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Chitosan-modified dry powder formulations for pulmonary gene delivery.

PURPOSE: Spray-drying is an effective process for preparing micron-dimensioned particles for pulmonary delivery. Previously, we have demonstrated enhanced dispersibility and fine particle fraction of spray-dried nonviral gene delivery formulations using amino acids or absorption enhancers as dispersibility-enhancing excipients. In this study, we investigate the use of the cationic polymer chitosan as a readily available and biocompatible dispersibility enhancer. METHODS: Lactose-lipid:polycation:pDNA (LPD) powders were prepared by spray-drying and post-mixed with chitosan or spray-dried chitosan. In addition, the water-soluble chitosan derivative, trimethyl chitosan, was added to the lactose-LPD formulation before spray-drying. RESULTS: Spray-dried chitosan particles, displaying an irregular surface morphology and diameter of less than 2 microm, readily adsorbed to lactose-LPD particles following mixing. In contrast with the smooth spherical surface of lactose-LPD particles, spray-dried trimethyl chitosan-lactose-LPD particles demonstrated increased surface roughness and a unimodal particle size distribution (mean diameter 3.4 microm), compared with the multimodal distribution for unmodified lactose-LPD powders (mean diameter 23.7 microm). The emitted dose and in vitro deposition of chitosan-modified powders was significantly greater than that of unmodified powders. Moreover, the inclusion of chitosan mediated an enhanced level of reporter gene expression. CONCLUSIONS: In summary, chitosan enhances the dispersibility and in vitro pulmonary deposition performance of spray-dried powders.

Biocompatible Materials↗

Moisture-dependent crystallization of amorphous lamotrigine mesylate.

A commercially available computer-controlled vacuum moisture balance was used for determining moisture sorption isotherms of freeze-dried and spray-dried lamotrigine mesylate drug substance and freeze dried drug product containing mannitol. The presence or absence of desorption hysteresis and the characteristics of the weight-versus-time profile as a sample was exposed to a defined relative humidity ramp were sensitive indicators of moisture-induced crystallization. Combination of the moisture sorption data with polarized light microscopy, differential scanning calorimetry, and X-ray powder diffraction provided qualitative verification of the crystallization with < 50 mg of sample. The normalized water loss during crystallization was used to detect as little as 2% amorphous content in physical mixtures of amorphous and crystalline lamotrigine mesylate. Moisture sorption, water plasticization, and crystallization properties of amorphous forms prepared by spray drying and freeze drying were nearly identical. Cofreeze-drying lamotrigine mesylate with D-mannitol resulted in a mixture of amorphous lamotrigine mesylate with properties similar to those of spray-dried or freeze-dried materials and crystalline D-mannitol. The amount of water needed for crystallization over a time scale observable in the moisture balance was considerably more than the amount needed to lower the glass transition temperature of the sample to the operating temperature of the instrument. This result illustrated the importance of time scale effects in determining critical moisture levels for crystallization from the amorphous state.

Crystallization↗

Preparation and characterization of ampicillin loaded methylpyrrolidinone chitosan and chitosan microspheres.

Ampicillin was embedded in microparticles made of a new derivative of chitosan: methylpyrrolidinone chitosan. They were prepared using different drug-to-polymer weight ratios and by a spray-drying technique. Spray-dried drug-loaded chitosan microspheres were prepared for comparison. The microparticles were characterized by scanning electron microscopy (SEM), particle size analysis, differential scanning calorimetry (DSC) and in vitro drug release. Microbiological assay was performed using different bacterial strains. Spray-dried microspheres of almost spherical shape, smooth surface and narrow size distribution were always obtained. Ampicillin loaded into both polymer matrices showed amorphous behaviour as determined by DSC. Drug-loaded microspheres resulted to control the drug release in a 30-120 min range, depending on chitosan type. Thermal denaturation of the microspheres does not modify drug release rate. The results of the microbiological assay show that the loading of ampicillin into chitosans is able to maintain or improve the anti-bacterial activity of the drug.

Ampicillin↗

Microparticle and liquid formulation of a novel HIV protease inhibitor.

