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Spray freeze drying for protein encapsulation: Impact of the formulation to morphology and stability

Alberto Baldelli, Yigong Guo, Hui Xin Ong, Aylin Cidem, Anika Singh, Daniela Traini, Anubhav Pratap-Singh

Abstract

Proteins, the building blocks of life, are increasingly being used as therapeutics for treating several diseases. Yet, there are challenges in the delivery of highly labile materials like proteins, which is often circumvented with the help of encapsulation for targeted delivery and enhanced stability. Spray drying technology has recently been employed for encapsulation due to its’ low cost and scale-up capabilities, yet the high temperatures of drying air makes the technology unsuitable for proteins. More recently, spray freeze drying has evolved as an emerging technology that combines spray drying with freeze drying by using low temperatures, and is thus suitable for maintaining the stability of proteins. This study investigates the correlation between formulation parameters and the properties of protein encapsulated microparticles prepared by spray freeze drying. Morphology was investigated using microscopic methods, and protein stability was examined using infrared and mass spectrometry. By using bovine serum albumin, we verify that increasing the total weight to 15 mg/ml results in microencapsulates with a projected area equivalent diameter of 100 µm larger. We demonstrate that some types of amino acids are essential for shell formation; however, glutamine generates an increase in dimer areas in mass spectra of 5.5. D-Mannitol is the suggested carrier for high encapsulation efficiency (above 90 %). The formulation containing polyvinylpyrrolidone, mannitol, and leucine (at 6, 9, and 2 mg/ml, respectively) produced the lowest reduction in the stability of a few types of proteins; deconvoluted infrared peaks show a difference of less than 2% compared to the free protein. Understanding the spray freeze drying phenomenon for protein encapsulation would allow the control over morphological and chemical properties of microparticles containing active proteins.

Keywords

Spray freeze drying, encapsulation, proteins, particle formation

Modulating the Pore Architecture of Ice-Templated Dextran Microparticles Using Molecular Weight and Concentration

Tero Kämäräinen, Kazunori Kadota, Jun Yee Tse, Hiromasa Uchiyama, Shinya Yamanaka, Yuichi Tozuka

Abstract

Spray freeze drying (SFD) is an ice templating method used to produce highly porous particles with complex pore architectures governed by ice nucleation and growth. SFD particles have been advanced as drug carrier systems, but the quantitative description of the morphology formation in the SFD process is still challenging. Here, the pore space dimensions of SFD particles prepared from aqueous dextran solutions of varying molecular weights (40-200 kDa) and concentrations (5-20%) are analyzed using scanning electron microscopy. Coexisting morphologies composed of cellular and dendritic motifs are obtained, which are attributed to variations in the ice growth mechanism determined by the SFD system and modulation of these mechanisms by given precursor solution properties leading to changes in their pore dimensions. Particles with low-aspect ratio cellular pores showing variation of around 0.5-1 μm in diameter with precursor composition but roughly constant with particle diameter are ascribed to a rapid growth regime with high nucleation site density. Image analysis suggests that the pore volume decreases with dextran solid content. Dendritic pores (≈2-20 μm in diameter) with often a central cellular region are identified with surface nucleation and growth followed by a slower growth regime, leading to the overall dendrite surface area scaling approximately linearly with the particle diameter. The dendrite lamellar spacing depends on the concentration according to an inverse power law but is not significantly influenced by molecular weight. Particles with highly elongated cellular pores without lamellar formation show intermediate pore dimensions between the above two limiting morphological types. Analysis of variance and post hoc tests indicate that dextran concentration is the most significant factor in affecting the pore dimensions. The SFD dextran particles herein described could find use in pulmonary drug delivery due to their high porosity and biocompatibility of the matrix material.

