydispersities are low, and block copolypeptides can be prepared. The scope and limitations of these initiators and their key features and mode of operation are described in detail in this highlight.
Block copolymer micelles with aldehyde functionality were prepared in aqueous medium by dialyzing the N,N-dimethylacetamide solution of α-acetoxy-poly(ethylene glycol)-poly(d,l-lactide) block copolymer (acetal-PEG–PDLLA) against water, followed by mild acid treatment to convert the acetal moiety of the micelle to the aldehyde group. Peptidyl ligands (phenylalanine (Phe) and tyrosyl–glutamic acid (Tyr–Glu)) were then chemically conjugated to the micelle through Schiff base formation and successive reductive amination using NaBH3CN. Micelles with peptidyl ligands thus prepared have a size of approximately 40 nm with extremely narrow distribution (μ2/2<0.1) based on cumulant analysis of dynamic light scattering. A maximum 53% of the PEG-chain end of the micelle could be converted into peptidyl groups. Zeta potential values of Tyr–Glu derivatized micelles were well correlated with the amount of conjugated ligands, controllable over the range of 0 to−9 mV in sodium phosphate buffer (pH 7.4, 10 mM). These micelles with peptidyl ligands may have a utility for exploring the effect of the surface charge on the pharmacokinetic behavior of particulate systems as well as for modulated drug delivery where cellular peptidyl receptors play a substantial role.
To estimate the feasibility of novel containers for drugs, poly-(ethylene oxide)–poly(β-benzyl L-aspartate) (PEO–PBLA) micelles were prepared by dialysis against water using different solvents. The solvent selected is very important because it drastically affects the stability of polymeric micelles. The critical micelle concentration (cmc) of the prepared micelles in distilled water was determined by a fluorescence probe technique using pyrene. Indomethacin (IMC) as a model drug was incorporated into the micelles by dialysis and an oil/water emulsion method. Characteristics of PEO–PBLA micelles without and with the physically trapped IMC in the inner core of the micelles (IMC/PEO–PBLA) were studied by dynamic light scattering and gel permeation chromatography/HPLC as well as an in vitro release test of IMC from the micelles. For the PEO–PBLA block copolymers, N,N-dimethylacetamide (DMAc) was found to be the best of the solvents tested to form stable polymeric micelles with a narrow size distribution and avoid its aggregation, and the cmc of PEO–PBLA micelles thus prepared was determined to be ca. 18 mg/L in water. The diameters of PEO–PBLA micelles and IMC/PEO–PBLA micelles in number averaged scale were observed to be ca. 19 and 25–29 nm, respectively. The release study of IMC from IMC/PEO–PBLA micelles in various buffer solutions at the pH range from 1.2 to 7.4 at 37 °C revealed that the release rate of IMC from the micelles was increased by increasing the pH of the medium and indicated that the release rate of IMC from the micelles are controlled by the partition coefficient of IMC based on the pH of the medium and interaction between IMC and the hydrophobic portion of the micelles.
This paper reviews ring-opening polymerization of lactones and lactides with different types of initiators and catalysts as
well as their use in the synthesis of macromolecules with advanced architecture. The purpose of this paper is to review the
latest developments within the coordination-insertion mechanism, and to describe the mechanisms and typical kinetic features.
Cationic and anionic ring-opening polymerizations are mentioned only briefly.
The entrapment of Adriamycin (ADR) in micelles composed of AB block copolymers (poly(ethylene oxide-co--benzyl L-aspartate) (PEO-PBLA)) was investigated. The loading process involved transfer of ADR and PEO-PBLA into an aqueous milieu from dimethyl-formamide (DMF) through a dialysis procedure. Evidence for the physical entrapment of ADR in the polymeric micelles was derived from fluorescence spectroscopy and gel permeation chromatography (GPC). The total fluorescence intensity of ADR was low, suggesting that the drug was self-associated in the micelles. In addition, quenching experiments, using a water-soluble quencher (iodide (I–)), showed that the fluorescence of ADR present in micellar solutions was largely unaffected by I–, whereas the fluorescence of free ADR was readily quenched. From Stern-Volmer plots, quenching constants (KSV) of 2.2 and 17 M–l were determined for ADR in micellar solutions and free ADR, respectively. As a result of the entrapment of ADR in the micelles, ADR binds only slightly serum albumin as evidenced by GPC. In contrast, ADR readily binds serum albumin in aqueous solutions. The findings suggest that ADR is stably entrapped in PEO-PBLA micelles. ADR entrapment in polymeric micelles is expected to affect markedly the pharmacokinetics of ADR.