The purpose of this study was to develop an improved dosage form of a novel protease inhibitor, LB71350. To overcome the dissolution rate-limiting step in oral absorption, amorphous LB71350 formulation was developed by spray drying ethanol solution of LB71350 and lecithin. Spray drying resulted in free flowing spherical microparticles with diameter < or = 5 microns. Powder X-ray diffraction confirmed that LB71350 in spray-dried microparticles and its aqueous dispersion was amorphous. In contrast to the aqueous suspension of crystalline LB71350, dispersion prepared from spray-dried microparticles showed significantly higher oral bioavailability in rat. Aqueous dispersion of spray-dried microparticles was more palatable than that of co-solvent solution formulation. Stability of dispersions depended on the concentration of dispersion and storage temperature. Dispersion containing 50 mg LB71350/mL was stable at 4 degrees C for 6 weeks without any significant physical or chemical changes. However, massive aggregation and crystallization of LB71350 occurred after 3 weeks at 25 degrees C. Dispersion containing 25 mg LB71350/mL showed sedimentation, which was re-dispersible by gentle shaking. When dispersion stored for 4 weeks at 4 degrees C was given orally to rat, plasma concentration profiles were similar to those obtained after administration of fresh sample. On the basis of these results, the dispersion can be stored at least for 4 weeks at 4 degrees C. Spray-dried microparticles have been stable for 1 year at 4 degrees C without drug crystallization and further study is in progress to establish long-term storage stability. The present study establishes the feasibility of LB71350 liquid dosage formulation that is composed of free flowing spray-dried microparticles and aqueous vehicle to be reconstituted at the time of dispensing.

Amides↗

Primary microparticles and agglomerates of morphine for nasal insufflation.

The aim of this work was to study the characteristics of powders of morphine HCl suitable for nasal administration to be employed for pain treatment as alternative to injection. Primary microparticles of morphine were prepared by spray drying of aqueous drug solutions using sugars or sugar derivatives as drying protectors and particle shapers. The spray drying procedure modified morphine crystallinity making the substance amorphous and affecting its stability in dependence on the excipient employed. A tendency of the spray-dried powders to turn to varying degrees of yellow was observed. Tumbling the powder in a rotating pan allowed the agglomeration of the primary microparticles. Agglomerates were also obtained by tumbling a mixture of morphine crystals and spray-dried microparticles of excipients, with advantages for the stability of the preparation. A nasal device quantitatively insufflated all the morphine agglomerates. The in vitro transport of morphine through rabbit nasal mucosa was faster using nasal powders than with the saturated solution of morphine. Lactose was the most effective excipient for agglomerate manufacturing and delivery of spray-dried morphine. The agglomerates of morphine crystals mixed with mannitol/lecithin microparticles showed superior stability. However, the drug permeation through rabbit mucosa was slower than with spray-dried morphine microparticle agglomerates.

Administration, Inhalation↗

Complexation with tolbutamide modifies the physicochemical and tableting properties of hydroxypropyl-beta-cyclodextrin.

The physicochemical and tableting properties of hydroxypropyl-beta-cyclodextrin (HP-beta-CD) and its tolbutamide (TBM) complex were studied. The kinetics of TBM/HP-beta-CD inclusion complex formation in solution were determined by the phase solubility method. Solid complexes were prepared by freeze-drying and spray-drying. Water sorption-desorption behaviour of the materials were studied and compacts were made using a compaction simulator. TBM and HP-beta-CD formed 1:1 inclusion complexes in aqueous solution with an apparent stability constant of 63 M(-1). HP-beta-CDs and TBM/HP-beta-CD complexes were amorphous whereas the freeze-dried and spray-dried TBMs were polymorphic forms II and I, respectively. Sorption-desorption studies showed that HP-beta-CDs were deliquescent at high relative humidities. TBM/HP-beta-CD complexes had slightly lower water contents at low relative humidities than the physical mixtures. However, at high humidities their water sorption and desorption behaviours were similar to those of corresponding physical mixtures, indicating a glass transition of the complexed materials. TBM/HP-beta-CD complexes demonstrated a worse compactability than similarly prepared HP-beta-CDs or physical mixtures. Also particle properties that resulted from these preparation methods affected the compactability of the materials. In conclusion, the physicochemical and tableting properties of HP-beta-CD were modified by complexation it with TBM.