Spray freeze-drying for inhalation application: process and formulation variables

Mostafa Rostamnezhad, Hossein Jafari, Farzad Moradikhah, Sara Bahrainian, Homa Faghihi, Reza Khalvati, Reza Bafkary, Alireza Vatanara

Abstract

High porous particles with specific aerodynamic properties were processed by the spray freeze-drying (SFD) method. Comprehensive knowledge about all aspects of the SFD method is required for particle engineering of various pharmaceutical products with good flow properties. In this review, different types of the SFD method, the most frequently employed excipients, properties of particles prepared by this method, and most recent approaches concerning SFD are summarized. Generally, this technique can prepare spherical-shaped particles with a highly porous interior structure, responsible for the very low density of powders. Increasing the solubility of spray freeze-dried formulations achieves the desired efficacy. Also, due to the high efficiency of SFD, by determining the different features of this method and optimizing the process by model-based studies, desirable results for various inhaled products can be achieved and significant progress can be made in the field of pulmonary drug delivery.

Keywords

Spray freeze-drying, Aerodynamic properties, Particle engineering, Pulmonary drug delivery, Inhaled products

Fabricating high-loading ultra-small PtCu3/rGO via a traceless protectant and spray-freeze-drying method

Qingyu Luo, Wei Xu, Shaolong Tang

Abstract

Exploring excellent catalysts for oxygen reduction reaction (ORR) with a facile and cost-effective method is desirable but remains challenging. Herein, ultra-small PtCu₃ nanoparticles (ca. 2.7 nm), immobilized on reduced graphene oxide (rGO), were synthesized via a novel and general strategy. Traceless protectant, NH₄OH, was used to resist the aggregation of graphene oxide (GO), and the spray-freeze-drying method ensures excellent dispersion of the Pt and Cu precursors, which could not be achieved by other reported drying methods. After annealing, the nanoparticles with the highest mass loading, 52%, among reported ordered Pt-based catalysts were obtained. The PtCu₃/rGO shows a remarkable electrocatalytic performance. Density functional theory calculations elucidate that PtCu₃ possess a lowered energy barrier of the rate-determining step, contributing to significantly improved ORR kinetics. This strategy was extended to the synthesis of other binary- and quaternary-metallic Pt-based nanoparticles, which proved its generality and applicability towards the potential commercialization of fuel cell technologies.

Keywords

Spray-freeze-drying, Traceless protectant, High-loading, Ultra-small, Oxygen reduction reaction

Designing highly porous amorphous celecoxib particles by spray freeze drying leads to accelerated drug absorption in-vivo

Dominic Lucas, Jan Kožák, Annika Rautenberg, Claire Chrétien, Yann Pellequer, Alf Lamprecht

Abstract

Poorly water-soluble drugs are still a major challenge to overcome in order to achieve sufficiently high oral bioavailability. Spray freeze drying (SFD) is proposed here as an alternative for the preparation of amorphous, free-flowing porous celecoxib spheres for enhanced drug dissolution. Tertiary butyl alcohol solutions of celecoxib + excipient (povidone, hydroxypropyl methylcellulose acetate succinate (HPMC-AS) and Soluplus®) at variable ratios were sprayed into a cooled spray tower, followed by vacuum freeze drying. Final porous particles were free-flowing, highly spherical (circularity ≥ 0.96) and mean diameters ranging from 210 to 800 µm, depending on excipient and drug content. XRPD measurements showed that Celecoxib was amorphous in all formulations and remained stable during 6 months storage. Kollidon 25 and HPMC-AS combinations resulted in the highest dissolution rates as well as dissolved drug amounts (30.4 ± 1.5 µg/ml and 41.8 ± 1.7 µg/ml) which in turn was 2-fold and 1.3-fold increase compared to film casted amorphous reference formulations, respectively. This phenomenon also translated into a faster onset of the drug absorption in-vivo, with significantly lower tmax values, while AUC values were non-significantly lowered compared to amorphous references. The high porosity of SFDs led to the advantageous accelerated dissolution which also translated into faster onset of absorption in-vivo.

