Drug-loaded nanocarriers: passive targeting and crossing of biological barriers.

Poor bioavailability and poor pharmacokinetic characteristics are some of the leading causes of drug development failure. Therefore, poorly-soluble drugs, fragile proteins or nucleic acid products may benefit from their encapsulation in nanosized vehicles, providing enhanced solubilization, protection against degradation, and increased access to pathological compartments. A key element for the success of drug-loaded nanocarriers is their ability to either cross biological barriers themselves, or allow loaded drugs to traverse them to achieve optimal pharmacological action at pathological sites. Depending on the mode of administration, nanocarriers may have to cross different physiological barriers in their journey towards their target. In this review, the crossing of biological barriers by passive targeting strategies will be presented for intravenous delivery (vascular endothelial lining, particularly for tumor vasculature and blood brain barrier targeting), oral administration (gastrointestinal lining), and upper airway administration (pulmonary epithelium). For each specific barrier, background information will be provided on the structure and biology of the tissues involved as well as available pathways for nano-objects or loaded drugs (diffusion and convection through fenestration, transcytosis, tight junction crossing, etc.). The determinants of passive targeting - size, shape, surface chemistry, surface patterning of nanovectors - will be discussed in light of current results. Perspectives on each mode of administration will be presented. The focus will be on polymeric nanoparticles and dendrimers, although advances in liposome technology will be also reported as they represent the largest body in the drug delivery literature.

Characteristics and properties of nanospheres co-loaded with lipophilic and hydrophilic drug models.

The biphasic nature of polymeric nanospheres prepared by the double emulsion method was exploited to co-encapsulate lipophilic and hydrophilic molecules. All-trans retinoic acid (RA) was selected as a lipophilic drug model whereas calf thymus DNA was chosen as a water-soluble model. Simultaneous quantification of the loaded ingredients was achieved by a second derivative spectrophotometric technique. In addition, prepared batches were fully characterized by atomic force microscopy, porosity measurement, and thermal analysis. Finally, the angiosuppressive action of loaded RA was assessed in a tissue culture model. A blend of either polycaprolactone-multiblock copolymer or the microemulsion technique improved DNA-loading, whereas RA-loading was decreased. DSC data were helpful in explaining the initial phase of RA release from the nanospheres. Along with affinity for the polymeric matrix, the microporosity of nanospheres seemed to play an important role in the diffusion rate and release profiles of both loaded drug models in aqueous medium. The anti-angiogenic effect of microencapsulated RA was generally more pronounced than that of the free drug, and its inhibitory action was maintained for the 14-day study period. Moreover, a relationship was observed between the release profiles and anti-angiogenic properties of the batches tested.

Microemulsion and diafiltration approaches: an attempt to maximize the global yield of DNA-loaded nanospheres.

The yield of DNA-loaded nanospheres in its widest definition includes encapsulation efficiency and the integrity of the loaded molecules plus the production yield of fabricated nanospheres. The former aspect could be considerably improved by adopting the microemulsion concept to enhance the stability of the primary emulsion during the preparation of nanospheres by the double emulsion solvent-removal method. The droplet size of the mentioned emulsion was monitored by means of photon electron correlation spectroscopy and could serve as an index for emulsion fineness and stability. DNA stability as a function of applied mechanical stress was monitored by horizontal agarose gel electrophoresis. The impact of the primary emulsion on nanosphere porosity was assessed as well. Regarding the second aspect of the global yield of nanospheres, i.e. production yield, a modified diafiltration technique was adopted for the washing and recovery processes in comparison with the traditional and for the conservation of particle size characteristics of the recovered nanospheres.

Preparation of hydrogel hollow particles for cell encapsulation by a method of polyester core degradation.