Purpose. The pharmacological activity and pharmacokinetics of cisplatin (CDDP)-loaded polymeric micelles were examined to reveal their usefulness as a novel tumor-directed drug carrier system of CDDP.
Methods. In biodistribution assay, free CDDP or CDDP-loaded micelles were administered intravenously to Lewis lung carcinoma-bearing mice. Antitumor activity and nephrotoxicity were respectively evaluated by the measurement of tumor size and plasma blood urea nitrogen (BUN) after single bolus i.v. administration of each drug.
Results. The time profile of the plasma Pt level after the injection of the micelles exhibited a time-modulated disappearance as observed in saline in vitro. The micelles exhibited 5.2- and 4.6-fold higher AUC of Pt in the plasma and tumor, respectively, with minimal change in the kidney, in comparison with free CDDP, suggesting that prolonged circulation of Pt in circulation and specific accumulation in the tumor were achieved utilizing the micellar drug carrier system. Administration of the micelles at the dose exhibiting antitumor activity similar to free CDDP did not increase the plasma BUN, whereas free CDDP induced its remarkable increase.
Conclusion. CDDP-loaded micelles restrained nephrotoxicity, which is the dose-limiting factor of CDDP, while exhibiting tumor-specific accumulation. Thus, CDDP-loaded micelles are expected to be a novel formulation of CDDP for clinical use.
Polymeric micelles of varying size in the range of 20 to 100 nm entrapping an antitumor drug, cis-dichlorodiammineplatinum(II) (cisplatin, CDDP), were prepared through the polymer–metal complex formation of CDDP with a mixture of poly(ethylene glycol)–poly(α,β-aspartic acid) block copolymer (PEG–P(Asp)) and poly(α,β-aspartic acid) homopolymer (P(Asp)) with the different feed ratio in distilled water. An increased ratio of P(Asp) to PEG–P(Asp) led to an increase in the micellar size in a controllable manner as well as prolongation in the induction period of the micellar decay accompanied by a sustained release of CDDP in physiological saline at 37°C. All of the CDDP-loaded micelles with a different incorporation ratio of P(Asp) exhibited appreciable in vitro cytotoxicity due to CDDP release from the micelles by prolonged incubation. These CDDP-loaded micelles are expected to have potential utility in tumor-directed delivery system of CDDP through the modulated in vivo biodisposition based on the EPR effect.
The limited solubility of hydrophobic drugs may hamper their potential investigation. Vehicles that can incorporate and effectively deliver such drugs are of interest. To this end, we have prepared micelles based on AB block copolymers of poly(ethylene oxide) (PEO) and poly(β-benzyl l-aspartate) (PBLA). The distribution of hydrophobic molecules into PEO-PBLA micelles was investigated by UV spectroscopy and the fluorescence probe technique using pyrene as a model drug. Further, the effects of temperature and sonication on pyrene partitioning into PEO-PBLA micelles were studied. The solubility of pyrene was enhanced in a linear fashion with respect to PEO-PBLA concentration. Weight-average partition coefficients of about 104 were determined for pyrene distribution in PEO-PBLA micellar solutions. The photophysical properties of pyrene, which are modified upon uptake within PEO-PBLA micelles, indicate a low polarity and pyrene mobility in the solubilization site (i.e. core region). The polarity of micellar cores (i.e. micropolarity) is apparently important for solute partitioning into polymeric micelles. PEO-PBLA micelles offer micropolarities slightly higher than those of the well-studied AB block copolymer micelles based on PEO and polystyrene (PS) on the pyrene scale. Nonetheless, block copolymer micelles, having cores consisting of a poly(amino acid), can be chemically tailored to adjust micropolarities for enhanced drug loading.
