Enhanced Dispersibility and Deposition of Spray-dried Powders for Pulmonary Gene Therapy

Spray-drying represents a viable alternative to freeze-drying for preparing dry powder dispersions for delivering macromolecules to the lung. The dispersibility of spray-dried powders is limited however, and needs to be enhanced to improve lung deposition and subsequent biological activity. In this study, we investigate the utility of leucine as a dry powder dispersibility enhancer when added prior to spray-drying a model non-viral gene therapy formulation (lipid:polycation:pDNA, LPD). Freeze-dried lactose–LPD, spray-dried lactose–LPD and spray-dried leucine–lactose–LPD powders were prepared. Scanning electron microscopy showed that leucine increased the surface roughness of spray-dried lactose particles. Particle size analysis revealed that leucine-containing spray-dried powders were unimodally dispersed with a mean particle diameter of 3.12 μm. Both gel electrophoresis and in vitro cell (A549) transfection showed that leucine may compromise the integrity and biological functionality of the gene therapy vector. The deposition of the leucine containing powder was however significantly enhanced as evidenced by an increase in gene expression mediated by dry powder collected at lower stages of a multistage liquid impinger (MSLI). Further studies are required to determine the potential of leucine as a ubiquitous dispersibility enhancer for a variety of pulmonary formulations.     

鲑降钙素吸入粉雾剂的制备及其药剂学性质研究鲑降钙素吸入粉雾剂的制备及其药剂学性质研究

目的喷雾干燥法制备鲑降钙素吸入粉雾剂,考察其主要药剂学特性。方法按中国药典2000年版方法测定鲑降钙素吸入粉雾剂排空率及有效部位沉积量,扫描电镜观察粉粒的形态,激光粒度测定仪测定粉粒的粒径大小及分布,差热分析及X射线粉末衍射考察吸入粉雾剂载体的晶型。结果有效部位鲑降钙素沉积量均在10%以上,排空率均在90%以上;相对湿度在52%以下,粉粒呈圆整的分散状态;当RH=75%时粉粒发生较多的粘连和聚集;粉体平均粒径为1.67 μm;差热分析结果显示:吸入粉雾剂中L-亮氨酸的熔融热较单一组分在喷雾干燥后显著下降;X射线粉末衍射图谱中,吸入粉雾剂载体的衍射峰强度较喷雾干燥前显著下降。结论在鲑降钙素吸入粉雾剂载体系统中加入甘露醇,粉粒圆整,加入L-亮氨酸可形成超低密度载体;扫描电镜的结果提示,应严格控制吸入粉雾剂贮存时环境的湿度;喷雾干燥复合载体更有助于无定形的形成。     

The Effect of Engineered Mannitol-Lactose Mixture on Dry Powder Inhaler Performance

Purpose

To co-crystallise mannitol and lactose with a view to obtaining crystals with more favourable morphological features than either lactose or mannitol alone, suitable for use as carriers in formulations for dry powder inhalers (DPIs) using simultaneous engineering of lactose-mannitol mixtures.

Methods

Mannitol and lactose individually and the two sugars with three different ratios were crystallised/co-crystallised using anti-solvent precipitation technique. Obtained crystals were sieved to separate 63?C90???m size fractions and then characterised by size, shape, density and in vitro aerosolisation performance. Solid state of crystallized samples was studied using FT-IR, XRPD and DSC.

Results

At unequal ratios of mannitol to lactose, the elongated shape dominated in the crystallisation process. However, lactose exerted an opposite effect to that of mannitol by reducing elongation ratio and increasing the crystals?? width and thickness. Crystallised ??-lactose showed different anomers compared to commercial lactose (??-lactose monohydrate). Crystallised ??-mannitol showed different polymorphic form compared to commercial mannitol (??-mannitol). Crystallised mannitol:lactose showed up to 5 transitions corresponding to ??-mannitol, ??-lactose monohydrate, ??-lactose, 5??-/3??-lactose and 4??-/1??-lactose. In vitro deposition assessments showed that crystallised carriers produced more efficient delivery of salbutamol sulphate compared to formulations containing commercial grade carriers.

Conclusion

The simultaneous crystallization of lactose-mannitol can be used as a new approach to improve the performance of DPI formulations.     