2-Hydroxypropyl-beta-cyclodextrin↗

Physicochemical characterization and in vitro dissolution behavior of nicardipine-cyclodextrins inclusion compounds.

Inclusion complexation between nicardipine hydrochloride (NC), a calcium-channel antagonist, and beta-cyclodextrin (beta-CD) or hydroxypropyl-beta-cyclodextrin (HPbetaCD) was evaluated in aqueous environment and in solid state. The phase solubility profiles with both cyclodextrins (CDs) were classified as A(L)-type, indicating the formation of 1:1 stoichiometric inclusion complexes. Stability constants (Ks) were calculated from the phase solubility diagrams and were found to be pH dependent. More stable NC:CDs complexes were formed in alkaline medium in which the drug is in its non-ionized form. Binary systems of NC with CDs, prepared experimentally by different techniques (kneading, evaporation, freeze-drying and spray-drying), were investigated by differential scanning calorimetry, Fourier transformation-infrared spectroscopy, X-ray diffractometry and scanning electron microscopy. From this analysis, evaporation, freeze-drying and spray-drying were found to produce inclusion complexes. In contrast, crystalline drug was still clearly detectable in the kneaded products. The dissolution profiles of the obtained powders were studied in order to define the most appropriate CD and preparation method to originate inclusion complexes, which will be used in the development of a new controlled release formulation of NC. Both the preparation and nature of carrier played an important role in the dissolution performance of the system. However, independently of the preparation technique, all the combinations with HPbetaCD were more effective in achieving the enhancement of the NC dissolution rate, yielding better performances than the corresponding ones with betaCD.

Calcium Channel Blockers↗

[Study on the preserving techniques of squeezed juice of fresh ginger and Glutinous rehannia].

OBJECTIVE: To study the preserve techniques of squeezed juice of Chinese medicinal materials. METHOD: The techniques of refrigeration, rapid freezing, 60Cogamma-ray sterilization, freeze-drying and spray-drying were used for preservation of squeezed juice of Ginger and Glutinous Rehannia. Different results were compared. RESULT: The period of preservation was half or one year. CONCLUSION: The rapid freezing, freeze-drying and spray-drying are suitable for preservation of squeezed juice of Chinese medicinal materials.

Cobalt Radioisotopes↗

Preparation of DNA dry powder for non-viral gene delivery by spray-freeze drying: effect of protective agents (polyethyleneimine and sugars) on the stability of DNA.

This study investigates the feasibility of using the process of spray-freeze drying (SFD) to produce DNA dry powders for non-viral gene delivery. The effect of protective agents was assessed on the stability of DNA dry powders after SFD. The process of SFD had adverse effects on the tertiary structure of DNA with the protective agents of sucrose, trehalose and mannitol. With the protection of these sugars, a band corresponding to the linear form of DNA was observed during gel electrophoresis between the supercoiled form (SC) and the open circular (OC) form. On the contrary, excess cationic condensing polyethyleneimine (PEI), in conjunction with the above sugars, had the ability to provide protection for DNA from degradation after SFD. This is indicated by the reservation in SC and OC forms of DNA during agarose gel electrophoresis. The electrostatic forces between PEI polymer and DNA are critical for providing protection against various stresses generated by the process of SFD. Furthermore, on rehydration, the particle size and zeta potential of PEI/DNA complexes at weight ratios 3:1 of SFD dry powders were well maintained. Also, no transfection activity loss of PEI/DNA complexes at weight ratios 3:1 on NIH/3T3 cells was observed for reconstituted powders as compared with untreated control solutions. These results give a better understanding of preparing stable DNA dry powders by the process of SFD.

Animals↗

An alternative source of blood protein for screwworm (Diptera: Calliphoridae) larval diet.