Star-block copolymer based on PBLG as the hydrophobic part and PEO as the hydrophilic one (as abbreviated GEG) was synthesized and characterized. Polymeric micelle was prepared by the diafiltration method. From the measurement of photon correlation spectroscopy, the nanoparticle sizes of GEG-1, GEG-2 and GEG-3 were 106.5±59.2, 43.8±0.7 and 13.5±1.0 nm in number average, respectively, indicating of the formation of polymeric micelle. Also, the nanoparticle sizes were dependent on the PBLG chain length, i.e. the more PBLG content in the copolymer, the larger the particle size. From the observation of transmission electron microscope(TEM), GEG-2 block copolymer had almost spherical shapes with size range about 20–70 nm, that was similar to particle size measurement. Fluorescence spectroscopy measurement indicated that GEG block copolymers associated in water to form polymeric micelles and critical micelle concentration (CMC) values of the block copolymers decreased with increasing PBLG chain length in the block copolymer. Characteristic peaks of the protons of the benzyl group in the PBLG and the methylene protons adjacent to the benzyl group of the PBLG segment in the GEG-2 nanoparticles appeared in 7.2∼7.4 and 5.0∼5.2 ppm, respectively, and disappeared in D2O, indicating the restricted motions of these protons within the micellar core and the very rigid structure of the PBLG core in the GEG polymeric micelles. Release of ADR from the polymeric micelles in vitro was slower in longer PBLG chain length and higher loading contents of ADR.
A water-insoluble anticancer drug, KRN 5500 (KRN), was incorporated into polymeric micelles forming from poly(ethylene glycol–poly(amino acid) block copolymers by physical entrapment utilizing hydrophobic interactions between this drug and the poly(amino acid) chain block of the block copolymers. Three block copolymers were examined for this incorporation; poly(ethylene glycol)–poly(β-benzyl l-aspartate) (PEG–PBLA) and its two derivatives obtained by partial hydrolysis at the β-benzyl l-aspartate (BLA) units (PEG–P(Asp, BLA)) and by partial cetyl ester substitution at the BLA units (PEG–P(C16, BLA)), respectively. Among these block copolymers, considerable effects of the cetyl esterification were seen on KRN yield and particle size. Considerable differences in the KRN incorporation yield and particle size were also observed between DMF and DMS used as solvent to dissolve KRN and the block copolymers. Sonication was turned out to be an effective method to obtain a polymer micelles fraction in high efficiency, and sonication was considered to work for separating intermicellar associates into dispersed micelles. A KRN incorporation procedure by dialysis using PEG–P(C16, BLA) and DMSO (as solvent) followed by sonication brought about polymeric micelles of 71 nm in weight–average diameter. This shows successful incorporation of a water-insoluble drug into polymeric micelles by optimizing block copolymer structure and incorporation conditions.
Five new polyethylenimines (PEI) were synthesized by polymerization of aziridine in aqueous solution and compared to several commercially available PEI used for gene transfer. Polymers were characterized by 13C NMR spectroscopy, capillary viscosimetry, potentiometric titration and Cu(II) complex formation to gain insight into structural and functional properties. 13C NMR analysis revealed differences in the extent of branching based on the ratio of primary, secondary and tertiary amino groups. An amino group ratio 1°:2°:3°=1:2:1 was obtained for the synthesized PEI, whereas commercially available PEI generally showed a higher degree of branching (1:1:1). Capillary viscosimetry of aqueous PEI solutions with a sufficient amount of salt gave Mark–Houwink parameters of α=0.26 and KV=1.00 cm3/g for the commercially available polymers. In case of the synthesized polymers, variation of reaction conditions yielded viscosity average molar masses (Mv) in the range of 8000–24 000 g/mol. PEI solutions were investigated by potentiometric titration analysis showing that their buffer capacity was not significantly influenced by molar mass or polymer structure. The pKa values (8.18–9.94) and the buffer capacity β (0.08–0.014 mol/l) were of comparable magnitude. This study highlights the necessity of more detailed characterization methods for PEI used in gene transfer protocols since physico-chemical properties do not reflect the vast differences found in transfection efficiencies.