Keywords

Spray freeze drying, Amorphous solid dispersions, Lyospheres, Porous particles, Poorly water-soluble

Dual targeting powder formulation of antiviral agent for customizable nasal and lung deposition profile through single intranasal administration

Han Cong Seow, Qiuying Liao, Andy T Y Lau, Susan W S Leung, Shuofeng Yuan, Jenny K W Lam

Abstract

Unpredictable outbreaks due to respiratory viral infections emphasize the need for new drug delivery strategies to the entire respiratory tract. As viral attack is not limited to a specific anatomic region, antiviral therapy that targets both the upper and lower respiratory tract would be most effective. This study aimed to formulate tamibarotene, a retinoid derivative previously reported to display broad-spectrum antiviral activity against influenza and severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), as a novel dual particle size powder formulation that targets both the nasal cavity and the lung by a single route of intranasal administration. Spray freeze drying (SFD) and spray drying (SD) techniques were employed to prepare tamibarotene powder formulations, and cyclodextrin was used as the sole excipient to enhance drug solubility. With the employment of appropriate atomizing nozzles, particles of size above 10 μm and below 5 μm could be produced for nasal and lung deposition, respectively. The aerosol performance of the powder was evaluated using Next Generation Impactor (NGI) coupled with a glass expansion chamber and the powder was dispersed with a nasal powder device. By blending powder of two different particle sizes, a single powder formulation with dual aerosol deposition characteristic in both the nasal and pulmonary regions was produced. The aerosol deposition fractions in the nasal cavity and pulmonary region could be modulated by varying the powder mixing ratio. All dry powder formulations exhibited spherical structures, amorphous characteristics and improved dissolution profile as compared to the unformulated tamibarotene. Overall, a novel dual targeting powder formulation of tamibarotene exhibiting customizable aerosol deposition profile was developed. This exceptional formulation strategy can be adopted to deliver other antimicrobial agents to the upper and lower airways for the prevention and treatment of human respiratory infections.

Keywords

Antiviral agent; COVID-19; Dry powder aerosol; Intranasal delivery; Pulmonary delivery; Spray drying; Spray freeze drying

Encapsulation of probiotics: past, present and future

R. Rajam & Parthasarathi Subramanian

Abstract – Background

Probiotics are live microbial supplements known for its health benefits. Consumption of probiotics reported to improve several health benefits including intestinal flora composition, resistance against pathogens. In the recent years, there is an increasing trend of probiotic-based food products in the market.

Keywords

Probiotics, Viability, Encapsulation, Spray drying, Digestion, Storage, Denaturation

PRELIMINARY STUDY OF INSULIN DRY POWDER FORMULATION: CRITICAL PROCESS PARAMETERS ON SPRAY-FREEZE-DRYING AND CRITICAL MATERIAL ATTRIBUTES OF TREHALOSE AND INULIN AS STABILIZER

CYNTHIA MARISCA MUNTU, SILVIA SURINI, CHRISTINA AVANTI, HAYUN, WOUTER HINRICHS

Abstract

Objective: The aim of this study was to obtain recommendations about critical process parameters (CPP) and optimal ratio of trehalose and inulin as critical material attribute (CMA) on insulin dry powder formulation with spray-freeze-drying (SFD) method. Methods: Inulin dry powder was formulated with the SFD method, which consisted of an atomization process and freeze-drying (FD). SFD processes were optimized in order to obtain dry powder and CPP was analyzed. All seven variations of formulas proceeded with physicochemical characterization to obtain the optimal formula. Results: In the early optimization, there was a slight time lag between the atomization process and FD; as a result, some of the powder coagulated and crystallized. Another critical parameter was that the FD process should not be interrupted for at least 50 h of FD. Dry powder proceeded with physicochemical characterization, a formula without inulin showed semicrystalline properties, while six formulas had amorphous properties due to its combination. All formulas had a spherulite shape and rough surface. Five formulas with the combination of trehalose and inulin obtained dry powders with a diameter range of 30-43 μm, moisture content below 3.5% and high encapsulation efficiency (EE). Formula with the ratio of 1:1 (F4) showed optimal properties with moisture content and EE of 2.62% and 99.68%, respectively. Conclusion: This study concluded that there were two critical process parameters in the SFD method. There should be no time lag in SFD process and FD time which should not be interrupted. The optimal ratio for trehalose and inulin was shown by F4 with ratio of 1:1.