Chitosan-based spray-dried respirable powders for sustained delivery of terbutaline sulfate.

In this study, we describe the preparation of highly dispersible dry powders for pulmonary drug delivery that display sustained drug release characteristics. Powders were prepared by spray-drying 30% v/v aqueous ethanol formulations containing terbutaline sulfate as a model drug, chitosan as a drug release modifier and leucine as an aerosolisation enhancer. The influence of chitosan molecular weight on the drug release profile was investigated by using low, medium and high molecular weight chitosan or combinations thereof. Following spray-drying, resultant powders were characterised using scanning electron microscopy, laser diffraction, tapped density analysis, differential scanning calorimetry and thermogravitational analysis. The in vitro aerosolisation performance and drug release profile were investigated using Multi-Stage Liquid Impinger analysis and modified USP II dissolution apparatus, respectively. The powders generated were of a suitable aerodynamic size for inhalation, had low moisture content and were amorphous in nature. The powders were highly dispersible, with emitted doses of over 90% and fine particle fractions of up to 82% of the total loaded dose, and mass median aerodynamic diameters of less than 2.5microm. A sustained drug release profile was observed during dissolution testing; increasing the molecular weight of the chitosan in the formulation increased the duration of drug release.     

Enhanced dispersibility and deposition of spray-dried powders for pulmonary gene therapy

Spray-drying represents a viable alternative to freeze-drying for preparing dry powder dispersions for delivering macromolecules to the lung. The dispersibility of spray-dried powders is limited however, and needs to be enhanced to improve lung deposition and subsequent biological activity. In this study, we investigate the utility of leucine as a dry powder dispersibility enhancer when added prior to spray-drying a model non-viral gene therapy formulation (lipid:polycation:pDNA, LPD). Freeze-dried lactose-LPD, spray-dried lactose-LPD and spray-dried leucine-lactose-LPD powders were prepared. Scanning electron microscopy showed that leucine increased the surface roughness of spray-dried lactose particles. Particle size analysis revealed that leucine-containing spray-dried powders were unimodally dispersed with a mean particle diameter of 3.12 microm. Both gel electrophoresis and in vitro cell (A549) transfection showed that leucine may compromise the integrity and biological functionality of the gene therapy vector. The deposition of the leucine containing powder was however significantly enhanced as evidenced by an increase in gene expression mediated by dry powder collected at lower stages of a multistage liquid impinger (MSLI). Further studies are required to determine the potential of leucine as a ubiquitous dispersibility enhancer for a variety of pulmonary formulations.     

Engineered Mannitol Ternary Additives Improve Dispersion of Lactose–Salbutamol Sulphate Dry Powder Inhalations

The aim of this study was to evaluate the influence of novel engineered fine mannitol particles (4.7%, w/w) on the performance of lactose–salbutamol sulphate dry powder inhaler (DPI) formulations to obtain promising aerosolisation properties. The results showed that the more elongated the fine mannitol particles, the weaker the drug–carrier adhesion, the better the drug content homogeneity, the higher the amount of drug expected to be delivered to the lower airways and the higher the total DPI formulation desirability. Linear relationships were established showing that mannitol particles with a more elongated shape generated powders with broader size distributions and that were less uniform in shape. The weaker the drug–carrier adhesion, the higher the fine particle fraction of the drug is upon aerosolisation. It is believed that more elongated fine mannitol particles reduce the number of drug–carrier and drug–drug physical contact points and increase the ability of the drug particles to travel into the lower airways. Additionally, a lower drug–carrier contact area, lower drug–carrier press-on forces and easier drug–carrier detachment are suggested in the case of formulations containing more elongated fine mannitol particles. Ternary ‘drug-coarse carrier-elongated fine ternary component’ DPI formulations were more favourable than both ‘drug-coarse carrier’ and ‘drug-elongated coarse carrier’ binary formulations. This study provides a comprehensive approach for formulators to overcome the undesirable properties of dry powder inhalers, as both improved aerosolisation performance and reasonable flow characteristics were obtained using only a small amount of elongated engineered fine mannitol particles.     