Larvae of screwworm, Cochliomyia hominivorax (Coquerel), were reared on diets with varying concentrations of spray dried animal blood cells, spray dried inedible egg product, and powdered milk substitute, and compared with those reared on spray dried whole bovine blood-based diet currently used for mass rearing. Mean larval weight, pupal weight, number of pupae, adult emergence, sex ratio, longevity, fecundity, and fertility obtained from the tests with animal blood cell based diets were similar to those from tests with whole bovine blood based diet. The animal blood cell based diet has potential for use in the mass production of screwworm larvae.

Animals↗

Characterization of the thermal properties of powder particles using microthermal analysis.

PURPOSE: Microthermal analysis is a recently introduced thermoanalytical technique whereby discrete regions of complex samples may be scrutinized at micron or submicron resolution. In this investigation, the novel use of the technique to study individual powder particles is described. METHODS: A TA Instruments microthermal analyzer was used in local thermal analysis mode using a heating rate of 10 degrees C/s. Powder samples of crystalline ibuprofen, spray-dried salbutamol sulphate, spray-dried and crystalline trehalose, and two polymorphic forms of indometacin were studied using differential scanning calorimetry as a supportive technique as appropriate. RESULTS: The ibuprofen showed a probe position discontinuity corresponding to the melting point of the material. Spray-dried salbutamol sulphate showed a discontinuity corresponding to decomposition but not to the glass transition, whereas both crystalline and amorphous trehalose showed a single discontinuity corresponding to physical collapse of the material. Studies using the alpha and gamma forms of indometacin showed that the technique was able to distinguish between the two polymorphic forms. CONCLUSION: The study suggests that micro-TA may be used to assess interparticulate composition and homogeneity.

Albuterol↗

Evaluation of animal protein supplements in diets of early-weaned pigs.

Three growth assays were conducted to determine the efficacy of replacing dried skim milk and(or) dried whey in diets of starting pigs with commercially available spray-dried porcine plasma, spray-dried porcine blood, spray-dried bovine plasma, or spray-dried extracted meat protein. In Exp. 1, 236, 24-d-old crossbred pigs were fed diets containing either skim milk and whey or porcine plasma from 0 to 14 d postweaning and whey or porcine plasma from 14 to 28 d. Although pigs fed diets containing porcine plasma had greater ADFI and ADG than those fed milk products from 0 to 7 d, no differences were observed from d 0 to 14 or from 14 to 28 d postweaning. In Exp. 2, 204, 21-d-old pigs were fed corn-soybean meal-based diets using the following supplemental protein source combinations: skim milk and whey; skim milk, whey, and casein; porcine plasma, whey, and lactose/starch (10%); porcine plasma and lactose/starch (24.4%); or whey. A common diet (1.25% lysine, 10% whey) was fed from 14 to 35 d postweaning. Pigs fed diets containing porcine plasma consumed more feed and had greater ADG than others (P < .05) from 0 to 14 d and from 0 to 35 d. Both ADG and ADFI were highest when the diet contained 10.3% porcine plasma, 20% whey, and 10% added lactose. Experiment 3 used 150, 21-d-old pigs to compare the inclusion of skim milk, porcine plasma, porcine blood, bovine plasma, or meat extract in diets fed from 0 to 14 d postweaning. A common diet (the same as in Exp. 2) was fed from 14 to 35 d. Pigs fed porcine plasma had greater ADFI (P < .05) from 0 to 14 d than pigs fed other treatments. Also, pigs fed porcine plasma had greater ADG (P < .05) from 0 to 14 d than pigs fed all other diets except porcine blood. Pigs fed porcine blood had the largest ADFI (P < .05) from 14 to 35 d compared with pigs fed other diets. These experiments indicate that porcine plasma is a protein supplement superior to skim milk in diets of starting pigs and that porcine blood has a positive influence on subsequent growth performance.

Animal Feed↗

Protein spray freeze drying. 2. Effect of formulation variables on particle size and stability.