Implantation of encapsulated cells in particles of less than 1 mm (micro-encapsulation) has been proposed as a cell synthesized bio-molecule delivery system. Encapsulation provides immuno-isolation, protecting foreign cells from host immune system while nutrients, oxygen and therapeutic products can diffuse freely across capsule walls. A new method is described for the synthesis of a new family of hollow microparticles for cell encapsulation. Unlike other micro-encapsulation methods, encapsulation in those devices will take place after capsule synthesis, by micro-injection. The microcapsules were prepared by a three-steps original procedure: first, synthesis of a core particle, followed by coating with a layer of epichlorohydrin cross-linked amylo-pectin gel and, finally, selective degradation of the core particle to create the cavity. Initial experiments make use of amylo-pectin cross-linked with trimetaphosphate as core particle material. However, selective degradation was difficult to achieve. In further essays, polyesters were used successfully for the preparation of core particles. Optimizations were carried out and the permeability and morphology of the hollow particles were investigated. The preliminary results show that the new method has the potential to become a standard procedure to obtain hydrogel hollow particles. Moreover, the permeability study seems to be in accordance with specifications for immuno-isolation.

Injectable nanospheres from a novel multiblock copolymer: cytocompatibility, degradation and in vitro release studies.

The aim was to develop and characterize nanospheres made from a newly synthesized poly (D,L-lactide-co-ethyleneglycol) (-PLA-PEG-PLA-)n multiblock copolymer. Nanospheres were prepared under optimized conditions of modified emulsion-solvent evaporation technique in a continuous flow process using rhodamine B as a drug model. They were characterized for size distribution, zeta (zeta) potential, porosity and morphology. Drug loading and yield were also determined. In vitro degradation studies of the copolymer were conducted in phosphate buffer (pH 7.4) at 37 degrees C. The cytotoxic properties of the polymer and vector were analysed by dimethylthiazoldiphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays on the B16 mouse cell line. Release of rhodamine B from the nanospheres was assayed in vitro using a dialysis bag in isotonic phosphate buffer (pH 7.4) at 37 degrees C. Spherical and non-porous nanospheres with mean size less than 800 nm could be prepared. The (zeta) potential was neutral. The average yield was approximately 70% with 7% rhodamine loading. A total of 50% of the multiblock underwent initial degradation after 4 weeks, while degradation was complete after 16 weeks. Cellular proliferation was not inhibited as no cytotoxicity was observed with the copolymers and nanospheres. Rhodamine B was released in a stepwise pattern. The initial burst was 20%, and release was prolonged thereafter for 29 days. Thus, injectable nanospheres with prolonged rhodamine B release have been designed and characterized as a potential drug-delivery system.

Structural modeling of drug release from biodegradable porous matrices based on a combined diffusion/erosion process.

Biodegradable, porous microspheres exhibit a wide range of release profiles. We propose in this paper a unifying approach based on the dual action of diffusion and erosion to establish which mechanisms are responsible for the variety of release kinetics observed during in vitro experiments. Our modeling procedure leads to the partitioning of the matrix into multiple, identical elements, thus simplifying significantly the mathematical and numerical treatment of the problem. The model equations cannot be solved analytically, since the domain contains a moving interface, and must therefore be solved numerically, using specific methods designed for that purpose. Our model confirms the major role that the relative dominance between diffusion and erosion plays in the release kinetics. In particular, the velocity of erosion, the effective diffusion coefficient of the drug molecule in the wetted polymer, the average pore length, and the initial pore diameter are sensitive parameters, whereas the porosity and the effective diffusion coefficient of the drug in the solvent-filled pores is seen to have little influence, if any, on the release kinetics. The model is confirmed by using release data from biodegradable microspheres with different ratios of low and high molecular weight PLA. Excellent goodness of fit is achieved by varying two parameters for all types of experimental kinetics: from the typical square root of time profile to zero-order kinetics to concave release curves. We are also able to predict, by interpolation, release curves from microspheres made of intermediate, untested ratios of PLA by using a relation between two model parameters.

Thermosensitive chitosan-based hydrogel containing liposomes for the delivery of hydrophilic molecules.