Influence of potential inhalation carriers on stability of thymopentin in rat bronchoalveolar lavage fluid

Abstract

In the present study, the stability of thymopentin (TP5) in bronchoalveolar lavage fluid (BALF) in presence of potential excipients in inhalation formulation was investigated. The content of TP5 was determined using HPLC method. Commonly used bulking agent, dispersibility enhancers and absorption enhancers in inhalation were investigated with respect to the stability of TP5 in BALF. Finally, the stability of TP5 in two inhalation formulations based on the screening experiments was tested in BALF. The results showed that TP5 alone degraded very rapidly in BALF and zero-order enzymatic reaction with a half-life of t_0.5?=?49.20?min was observed using 10 times diluted BALF. Among the amino acids examined, leucine and phenylalanine effectively inhibited the enzymolysis of TP5 with prolonged half-life of 112.7?min and 136.2?min, respectively. Nevertheless, slight but insignificant inhibition effect was witnessed for tyrosine, aspartic acid, and lysine; and negligible prevention on the degradation process of TP5 were found for lactose and mannitol. Regarding chitosan, irrespective with molecular weight, the formation of chitosan-TP5 complex improved the stability of TP5 with prolonged t_0.5 by 1.8 times. However, along with the improved stability of TP5 in spray-dried chitosan microspheres, the content of TP5 in formulations was reduced to about 75% during preparation process. Thus, leucine was proved to be a prior candidate for inhalation formulation of TP5. Consequently, the results indicate the potential of leucine as carrier for pulmonary delivery of TP5 serving as both stabilizer and dispersibility enhancer.     

硫酸特布他林干粉吸入剂制备工艺的优化研究

目的:优化硫酸特布他林干粉吸入剂的制备工艺。方法:采用喷雾干燥技术制备硫酸特布他林干粉吸入剂,采用双层液体碰撞器测定其体外肺沉积率,扫描电镜观察干粉的表观形貌,热重分析仪测定干粉的水分含量,激光粒度测定仪测定粒径大小,以产品收率、水分含量、粉末的空气动力学粒径及体外肺沉积率为考察指标,通过正交设计结合多指标综合评价法优化最佳制备工艺。结果:通过正交试验-多指标综合评价,最佳制备工艺为:喷雾压力190 kPa,干燥风速0.7 m3.min-1,供液速度7.0 mL.min-1,入口温度120℃。结论:按最佳制备工艺制得的干粉收率为50.54%,水分含量为0.467%,空气动力学粒径为1.80μm,体外肺沉积率为55.19%。正交试验结合多指标综合评价法用于硫酸特布他林干粉吸入剂制备工艺的优化有效可用。     

Preparation and characterization of spray dried inhalable powders containing chitosan nanoparticles for pulmonary delivery of isoniazid

The aim of this study was to prepare spray dried inhalable powders containing isoniazid-loaded chitosan/tripolyphosphate (TPP) nanoparticles for sustained delivery of the drug to the lung. Nanoparticles were prepared by ionic gelation method. In-vitro drug release study indicated that the rate of drug release from nanoparticles was decreased by increasing the amount of chitosan. Entrapment of isoniazid into chitosan/TPP nanoparticles decreased minimum inhibitory concentrations (MIC) of the drug against mycobacterium avium intracellulare. Nanoparticles were spray dried using excipients such as lactose, mannitol and maltodextrin alone or with leucine. Results showed that the obtained powders had different aerosolization property. It was observed that by adding leucine, the particle size of microparticles deceased and the process yield and fine particle fraction (FPF) increased significantly. The in-vitro deposition data indicated that spray drying of isoniazid-loaded nanoparticles with lactose in the presence of leucine resulted in the production of inhalable powders with the highest FPF (45%).     