Spray freeze drying produces protein particles suitable for microencapsulation into polymeric microspheres intended for sustained release. Accessibility of encapsulated protein particles to the microsphere surface increases as the protein particle size is increased. Thus, it is desirable that the encapsulated protein particle size be minimized to limit initial release. We have investigated the effect of formulation on spray freeze-dried bovine serum albumin (BSA) as a model protein. Atomization conditions were fixed such that in the absence of excipient, the particle size of the sonicated powder was submicron, and there was substantial protein degradation (loss of monomer). Addition of low concentrations of surfactants (up to the CMC) or mannitol (up to the point where it tended to crystallize upon dehydration) resulted in partial stabilization without impacting particle size. Trehalose was successful in stabilizing the protein; however, there was a marked increase in particle size at the highest levels tested. Ammonium sulfate provided partial stabilization, but also tended to form crystals and increase particle size. FTIR measurements showed a loss of native secondary structure upon spray freeze drying that was ameliorated by addition of trehalose. Other excipients did not prevent structural perturbations. In general, stabilization of spray freeze-dried BSA was related to lowering of the specific surface area in the powder. A balance must be achieved when spray freeze drying proteins intended for encapsulation in sustained-release systems.

Chemistry, Pharmaceutical↗

Optimization of topical cidofovir penetration using microparticles.

Cidofovir is a new class of antiviral agent with potent in vitro and in vivo activity against a broad spectrum of herpes viruses. The aim of this work was to obtain a prolonged therapeutic effect of cidofovir in the basal epidermis after its topical application. For this purpose, poly(lactide-co-glycolide) (PLGA) microparticles were prepared by solvent evaporation and spray-drying methods. Microparticles prepared by spray-drying showed a encapsulation efficiency of 80%. Conversely, for all the microspheres prepared by the W/O/W solvent evaporation method the encapsulation efficiency was low. Also, microparticles prepared by spray-drying showed a higher burst release. Skin penetration and distribution experiments were carried out with cidofovir-loaded microparticles prepared by spray-drying, since these carriers presented the best characteristics in terms of size and encapsulation efficiency. A cidofovir solution in 0.2% PVA served for comparison. Penetration experiments were carried out in Franz type diffusion cells with an available diffusion area of 1.76 cm(2), using porcine skin. The results obtained showed that the amount of cidofovir penetrated, over a 24 h time period, was higher with the drug solution than with microparticles. Cidofovir distribution in porcine skin, after topical application of microparticles and drug solution for 24 h, was determined by horizontal slicing of the skin. The profiles obtained for the two formulations showed that the quantity of cidofovir retained in the skin decreased with the depth. Besides the amount of cidofovir found in the basal epidermis (120-150 microm) was much higher with microparticles than with the control solution. These data showed that cidofovir-loaded microparticles could improve cidofovir topical therapy since these vehicles increased drug retention in the basal epidermis and decreased its penetration through the skin.

Administration, Topical↗

A necessary modification to the preparation of papain from any high-quality latex of Carica papaya and evidence for the structural integrity of the enzyme produced by traditional methods.

A method of preparation of papain (EC 3.4.22.2) from relatively soluble types of latex of Carica papaya, including spray-dried latex produced by a controlled and relatively mild process, was devised. Spray-dried latex dissolves easily in water up to 350mg/ml at 22 degrees C, which corresponds to approx. 230mg of protein/ml. When the usual method of preparation of crystalline papain contaminated only by its oxidation products, developed by Kimmel & Smith [J. Biol. Chem. (1954) 207, 515-531], is applied to spray-dried latex, the result is a preparation of papain heavily contaminated by chymopapains A and B (EC 3.4.22.6), and to a lesser extent by papaya peptidase A. This applies also to other types of papaya-latex currently commercially available, which, though less soluble than spray-dried latex, are more soluble than the types of latex available when the method of Kimmel & Smith (1954) was developed. This contamination is avoided by adjusting the concentration of the initial latex extract to 65mg of protein/ml (or less) before salt fractionation. For spray-dried latex this corresponds to 100mg of latex/ml. Papain isolated from spray-dried latex was characterized by using 2,2'-dipyridyl disulphide and 4-chloro-7-nitrobenzofurazan as thiol-specific reactivity probes and alpha-N-benzoyl-l-arginine ethyl ester as substrate. Papain isolated from this source appears to have the same catalytic-centre characteristics as papain isolated previously from latex produced by harsher methods. The catalysis of the hydrolysis of alpha-N-benzoyl-l-arginine ethyl ester by the mixture of thiol proteinases extracted from spray-dried latex by application of the method of Kimmel & Smith (1954) appears to obey Michaelis-Menten kinetics. The presence of the other enzymes results in an increase in the value of K(m) and a decrease in the catalytic-centre activity (k(cat.)) relative to the values for the catalysis by papain.