A novel injectable in situ gelling thermosensitive chitosan-beta-glycerophosphate (C-GP) formulation has been recently proposed for tissue repair and drug delivery. The system can sustain the release of macromolecules over a period of several hours to a few days. However, with low-molecular-weight hydrophilic compounds, the release is generally completed within 24 h. In this study, liposomes were added to the C-GP solution and their effect on the viscoelastic properties of the system and release kinetics of encapsulated carboxyfluorescein was investigated. The gelation rate and gel strength were slightly increased by the presence of the liposomes. The in vitro release profiles demonstrated controlled delivery over at least 2 weeks. The release rate strongly depended on the liposome size and composition (i.e. addition of cholesterol), and on the presence of phospholipase in the release medium. The kinetics was not substantially modified when using liposomes prepared with a negatively-charged lipid or a lipid having a high phase transition temperature. These results indicate that the liposome-C-GP system rapidly gels at body temperature, and can sustain the delivery of low-molecular-weight hydrophilic compounds. A mathematical model was proposed to characterize the release kinetics.

Factors affecting the sterility of work areas in barrier isolators and a biological safety cabinet.

Factors affecting the sterility of the work area in barrier isolators and a biological safety cabinet (BSC) were studied. A study in a mother-and-child tertiary care teaching hospital in Canada examined the effects of a range of variables on the sterility of work areas in four barrier isolators and a standard BSC. The variables consisted of type of equipment (barrier isolator or BSC), day of the week (Monday through Thursday), time of day (0800-1000, 1000-1200, 1200-1400, and 1400-1600), sampling site (16 surfaces and 5 air sedimentation zones), type of product prepared (antimicrobial, total parenteral nutrient solution, etc.), cleaning procedure (before or after primary cleaning), and level of product preparation activity (none to intense). A total of 657 surface and air sedimentation samples, 327 plated onto Trypticase soy agar (TSA) and 330 onto Sabouraud dextrose agar (SAB-D), were taken during a 20-day period. Thirty-three (5%) of the samples yielded microbial growth when cultured (24 on TSA and 9 on SAB-D). A total of 74 isolates were identified, including Bacillus, Staphylococcus, Penicillium, Micrococcus, Corynebacterium, and Mucor species. Single-variable analysis showed that sampling site, sample type, the time of day samples were taken, and the types of equipment contributed significantly to microbial growth in the samples taken. Several variables were associated with microbial growth in samples from the work areas of barrier isolators and a BSC. More study is needed to compare BSCs and barrier isolators with respect to sterility.

Stability of Morphine-Ketamine Mixtures in 0.9% Sodium Chloride Injection Packaged in Syringes, Plastic Bags and MEDICATION CASSETTE Reservoirs.

The purpose of this study was to determine the stablilty of different intravenous mixtures of morphine and ketamine in 0.9% sodium chloride injection, USP packaged in plastic syringes, plastic bags and MEDICATION CASSETTE reservoirs (SIMS-Deltec, Inc., Minneapolis, MN). Nine morphine (1-,10-,and 25-mg/mL) and ketamine (1-,10-,and 25-mg/mL) mixtures in syringes; two morphine and ketamine mixutres (1 mg/mL and 25 mg/mL of each drug) in plastic bags: and one morpnhine and ketamine mixture (25 mg/mL of each drug) in a MEDICATION CASSETTE reservoir were investigated using a high-performance liquid chromatography analytical technique to measure drug concentrations after one to six days at room temperatue. No loss of drug was observed during the study.

Influence of the Fractal Character of Model Substances on their Reactivity at Solid-Liquid Interfaces.