Formulation and characterization of spray-dried powders containing nanoparticles for aerosol delivery to the lung

Spray-drying is a common practice of powder preparation for a wide range of drugs. Spray-dried powders can be used to deliver particles to the lungs via a dry powder inhaler (DPI). The present study investigated the feasibility of developing a platform for aerosol delivery of nanoparticles. Lactose was used as the excipient and spray-dried with two different types of nanoparticles: gelatin and polybutylcyanoacrylate nanoparticles. Results showed that some carrier particles were hollow while others had a continuous matrix. Gelatin nanoparticles were incorporated throughout the matrix and sometimes accumulated at one end of the lactose. Polycyanoacrylate nanoparticles mostly clustered in different spots within the lactose carriers. The mean sizes of both nanoparticle types were characterized at two different times: before they were spray-dried and after they were redissolved from the spray-dried powders. Both nanoparticle types remained in the nano-range size after spray-drying. The mean nanoparticle sizes were increased by approximately 30% after spray-drying, though this increase was statistically significant only for the gelatin nanoparticles. Dispersion of the powder with an in-house passive dry powder inhaler and subsequent cascade impaction measurements showed that incorporation of the nanoparticles did not affect the fine particle fraction (FPF) or mass median aerodynamic diameter (MMAD) of the powders. FPF was approximately 40% while MMAD was 3.0+/-0.2 microm, indicating the present formulations yield aerosols of a suitable particle size for efficient lung delivery of nanoparticles.The present work demonstrates that nanoparticles can be delivered to the lungs via carrier particles that dissolve after coming in contact with the aqueous environment of the lung epithelium. This opens the way for new drug-targeting strategies using nanoparticles for pulmonary delivery of drugs and diagnostics.     

Preparation of honokiol-loaded chitosan microparticles via spray-drying method intended for pulmonary delivery

It has been demonstrated that spray-drying is a powerful method to prepare dry powders for pulmonary delivery. This paper prepared dispersible dry powders based on chitosan and mannitol containing honokiol nanoparticles as model drug. The results showed that the prepared microparticles are almost spherical and have appropriate aerodynamic properties for pulmonary delivery (aerodynamic diameters was between 2.8–3.3 μm and tapped density ranging from 0.14–0.?18?g/cm³). Moreover, surface morphology and aerodynamic properties of the powders were strongly affected by the content of mannitol. Fourier transform infra-red (FTIR) spectrum of powders indicated that the honokiol nanoparticles were successfully incorporated into microparticles. In vitro drug release profile was also observed. The content of mannitol in powders significantly influenced the release rate of honokiol from matrices.     

载体颗粒表面修饰对其表面能及粉雾剂性能的影响

目的研究载体的表面修饰对粉雾剂性能的影响。方法用不同的润滑剂对乳糖颗粒进行了表面修饰,并利用反向气相法测定了乳糖修饰前后的表面能,考察了表面能对干扰素α-2b粉雾剂沉积性能的影响。结果表面修饰后乳糖颗粒的流动性、表面光滑度都得到较大程度的改善,表面能也相应降低,而粉雾剂的性能得以提高。结论利用反向气相色谱法测定载体的表面能来预测粉雾剂的性能,可以为粉雾剂的处方设计和筛选提供一种快速有力的工具。     

Feasibility of spray drying bacteriophages into respirable powders to combat pulmonary bacterial infections

The use of bacterial viruses for antibacterial treatment (bacteriophage therapy) is currently being reevaluated. In this study, we analyze the potential of processing bacteriophages in a dry powder formulation, using a laboratory spray dryer. The phages were dried in the presence of lactose, trehalose or dextran 35, serving as an excipient to give the resulting powder the necessary bulk mass and offer protection to the delicate phage structure. Out of the three excipients tested, trehalose was found to be the most efficient in protecting the phages from temperature and shear stress throughout the spray drying process. A low inlet air temperature and atomizing force appeared to be the best parameter conditions for phage survival. Pseudomonas podovirus LUZ19 was remarkably stable, suffering less than 1 logarithmic unit reduction in phage titer. The phage titer of Staphyloccus phage Romulus-containing powders, a member of the Myoviridae family, showed more than 2.5 logarithmic units reduction. On the other hand, Romulus-containing powders showed more favorable characteristics for pulmonary delivery, with a high percentage of dry powder particles in the pulmonary deposition fraction (1–5 μm particle diameter). Even though the parameters were not optimized for spray drying all phages, it was demonstrated that spray drying phages with this industrial relevant and scalable set up was possible. The resulting powders had desirable size ranges for pulmonary delivery of phages with dry powder inhalers (DPIs).     