2,2'-Dipyridyl↗

Particle size and density in spray drying-effects of carbohydrate properties.

The purpose of this study was to examine some fundamental aspects of the particle formation during spray drying, related to particle size and density. Particles were prepared in a laboratory spray dryer from carbohydrates with different solubility and crystallization propensity, such as lactose, mannitol, and sucrose/dextran 4:1. The feed concentrations ranged from 1% w/w to saturated and the size of droplets and particles were measured by laser diffraction. Particles were also characterized by various microscopy techniques (i.e., scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), and light microscopy), differential scanning calorimetry (DSC), gas adsorption, and gas pycnometry. As demonstrated larger particles could be obtained by either increasing the droplet size during atomization; increasing the concentration of the feed solution; or decreasing the solubility of the solute. The apparent particle density, measured by gas pycnometry, was found negatively correlated to the feed concentration. Due to the nonlinear relationship between the feed concentration and the particle size, it was concluded that higher solids load would cause an increase in the effective particle density and that the reduction in the apparent particle density was a result of a gradually less permeable particle surface. Further, the crystallization propensity of the carbohydrate influenced the particle formation and resulted in either hollow or porous particles.

Aerosols↗

Effect of drying methods on retention of moist sucralfate gel properties.

The aim of this work was to find a drying procedure for moist sucralfate gel capable of producing dried sucralfate gel that retains the original gel properties of bioadhesion, rheology, and micromeritics. Spray-drying and microwave-drying procedures were employed. Mannitol was used as a gel-protective substance during the drying processes. The spray drying of moist sucralfate gel gave rise to a powder whose water suspensions showed significantly reduced viscosity. The bioadhesion of spray-dried sucralfate gel was strongly reduced by drying. When mannitol was used as a gel protector, the spray-dried sucralfate in part maintained the original bioadhesion of moist sucralfate gel. The preparation of a dried sucralfate gel retaining the bioadhesion characteristics, avoiding the use of mannitol, was made possible using the microwave-drying procedure. The microwave-dried product possesses a granular morphology suitable for direct compression because it is a free flowing and strongly coherent granular powder.

Acid-Base Equilibrium↗

Buckling and crumpling of drying droplets of colloid-polymer suspensions.

Spray drying of complex liquids to form solid powders is important in many industrial applications. One of the challenges associated with spray drying is controlling the morphologies of the powders produced; this requires an understanding of how drying mechanics depend on the ingredients and conditions. We demonstrate that the morphology of powders produced by spray drying colloidal polystyrene (PS) suspensions can be significantly altered by changing the molecular weight of dissolved poly(ethylene oxide) (PEO). Samples containing high-molecular-weight PEO produce powders with more crumpled morphologies than those containing low-molecular-weight PEO. Observations of drying droplets suspended by a thin film of vapor suggest that this occurs because the samples with high-molecular-weight PEO buckle earlier in the drying process when the droplets are larger. Earlier buckling times are likely caused by the decreased stability, demonstrated by bulk rheology experiments, of PS particles in the presence of high-molecular-weight PEO at elevated temperatures. We present a consistent picture in which decreased particle stability hastens droplet buckling and leads to more crumpled powder morphologies; this underscores the importance of interparticle forces in determining the buckling of particle-laden droplets.

Journal Article↗