Nonnative aggregation of an IgG1 antibody in acidic conditions, part 2: nucleation and growth kinetics with competing growth mechanisms

Aggregation mechanisms as a function of pH were assessed for the IgG1 antibody described in Part 1 (Brummitt RK, Nesta DP, Chang L, Chase SF, Laue TM, Roberts CJ. Non-native aggregation of an IGG1 antibody in acidic conditions: 1. Unfolding, colloidal interactions, and high molecular weight aggregate formation. J Pharm Sci. In press). Aggregation kinetics along with static light scattering and size-exclusion chromatography indicated that the aggregate nucleus was a dimer for all conditions tested, and this was semiquantitatively consistent with scaling of the characteristic time scale for nucleation (τ(n)) versus protein concentration at pH 4.5 and pH 5.5. Changing pH significantly altered the mechanism of aggregate growth, as well as the size and solubility of aggregates that were formed. Aggregates at pH 3.5 grew primarily by monomer addition and remained small and soluble. Aggregates at pH 4.5 grew first by chain polymerization, followed by condensation polymerization, leading ultimately to large insoluble particles. At pH 5.5, monomer loss resulted primarily in insoluble aggregate formation, with only low levels of soluble aggregate intermediates detected at early times. The influence of pH on aggregate solubility and the reversibility of aggregate phase separation were confirmed via cloud point titrations. Qualitatively, the global aggregation behavior was consistent with reduction of charge-charge repulsions as a primary factor in promoting larger aggregates and aggregate phase separation.     

Entrapment of bioactive molecules in poly (alkylcyanoacrylate) nanoparticles

Poly (alkylcyanoacrylate) [PACA] nanoparticles have been studied since the early 1980s as possible colloidal drug delivery systems. Several excellent general reviews have since been published on this subject. This review focuses on the use of the two different methods (encapsulation and sorption) for the entrapment of drugs and model compounds in PACA nanoparticles. The term encapsulation is used when the drug or model compound is added at the same time or before the monomer to the polymerization template. The term sorption is used when the compound is added after the polymerization has taken place. High drug entrapment can be achieved with both methods and the method of entrapment (encapsulation or sorption) should be chosen depending on the type of drug to be entrapped and the method of particle preparation (interfacial polymerization of a coarse emulsion or a microemulsion or micellar polymerization). The type, chain length, and amount of monomer used for the polymerization as well as possible interactions of the compound with the monomer during polymerization should also be considered in the choice of entrapment method and these can also influence the extent of encapsulation.     

Evidence for restrictive parameters in formulation of insulin-loaded nanocapsules.

Poly(isobutylcyanoacrylate) nanocapsules with an oily core were originally proposed for lipophilic drug encapsulation [Int. J. Pharm. 28 (1986) 125] but insulin, a hydrosoluble protein, has also been successfully encapsulated by Damgé et al. [Diabetes 37 (1988) 246]. The aim of this work was to understand if several parameters were restrictive for the encapsulation of insulin into the oily core of the nanocapsules prepared by interfacial polymerization. The encapsulation efficiency of insulin was not affected by the type of insulin since the peptides adopted the same association state after their addition to the organic phase. Formulation parameters mainly affected the size of the nanocapsules obtained but did not influence the insulin encapsulation efficiency. In contrast, the order of introduction of insulin and of the monomer in the organic phase was shown to control the formation and the characteristics of the nanocapsules. The key parameters, which were found to clearly influence the encapsulation efficiency of insulin, were the pH of the aqueous insulin solution and the origin of the monomer. Both of these parameters can affect the rate of the interfacial polymerization. Consequently, the ability of insulin to be entrapped into the oil containing nanocapsules appeared to be governed more by the rate of the monomer polymerization.     

The effect of monomers on the formulation of polymeric nanocapsules based on polyureas and polyamides

Formulation of nanocapsules based on polyureas and polyamides have been tested using a patented process. This method based on polycondensation reaction of two complementary monomers and spontaneous formation of oil in water emulsion, is an alternative concept to the known technique based on the same type of reaction used for the formulation of microcapsules, and in which the lipophilic monomer was emulsified in the organic phase before the formation of the polymeric membrane. Nanocapsules can be prepared from different monomers. Wall based on cross-linked polymer contributes to the stability of nanocapsules during and after formulation. The permeability of the polymeric wall is related to its crystallinity and contributes to the growth of nanocapsule membrane by the diffusion of the hydrophilic monomers to get stable colloidal suspensions.     

Atomic force microscopy imaging of novel type of polymeric colloidal nanostructures.