Formulation of pH responsive peptides as inhalable dry powders for pulmonary delivery of nucleic acids

Nucleic acids have the potential to be used as therapies or vaccines for many different types of disease, but delivery remains the most significant challenge to their clinical adoption. pH responsive peptides containing either histidine or derivatives of 2,3-diaminopropionic acid (Dap) can mediate effective DNA transfection in lung epithelial cells with the latter remaining effective even in the presence of lung surfactant containing bronchoalveolar lavage fluid (BALF), making this class of peptides attractive candidates for delivering nucleic acids to lung tissues. To further assess the suitability of pH responsive peptides for pulmonary delivery by inhalation, dry powder formulations of pH responsive peptides and plasmid DNA, with mannitol as carrier, were produced by either spray drying (SD) or spray freeze drying (SFD). The properties of the two types of powders were characterised and compared using scanning electron microscopy (SEM), next generation impactor (NGI), gel retardation and in vitro transfection via a twin stage impinger (TSI) following aerosolisation by a dry powder inhaler (Osmohaler™). Although the aerodynamic performance and transfection efficacy of both powders were good, the overall performance revealed SD powders to have a number of advantages over SFD powders and are the more effective formulation with potential for efficient nucleic acid delivery through inhalation.     

Alternative sugars as potential carriers for dry powder inhalations

Most dry powder inhaler (DPI) formulations rely on lactose monohydrate as a carrier in the drug powder blends. However, lactose cannot be used for compounds that interact with the reducing sugar function of the lactose, such as formoterol, budesonide or peptides and proteins. In this study, alternative carriers like mannitol, glucose, sorbitol, maltitol and xylitol have therefore been evaluated for their potential use in DPI formulations. Raw materials were characterised physico-chemically and blends with the model drug substance budesonide were tested with respect to the aerosolization behaviour of the powders.

It was found out that similarly to the problems known for lactose monohydrate, such as supplier variability, variability between different qualities of one supplier, the same difficulties apply to the alternative carriers investigated. Different sources and qualities of mannitol led to significant differences in the fine particle fraction (FPF), varying from 15 to 50% for two different qualities of mannitol. Similar observations were made for the other carrier materials studied. Also, the influence of conditioning the raw material at different relative humidity was found to have substantial influence on the performance of drug/carrier blends which is characterised by a strong decrease in the FPF. In summary, mannitol showed potential as a drug carrier to be used in DPIs whereas the more hygroscopic sugars only showed poor dispersibility.     

Use of recrystallized lactose as carrier for inhalation powder of interferon a2b

The aim of the present investigation was to evaluate the effects of physical properties of the carrier on the in vitro deposition performance of dry powder inhalations (DPIs) of recombination human interferon a2b (IFN a2b). Recrystallized lactose was used as the carrier. Inverse gas chromatography (IGC) was used to assess the surface energy, and atomic force microscopy (AFM) was used to assess the roughness and topography of the carrier. In vitro performance of the powder blends was strongly correlated to the physical properties of the carrier. Plotting emitted dose (%) vs. flow rate and fine particle fraction vs. surface energy, yielded an R2 value of 0.9621 and 0.9146, respectively.     

Excipient-free nanoporous microparticles of budesonide for pulmonary delivery

The aim of this study was to investigate the application of a spray-drying process for the production of nanoporous microparticles (NPMPs) to budesonide, and to characterise the particles produced in terms of their suitability for pulmonary delivery.Budesonide was spray dried with and without ammonium carbonate from ethanol/water or methanol/water solutions. The solid-state characteristics and micromeritic (particle size, density, surface area) properties of spray dried powders were assessed. In vitro deposition studies were performed to assess aerosol performance.The densities of the NPMPs were significantly lower and the surface areas significantly higher than for non-porous spray dried or micronised material. NPMPs of budesonide demonstrated improved aerosolisation properties compared to spray dried non-porous, micronised material and two budesonide commercial products. All spray dried materials were amorphous in nature. The glass transition temperature (90 °C) was sufficiently high to suggest good physical stability at room temperature. When stored at 25 °C/60% RH NPMPs showed a reduced tendency to recrystallise compared to the equivalent non-porous spray dried powder. The physical stability and amorphous nature of NPMPs was retained, under these storage conditions for at least one year and the in vitro aerosolisation properties were not affected by the storage conditions.Excipient-free porous microparticles, prepared by the novel process described, show good potential for drug delivery by oral inhalation with improved in vitro deposition properties compared to non-porous particles.     