The fractal theory has been applied to study surface complexity where surface aspects influence physical properties and play a role in controlling heterogeneous reactions at interfaces. In this work, the influence of the fractal character of some selected antacids was investigated in regard to their neutralizing activity. The materials used were magnesium trisilicate, magnesium hydroxide, and heavy and light magnesium oxide, each from three different manufacturers. Surface area, total pore volume, and particle size were measured. Fractal dimension was determined from gas adsorption data according to pore size distribution, the Frenkel-Halsey-Hill, and thermodynamic methods. The results obtained show a correlation between neutralization activity and fractal character rather than total surface area or particle size. Also, the effect of porosity in terms of total pore volume was modified by the structure of the porous network. The complexity of the pore network played a major role in controlling reactivity. However, the effect of surface roughness was only demonstrated when adsorption on the surface was a rate-limiting step in the reaction. Copyright 1999 Academic Press.

Influence of surface properties at biodegradable microsphere surfaces: effects on plasma protein adsorption and phagocytosis.

OBJECTIVE: The objective of this work was to determine plasma protein adsorption and macrophage phagocytosis of biodegradable polyanhydride, polylactic acid and polylactic-co-glycolic acid microspheres prepared by both spray-drying and solvent evaporation techniques. METHODS: Microspheres were characterized by scanning electron microscopy (SEM), confocal laser microscopy, particle size distribution and zeta (zeta) potential determination. Plasma protein adsorption onto the microspheres was determined using a fluoroaldehyde reagent. Phagocytosis was evaluated by incubating microspheres containing the angiotensin II antagonist, L-158,809, with the macrophages in the presence or absence of the phagocytosis inhibitor cythochalasin D. The extent of phagocytosis was established by fluorescence determination of L-158,809 and by optical microscopy. The effect of amphiphilic poly(ethylene glycol) (PEG) derivatives on phagocytosis was determined using PEG-distearate incorporated into the microspheres. RESULTS: The average diameter of the microspheres, which depended on the polymer and the initial formulation, ranged from 0.9 to 3.2 micrometers. Zeta potential studies showed strong negative values irrespective of the polymer used for the spray-dried formulations. The zeta potential was masked by the incorporation of PEG 400- or PEG 1,400-distearate in the formulation. Confocal laser microscopy showed a homogenous dispersion of PEG (measured as PEG-fluorescein) in the microspheres. Protein adsorption was not observed for any of the microsphere formulations following incubation with bovine serum. Incubation of microspheres with murine macrophages showed that PEG-distearate inhibited phagocytosis at appropriate levels (0.1% w/w). Higher levels > 1% w/w of PEG-distearate) resulted in enhanced association with macrophages, despite the presence of the phagocytosis inhibitor cytochalasin D, indicating fusion between the microspheres and the plasma membrane. CONCLUSIONS: These results demonstrate that spray-dried PEG-containing microspheres can be manufactured and that an appropriate concentration of this excipient in microspheres results in decreased phagocytosis.

Improved activity of a new angiotensin receptor antagonist by an injectable spray-dried polymer microsphere preparation.

PURPOSE: To characterize and evaluate in vitro and in vivo the release mechanisms involved in spray-dried biodegradable microspheres having different Poly(D,L-lactide) blend formulations and containing an antihypertensive drug (L-158,809). METHODS: Microspheres and blended polymers were characterized by DSC, SEM, confocal laser microscopy and size analysis. In vitro release studies were evaluated by using microspheres made from various blends of high and low molecular weight polymer. In vivo studies were evaluated by L-158,809 antagonist AT1 function versus the shift of the normal dose-response curve of blood pressure induced by Angiotensine II. RESULTS: The average yield of L-158,809 microspheres (10% (w/w)) was 95% of the theoretical loading. The average diameter of the microspheres was from 1 to 3 micrometers. In all release experiments, a significant burst effect (< 15%) was observed followed by a near zero-order release kinetics. In vivo studies with two different formulations show a strong shift of angiotensin II dose-response curve. CONCLUSIONS: The release kinetics and photomicrographs suggest that the system is best described as a multi-parameter controlled released system in which the drug is molecularly dispersed. In vivo results demonstrating the controlled release of L-158,809.