Polymeric nanoparticles were prepared by the interfacial poly-condensation of the lipophilic monomer, phtaloyldichloride and the hydrophilic monomer, diethylenetriamine, in the presence and absence of the surfactant Pluronic F68. The colloidal systems were analysed by dynamic light scattering and atomic force microscopy, the structures formed have two populations (150 and 350 nm) in the presence of the surfactant and one population (450 nm) in the absence of the surfactant. The results can be interpreted in terms of the formation of hollow nanocapsules that collapse on deposition and drying.     

Methods of obtaining and formation mechanisms of polymer nanoparticles

Severals processes exist today for manufacturing colloidal systems of nanospheres or nanocapsules. These nanoparticles have applications in various industrial fields such as cosmetic, pharmacy, food industry, agrochemicals.... The formation of nanoparticles allows the protection of an active molecule by a polymeric coating and the release of this product following a perfectly defined profile, which depends on the nature of the polymer, the type of particle and the field of use. The purpose of this article is to expose the different methods of preparation of nanoparticular systems. These methods are classified in two mains categories. The first gathers most of the methods which are based on polymerization reactions, the second presents the use of preformed polymers (of natural or synthetic origin). A thorough study is devoted to the mechanisms of formation, in order to find the advantages and the disadvantages of each technique to support the development of manufactoring processes.     

Nanocapsule technology: a review

Nanocapsules are submicroscopic colloidal drug carrier systems composed of an oily or an aqueous core surrounded by a thin polymer membrane. Two technologies can be used to obtain such nanocapsules: the interfacial polymerization of a monomer or the interfacial nanodeposition of a preformed polymer. This article is an extended review of these nanocapsule technologies and their applications for the treatment of various diseases (including cancer and infections).     

Microemulsions as colloidal vehicle systems for dermal drug delivery. Part IV: Investigation of microemulsion systems based on a eutectic mixture of lidocaine and prilocaine as the colloidal phase by dynamic light scattering

Stable oil-in-water (o/w) microemulsions used as vehicles for dermal drug delivery have been developed using lidocaine (lignocaine) and prilocaine in oil form (eutectic mixture), a blend of a high (Tween 80, hydrophilic-lipophilic balance (HLB) = 15.0) and a low (Poloxamer 331, HLB = 1.0) HLB surfactant and propylene glycol-water as hydrophilic phase. These microemulsions were able to solubilize up to 20% eutectic mixture of lidocaine and prilocaine without phase separation. The dispersity of the oil phase was investigated by dynamic light scattering. Small colloidal droplets for stable microemulsions of 5~10 nm were observed. At constant surfactant and hydrophilic phase concentration, increasing the total drug concentration in the microemulsion resulted in an increase in the droplet size of the dispersed, colloidal phase. It was observed that a monolayer of surfactant surrounds the oil (eutectic mixture) core. Colloidal droplets of the microemulsion interact via hard sphere with supplementary attractive interaction. This observed interparticle attractive interaction could explain the observed phase behaviour with respect to change in the basicity of the hydrophilic phase as well as the increase in volume fraction of the dispersed, colloidal phase. It was also observed that the stability and size of this dispersed phase depends on the pH of the composition. Because these microemulsions formed stable, isotropic systems in the range of pH 9.5 to 10.4 with alkali buffer or NaOH solution instead of water as hydrophilic phase, so one can produce microemulsions in this pH area.     

Prediction of a Stable Microemulsion Formulation for the Oral Delivery of a Combination of Antitubercular Drugs Using ANN Methodology

PURPOSE: The aim of this project was to develop a colloidal dosage form for the oral delivery of rifampicin and isoniazid in combination with the aid of artificial neural network (ANN) data modeling. METHODS: Data from the 20 pseudoternary phase triangles containing miglyol 812 as the oil component and a mixture of surfactants or a surfactant/cosurfactant blend were used to train, test, and validate the ANN model. The weight ratios of individual components were correlated with the observed phase behavior using radial basis function (RBF) network architecture. The criterion for judging the best model was the percentage success of the model prediction. RESULTS: The best model successfully predicted the microemulsion region as well as the coarse emulsion region but failed to predict the multiphase liquid crystalline phase for cosurfactant-free systems indicating the difference in microemulsion behavior on dilution with water. CONCLUSIONS: A novel microemulsion formulation capable of delivering rifampicin and isoniazid in combination was created to allow for their differences in solubility and potential for chemical reaction. The developed model allowed better understanding of the process of microemulsion formation and stability within pseudoternary colloidal systems.     