Preparation and characterization of spray dried inhalable powders containing chitosan nanoparticles for pulmonary delivery of isoniazid

The aim of this study was to prepare spray dried inhalable powders containing isoniazid-loaded chitosan/tripolyphosphate (TPP) nanoparticles for sustained delivery of the drug to the lung. Nanoparticles were prepared by ionic gelation method. In-vitro drug release study indicated that the rate of drug release from nanoparticles was decreased by increasing the amount of chitosan. Entrapment of isoniazid into chitosan/TPP nanoparticles decreased minimum inhibitory concentrations (MIC) of the drug against mycobacterium avium intracellulare. Nanoparticles were spray dried using excipients such as lactose, mannitol and maltodextrin alone or with leucine. Results showed that the obtained powders had different aerosolization property. It was observed that by adding leucine, the particle size of microparticles deceased and the process yield and fine particle fraction (FPF) increased significantly. The in-vitro deposition data indicated that spray drying of isoniazid-loaded nanoparticles with lactose in the presence of leucine resulted in the production of inhalable powders with the highest FPF (45%).     

Comparison of particle sizing techniques in the case of inhalation dry powders.

The objectives of this work were (i) to validate electrical zone sensing and laser diffraction for the analysis of primary particle size in the case of inhalation dry powders and (ii) to study the influence of the aggregation state of the powder on the sizing techniques. Free-flowing dry powders were prepared by spray-drying with a combination of albumin, lactose, and dipalmitoylphosphatidylcholine. The replacement of lactose by mannitol, the removal of albumin, and the atomization at high relative humidity all increased powder cohesion. Automated measurements were compared with primary particle sizes collected by light and electron microscopy. The mass mode obtained by electrical zone sensing and the mass median diameter measured by laser diffraction following dispersion with compressed air at a pressure of 3 bar or following suspension in water and ultrasonic dispersion at a power of 60 W for 30 s each provided primary particle sizes close to microscopy measurements. However, these conditions only applied in the case of slightly to moderately aggregated powders. For strongly agglomerated powders, an exact measurement of the size was only collected by laser diffraction in the wet state combined with ultrasonic dispersion. Our study underlies how measurement of primary particle size highly depends on both powder material and proper particle dispersion.     

Improved lung delivery from a passive dry powder inhaler using an Engineered PulmoSphere powder

Purpose. To assess the pulmonary deposition and pharmacokinetics of an engineered PulmoSphere® powder relative to standard micronized drug when delivered from passive dry powder inhalers (DPIs). Methods. Budesonide PulmoSphere (PS_bud) powder was manufactured using an emulsion-based spray-drying process. Eight healthy subjects completed 3 treatments in crossover fashion: 370 g budesonide PulmoSphere inhaled from Eclipse® DPI at target PIF of 25 L·min^-1 (PS_bud25), and 50 L·min^-1 (PS_bud50), and 800 g of pelletized budesonide from Pulmicort® Turbuhaler® at 60 L·min^-1(TH_bud60). PS_bud powder was radiolabeled with ^99mTc and lung deposition determined scintigraphically. Plasma budesonide concentrations were measured for 12 h after inhalation. Results. Pulmonary deposition (mean ± sd) of PS_bud was 57 ± 7% and 58 ± 8% of the nominal dose at 25 and 50 L·min^-1, respectively. Mean peak plasma budesonide levels were 4.7 (PS_bud25), 4.0 (PS_bud50), and 2.2 ng·ml^-1 (TH_bud60). Median t_max was 5 min after both PS_bud inhalations compared to 20 min for Turbuhaler (P < 0.05). Mean AUCs were comparable after all inhalations, 5.1 (PS_bud25), 5.9 (PS_bud50), and 6.0 (TH_bud60) ng·h·ml^-1. The engineered PS_bud powder delivered at both flow rates from the Eclipse® DPI was twice as efficiently deposited as pelletized budesonide delivered at 60 L·min^-1 from the Turbuhaler. Intersubject variability was also dramatically decreased for PS_bud relative to TH_bud. Conclusion. Delivery of an engineered PulmoSphere formulation is more efficient and reproducible than delivery of micronized drug from passive DPIs.