Keywords

Process Parameters, Freeze Drying, Time Lag, Physicochemical Characterization, Dry Powder, Critical Material, Atomization Process, Critical Process Parameters, Spray Freeze Drying, Critical Process

Emerging Technologies and Coating Materials for Improved Probiotication in Food Products: a Review

Sourav Misra, Pooja Pandey, Chandrakant Genu Dalbhagat, Hari Niwas Mishra

Abstract

From the past few decades, consumers’ demand for probiotic-based functional and healthy food products is rising exponentially. Encapsulation is an emerging field to protect probiotics from unfavorable conditions and to deliver probiotics at the target place while maintaining the controlled release in the colon. Probiotics have been encapsulated for decades using different encapsulation methods to maintain their viability during processing, storage, and digestion and to give health benefits. This review focuses on novel microencapsulation techniques of probiotic bacteria including vacuum drying, microwave drying, spray freeze drying, fluidized bed drying, impinging aerosol technology, hybridization system, ultrasonication with their recent advancement, and characteristics of the commonly used polymers have been briefly discussed. Other than novel techniques, characterization of microcapsules along with their mechanism of release and stability have shown great interest recently in developing novel functional food products with synergetic effects, especially in COVID-19 outbreak. A thorough discussion of novel processing technologies and applications in food products with the incorporation of recent research works is the novelty and highlight of this review paper.

Keywords

Drying; Functional foods; Microencapsulation; Packaging condition; Probiotics; Storage

Complete Protection Against Yersinia pestis in BALB/c Mouse Model Elicited by Immunization With Inhalable Formulations of rF1-V10 Fusion Protein via Aerosolized Intratracheal Inoculation

Wei Zhang, Xiaolin Song, Lina Zhai, Jianshu Guo, Xinying Zheng, Lili Zhang, Meng Lv, Lingfei Hu, Dongsheng Zhou, Xiaolu Xiong and Wenhui Yang

Abstract

Pneumonic plague, caused by Yersinia pestis, is an infectious disease with high mortality rates unless treated early with antibiotics. Currently, no FDA-approved vaccine against plague is available for human use. The capsular antigen F1, the low-calcium-response V antigen (LcrV), and the recombinant fusion protein (rF1-LcrV) of Y. pestis are leading subunit vaccine candidates under intense investigation; however, the inability of recombinant antigens to provide complete protection against pneumonic plague in animal models remains a significant concern. In this study, we compared immunoprotection against pneumonic plague provided by rF1, rV10 (a truncation of LcrV), and rF1-V10, and vaccinations delivered via aerosolized intratracheal (i.t.) inoculation or subcutaneous (s.c.) injection. We further considered three vaccine formulations: conventional liquid, dry powder produced by spray freeze drying, or dry powder reconstituted in PBS. The main findings are: (i) rF1-V10 immunization with any formulation via i.t. or s.c. routes conferred 100% protection against Y. pestis i.t. infection; (ii) rF1 or rV10 immunization using i.t. delivery provided significantly stronger protection than rF1 or rV10 immunization via s.c. delivery; and (iii) powder formulations of subunit vaccines induced immune responses and provided protection equivalent to those elicited by unprocessed liquid formulations of vaccines. Our data indicate that immunization with a powder formulation of rF1-V10 vaccines via an i.t. route may be a promising vaccination strategy for providing protective immunity against pneumonic plague.

Keywords

Yersinia pestis, pneumonic plague, subunit vaccine, rF1-V10, dry powder formulation, aerosolized intratracheal inoculation, mucosal immune response