Pseudo-ternary phase diagrams of aqueous mixtures of Quil A, cholesterol and phospholipid prepared by the lipid-film hydration method

Pseudo-ternary phase diagrams of the polar lipids Quil A, cholesterol (Chol) and phosphatidylcholine (PC) in aqueous mixtures prepared by the lipid film hydration method (where dried lipid film of phospholipids and cholesterol are hydrated by an aqueous solution of Quil A) were investigated in terms of the types of particulate structures formed therein. Negative staining transmission electron microscopy and polarized light microscopy were used to characterize the colloidal and coarse dispersed particles present in the systems. Pseudo-ternary phase diagrams were established for lipid mixtures hydrated in water and in Tris buffer (pH 7.4). The effect of equilibration time was also studied with respect to systems hydrated in water where the samples were stored for 2 months at 4 °C. Depending on the mass ratio of Quil A, Chol and PC in the systems, various colloidal particles including ISCOM matrices, liposomes, ring-like micelles and worm-like micelles were observed. Other colloidal particles were also observed as minor structures in the presence of these predominant colloids including helices, layered structures and lamellae (hexagonal pattern of ring-like micelles). In terms of the conditions which appeared to promote the formation of ISCOM matrices, the area of the phase diagrams associated with systems containing these structures increased in the order: hydrated in water/short equilibration period相似文献   

Physico-chemical characterization of insulin-loaded poly(isobutylcyanoacrylate) nanocapsules obtained by interfacial polymerization.

Insulin could be encapsulated very efficiently in oily containing poly(isobutylcyanoacrylate) nanocapsules obtained by interfacial polymerization. In addition, these nanocapsules showed unexpected biological activity after intragastric administration. The hypoglycemic effect was characterized by a lag time period of 2 days and a prolonged effect over a period of 20 days. To explain, the high encapsulation rate of insulin achieved in these nanocapsules and the biological effect, this work was focused on the characterization of the nanocapsules and on the study of the mechanism of nanocapsule formation. Results showed that insulin was found unmodified during the nanoencapsulation process. This was due to the large amount of ethanol used in the preparation of the nanocapsules that initiated the polymerization of isobutylcyanoacrylate preserving the peptide from a reaction with the monomer. Results also showed that insulin was located inside the core of the nanocapsules and not simply adsorbed onto their surface.     

Preparation of uniform titanium dioxide (TiO2) polystyrene-based composite particles using the glass membrane emulsification process with a subsequent suspension polymerization

Uniform titanium dioxide (TiO(2))-polystyrene-based composite particles were prepared using the glass membrane emulsification process followed by a subsequent suspension polymerization. The oil phase, consisting of anatase TiO(2) fine powder, monomers, methyl laurate as the hydrophobic additive, Disperbyk-180 and the poly(styrene-co-2-ethyl hexylacrylate) were emulsified through the membrane pores into the aqueous phase containing stabilizers to form a (solid-in-oil)-in-water (S/O/W) emulsion of monomer droplets. The suspension polymerization was carried out at 343 K for 24 h under a nitrogen atmosphere. An SPG membrane with a pore size of 5.25 microm was employed and 20-25 microm TiO(2)-polystyrene based composite particles were obtained depending on the composition of polymerizing oil phase. The effects of the co-monomer, 2-ethylhexyl acrylate and the cross-linking agent, divinyl benzene on the dispersion stability of TiO(2) in the oil phase, the surface feature of the particle and the encapsulation loading were investigated in this study. The membrane emulsification process was capable of preparing the composite particles with approximately 5 wt% of TiO(2) encapsulated, which accounts for with at least 85 wt% of TiO(2) in the oil phase.     

Preparation and characterization of two-phase melt systems of lidocaine

The melting point of lidocaine was significantly lowered when mixed with thymol and/or aqueous ethanol. Mixtures of lidocaine and thymol at ratios within the range of 30:70-70:30 (w:w) became homogeneous oils at 25 degrees C. In a pH 9.2 carbonate buffer containing 25% ethanol, lidocaine (5%, w:w) also liquefied at 25 degrees C. The studies led to the development of novel two-phase melt systems of lidocaine (TMS) which consisted of a highly concentrated oil phase of lidocaine and an alcoholic aqueous phase. A compositional phase diagram showed that in aqueous dispersions of lidocaine, concurrent use of thymol and ethanol depressed the melting point of lidocaine more effectively than when they were used individually. Both thymol and aqueous ethanol were necessary as melting point depressing agents to achieve the highest possible lidocaine concentration of 87% (w:w) in the oil phase of a TMS at 25 degrees C. Containing an internal oil phase and an external aqueous phase at ambient temperature, such a TMS can be readily formulated into topical O/W cream after addition of proper surfactants and thickening agents. In an anesthetic activity test using mouse tail-flick model, a 5% lidocaine cream prepared was highly effective as shown by the prolonged latency time of the mice to a heat stimulus as compared with a placebo (P<0.05).     