The conjugation reaction of adriamycin (ADR) to poly (ethylene oxide)-poly (aspartic acid) block copolymer (PEO-P(Asp)) was optimized in order to inhibit side reactions, which damage the ADR residues. By diminishing alkaline components and lowering temperature, ADR was successfully conjugated to PEO-P(Asp) without side reactions. The obtained ADR-conjugated poly(ethylene oxide)-poly(aspartic acid) block copolymer [PEO-P(Asp(ADR))] was observed to form a micellar structure with a narrower distribution in size than the PEO-P(Asp(ADR)) synthesized by the previously reported method. Unconjugated ADR remained in the inner core of the PEO-P(Asp(ADR)) micelles was removed by dialysis in organic solvents to obtain the micelle without free ADR. Regulated amount of free ADR was then successfully entrapped into the inner core of these purified micelles. In vitro cytotoxicity against P388D1 cells of PEO-P(Asp(ADR)) micelle without free ADR was assayed to estimate the contribution of physically entrapped ADR on anti-tumor activity of the micelle-forming polymeric drug, PEO-P(Asp(ADR)).
The goal of this study was to assess a solvent evaporation method for the encapsulation of amphotericin B (AmB) in poly(ethylene oxide)-block-poly(N-hexyl stearate l-aspartamide) (PEO-b-PHSA) micelles. By the solvent evaporation method, PEO-b-PHSA self-assembled into small spherical micelles with a high AmB content based on transmission electron microscopy, size exclusion chromatography and absorption spectroscopy. The encapsulation of AmB was slightly better than an earlier method based on dialysis. Importantly, AmB in PEO-b-PHSA micelles encapsulated by the solvent evaporation method was non-haemolytic at 15 μg/ml, whereas AmB in PEO-b-PHSA micelles encapsulated by the dialysis method caused 50% haemolysis at the level of 3.8 μg/ml, and AmB itself caused 100% haemolysis at 1.0 μg/ml. Thus, PEO-b-PHSA micelles could effectively encapsulate AmB, increase the overall water solubility of AmB and reduce the toxicity of the membrane-acting drug, particularly by a solvent evaporation method.
This work describes a simple, versatile solid-phase peptide-synthesis (SPPS) method for preparing micelle-forming poly(ethylene oxide)-block-peptide block copolymers for drug delivery. To demonstrate its utility, this SPPS method was used to construct two series of micelle-forming block copolymers (one of constant core-composition and variable length; the other of constant core length and variable composition). The block copolymers were then used to study in detail the effect of size and composition on micellization. The various block copolymers were prepared by a combination of SPPS for the peptide block, followed by solution–phase conjugation of the peptide block with a proprionic acid derivative of poly(ethylene oxide) (PEO) to form the PEO-b-peptide block copolymer. The composition of each block component was characterized by mass spectrometry (MALDI and ES-MS). Block copolymer compositions were characterized by 1H NMR. All the block copolymers were found to form micelles as judged by transmission electron microscopy (TEM) and light scattering analysis. To demonstrate their potential as drug delivery systems, micelles prepared from one member of the PEO-b-peptide block copolymer series were physically loaded with the anticancer drug doxorubicin (DOX). Micelle static and dynamic stability were found to correlate strongly with micelle core length. In contrast, these same micellization properties appear to be a complex function of core composition, and no clear trends could be identified from among the set of compositionally varying, fixed length block copolymer micelles. We conclude that SPPS can be used to construct biocompatible block copolymers with well-defined core lengths and compositions, which in turn can be used to study and to tailor the behavior of block copolymer micelles.
Doxorubicin (DOX) was physically loaded into micelles prepared from poly(ethylene glycol)–poly(β-benzyl-l-aspartate) block copolymer (PEG–PBLA) by an o/w emulsion method with a substantial drug loading level (15 to 20 w/w%). DOX-loaded micelles were narrowly distributed in size with diameters of approximately 50–70 nm. Dimer derivatives of DOX as well as DOX itself were revealed to be entrapped in the micelle, the former seems to improve micelle stability due to its low water solubility and possible interaction with benzyl residues of PBLA segments through π–π stacking. Release of DOX compounds from the micelles proceeded in two stages: an initial rapid release was followed by a stage of slow and long-lasting release of DOX. Acceleration of DOX release can be obtained by lowering the surrounding pH from 7.4 to 5.0, suggesting a pH-sensitive release of DOX from the micelles. A remarkable improvement in blood circulation of DOX was achieved by use of PEG–PBLA micelle as a carrier presumably due to the reduced reticuloendothelial system uptake of the micelles through a steric stabilization mechanism. Finally, DOX loaded in the micelle showed a considerably higher antitumor activity compared to free DOX against mouse C26 tumor by i.v. injection, indicating a promising feature for PEG–PBLA micelle as a long-circulating carrier system useful in modulated drug delivery.