Association of cyclosporin to isohexylcyanoacrylate nanospheres and subsequent release in human plasma in vitro.

Polyisohexylcyanoacrylate nanocapsules containing cyclosporin were prepared by mixing in a 1:2 ratio an oil/ethanol solution of monomer and drug with an aqueous phase. Drug nanoencapsulation rate was controlled by its partition coefficient between the inner (organic) and outer (aqueous) phases. Thus highest encapsulation yields (88 per cent) were achieved by reducing cyclosporin solubility in the aqueous phase, i.e. by reducing ethanol concentration under reduced pressure, achieving a 3-fold volume reduction. Due to the relative insolubility of cyclosporin in water, no drug was released from the nanocapsules during storage in this injectable vehicle. Upon a 1/5 dilution in human plasma at 37 degrees C in vitro around 40 per cent of the initially encapsulated cyclosporin diffused quickly out of the capsules and an equilibrium was reached, the drug being most likely dissolved in the fatty compartment of the plasma such as lipoproteins, etc. This release mechanism is different from plain polymeric nanoparticles. Indeed, in this case the drug was released in two phases: an initial burst (around 60 per cent) of adsorbed drug as a result of the dilution, followed by a slow release (around 20 per cent over 3 h) which is likely to result from the progressive enzymatic erosion of the polymer. The initial burst was markedly more pronounced (around 80 per cent) when nanoparticle suspensions were evaporated to 1/3 of their initial volume under reduced pressure. Finally, experiments performed at 0 degree C allowed a reduction of the fraction released immediately from both types of nanospheres, probably because of a reduced solubility in plasma. In the case of nanoparticles the second phase of slow release is also inhibited at 0 degree C, in agreement with an enzymatically controlled release mechanism.     

Vidarabine-Loaded Nanoparticles: A Physicochemical Study

The chemical reaction of vidarabine (VIDA) with isohexyl cyanoacrylate nanoparticles in a pH-dependent fashion occurs only in the presence of dioctylsulfosuccinate (DOSS). The formation of an ion pair with DOSS allows a better contact of VIDA with the monomer during the polymerization process taking place in micelles. On the basis of molecular weight profiles of the polymer, determined by gel permeation chromatography (GPC), it is proposed that VIDA induces the polymerization of cyanoacrylic monomers through a zwitterionic pathway. This mechanism allows the covalent linkage of the drug with the polymer, which is consistent with NMR experiments. The present study illustrates the need for physicochemical studies in the design of new colloidal drug delivery formulations.     

Preparation of poly (alkylcyanoacrylate) nanoparticles by polymerization of water-free microemulsions

Phase diagrams of the pseudoternary systems ethyloleate, polyoxyethylene 20 sorbitan mono-oleate/sorbitan monolaurate and propylene glycol with and without butanol as a co-surfactant were prepared. Areas containing optically isotropic, one-phase systems were identified and samples therein designated as droplet, bicontinuous or solution type microemulsions using conductivity, viscosity and self-diffusion NMR. Nanoparticles were prepared by polymerization of selected microemulsions with ethyl-2-cyanoacrylate and the morphology of the particles was investigated. Addition of monomer to all types of microemulsions led to the formation of nanoparticles, which had an average size of 244 +/- 25 nm, an average polydispersity index of 0.15 +/- 0.04 and a zeta-potential of -17 +/- 3 mV. The formation of particles from water-free microemulsions of different types is surprising, particularly considering that polymerization is expected to occur at a water-oil interface by base-catalysed polymerization. It would appear that propylene glycol is sufficiently nucleophilic to initiate the polymerization. The use of water-free microemulsions as templates for the preparation of poly (alkylcyanoacrylate) nanoparticles opens up interesting opportunities for the encapsulation of bioactives which do not have suitable properties for encapsulation on the basis of water-containing microemulsions.     

Immuno-stimulating complexes prepared by ethanol injection

This study describes the formulation of immuno-stimulating complexes (ISCOMs) utilising the ethanol injection technique. Cholesterol and phosphatidylcholine were dissolved in ethanol and the resulting solution was rapidly injected into a stirred, aqueous solution of the triterpene-saponin mixture Quil-A. The reversed experiment was also carried out by adding the aqueous Quil-A solution to a solution of cholesterol/phosphatidylcholine dissolved in ethanol. This was done by either rapid injection or dropwise addition of the aqueous Quil-A solution. The colloidal dispersions obtained by ethanol injection and reversed addition were compared with formulations obtained by the dialysis and lipid-film hydration methods. In a further experiment, the preparation of ISCOMs from liposomes as precursor structures was investigated. Transmission electron microscopy was used to analyse the resulting colloidal dispersions. Samples were also compared with respect to homogeneity of obtained particle species. The ethanol injection technique led to formation of ISCOMs in high numbers within 2 h post formulation. The reversed rapid injection resulted in a similar colloidal dispersion, differing from the former mainly due to the presence of some helical micellar structures. The reversed, dropwise addition led to the formation of helices as the predominant colloidal structure. Of the three previously established methods, only dialysis led to the formation of ISCOMs within 48 h. The lipid-film hydration method and the approach using liposomes as precursor structures did not produce ISCOMs under the conditions and within the time periods investigated. However, it is known that dispersions prepared by the hydration method equilibrate towards ISCOMs after longer storage. Ethanol injection and reversed rapid injection are simple, cost-effective and quick methods to produce ISCOMs.     

Organic solvent-free polymeric microspheres prepared from aqueous colloidal polymer dispersions by a w/o-emulsion technique

Polymeric microspheres were prepared from water-insoluble polymers by a novel technique without the use of organic solvents. Aqueous colloidal polymer dispersions (latexes or pseudolatexes) were emulsified into a heated external oil phase to form a w/o emulsion. The colloidal polymer particles fused (coalesced) into homogeneous polymeric microspheres at temperatures above the minimum film formation temperature upon removal of water. The formation of the microspheres was affected by the glass transition temperature of the polymer, the type of oil and surfactant, the heating temperature and time, and the addition of plasticizers. Plasticizers had to be added to colloidal dispersions with high minimum film formation temperatures. The resulting microspheres were spherical with a smooth surface and non-agglomerated. The particle size could be varied between 5 and 250 μm. Water-soluble compounds such as propranolol HC1 could be entrapped with drug loadings up to 40% within the microspheres by dissolving the drug in the aqueous polymer dispersion prior to the emulsification step. The drug release was sustained over a 6-h period with microspheres prepared with the acrylic pseudolatex, Eudragit RS 30D.     

Microemulsions as colloidal vehicle systems for dermal drug delivery. Part V: Microemulsions without and with glycolipid as penetration enhancer

The aim of this study was to investigate the dermal administration of a highly hydrophilic model drug, diphenhydramine (DPH), in colloidal systems with an aqueous colloidal phase in the presence of a glycolipid (GL) as a penetration modifier. Dermal penetration of DPH, GL, and isopropylpalmitate (IPP) from ME systems without GL and with GL as well as from a hydrogel used as standard formulation were estimated in vitro using human skin. The penetration of the drug, the oil (IPP), and the GL was measured with highly sensitive HPLC, HPLC-MS, and GC-MS assays, respectively. It could be shown that penetration modifier GL is penetrating very fast, and to a high extent into and through the human skin. In contrast, the penetration of IPP used as oily phase in the ME is limited. When incorporated in the ME systems GL and DPH was accumulated in the viable epidermis and in the dermis. Using ME containing a penetration modifier such as GL, a slight additional enhancing effect could be observed, particularly concerning the penetration of DPH into the acceptor fluid when a highly hydrophilic drug such as DPH was applied.     

Characterisation of drug release from cubosomes using the pressure ultrafiltration method

Cubosomes have been proposed as a controlled release, intravenous drug delivery system. The objective of this study was to characterise cubosomes as either a therapeutically useful, controlled release delivery system, or as a burst release carrier such as submicron emulsions. The pressure ultrafiltration method and equilibrium dialysis were used to elucidate the in vitro drug release mechanisms. On dilution of cubosomes, lipophilic compounds were released rapidly when studied by the pressure ultrafiltration method. This agrees with the behaviour predicted from simple diffusion from the bulk non-dispersed cubic phase. In contrast, equilibrium dialysis incorrectly indicated sustained drug release from cubosomes. This study illustrates that cubosomes should be classified as a burst release delivery system where drug is released by diffusion from the cubic phase matrix, and that pressure ultrafiltration may have benefits over dialysis methods for measurement of drug release from colloidal particle-based drug delivery systems.