Inhalable, Spray-Dried Terbinafine Microparticles for Management of Pulmonary Fungal Infections: Optimization of the Excipient Composition and Selection of an Inhalation Device

Terbinafine is a broad-spectrum antifungal agent with therapeutic potential against pulmonary aspergillosis. The main aim of the current study was to investigate the potential of l-leucine, alone and in combination with mannitol, to improve the performance of spray-dried terbinafine microparticles for inhalation. The study also aimed to investigate the potential of the low resistance Cyclohaler® and the high resistance Handihaler® as inhalation devices for spray-dried microparticles. To this end, eight powder inhalation formulations of terbinafine were prepared by nano spray drying via a factorial experimental design. The formulations were evaluated in vitro for their potential to deliver the antifungal drug to the lungs using the Cyclohaler® and the Handihaler®. Leucine was superior as an excipient to mannitol and to mixtures of leucine and mannitol. Using leucine as an excipient resulted in formulations with fine particle fractions of up to 60.84 ± 0.67% w/w and particle mass median aerodynamic diameters of down to 1.90 ± 0.20 μm, whereas using mannitol as an excipient resulted in formulations with fine particle fractions of up to 18.75 ± 3.46% w/w and particle mass median aerodynamic diameters of down to 6.79 ± 0.82 μm. When leucine was used as an excipient, using 50% w/w rather than 25% w/w ethanol in water as a spray solvent enhanced the dispersibility of the particles, with a mean absolute increase in the formulation fine particle fraction of 9.57% w/w (95% confidence interval = 6.40–12.73% w/w). This was potentially underlain by enrichment of the particle surfaces with leucine. The Cyclohaler® outperformed the Handihaler® as an inhalation device for the developed formulations, with a mean absolute increase in the fine particle fraction of 9.17% w/w (95% confidence interval = 8.17–10.16% w/w).

Systemic Delivery of hGhrelin Derivative by Lyophilizate for Dry Powder Inhalation System in Monkeys

Ghrelin is the peptide that increases the hunger sensation and food intake and is expected to be clinically applied for treatment of diseases such as cachexia and anorexia nervosa. In the clinical application of ghrelin, injections are problematic in that they are invasive and inconvenient. Thus, we aimed to develop a formulation that can eliminate the need for injections and can be applied clinically. We prepared formulations of an hGhrelin derivative, in which the octanoyl group essential for expression of activity is modified to avoid rapid des-acylation, using lyophilizate for a dry powder inhalation (LDPI) system. The formulation of hGhrelin derivative was optimized by the addition of phenylalanine, of which the fine particle fraction of 5 µm or less was 41.7 ± 3.8%. We also performed pharmacokinetic/pharmacodynamic tests in monkeys using the optimum formulation that can be applied clinically. The absolute bioavailability of inhaled hGhrelin derivative with respect to that intravenously injected was 16.9 ± 2.6%. An increase in growth hormone was shown as an effect of the inhaled hGhrelin derivative similar to intravenous injection. The LDPI formulation can deliver the hGhrelin derivative systemically, and it is expected to be applied clinically as a substitute for injections.

Providing calves different feed source early in life would not affect feed sorting and rumen fermentation for a long term

This study aimed to investigate the short and long-term effect of early in life exposure to different feed sources on feed sorting and rumen fermentation of calves. Forty newborn female Holstein calves were randomly divided based on supplementation of concentrate (CON) or hay (HAY) only during the milk-feeding stage (d 1 to 56). After that, all calves were offered a total mixed ration (TMR) containing 43 % of forage and 57 % of concentrate until the end of the experiment (d 57 to 196). Rectal temperature, feed intake, body weight, body structural measurement, fecal and cough score were recorded ranges from day 1 to 196. Fresh and orts feed were sampled daily at the first two weeks (d 57-70) once TMR was offered and at the last week of the experiment (d 190 to 196) for analysis of feed sorting. Rumen fluid samples were collected at ranges from d 14-190 for detecting rumen pH and volatile fatty acids (VFA) concentrations. Our findings showed that early feed exposure did not influence calf health and growth performance but affected feed sorting in a short period. Upon transition to the same mixed diet, differences were found between treatments in the sorting of the different particle fractions. At week 9 and 10, calves early exposed to concentrates began to sort for fine particle fraction, and against the long particle fraction, whereas calves early fed hay sorted for the long particle fraction, and against the fine particle fraction. However, no carryover effect was found, the differences between treatments fade away when detecting feed sorting at the last week, all calves sorted for short and fine particle fractions, and sort against long and medium particle fractions. Although the pattern of feed sorting initially differed between treatments, the rumen pH and VFA concentration were not altered once all calves transitioned to a TMR. These results suggested that the feed familiarity established early in life would not affect diet selection and rumen fermentation in heifer later in life, the dietary experience after transition to same TMR would eventually override the effect of early feed experience.

Providing calves different feed source early in life would not affect feed sorting and rumen fermentation for a long term

This study aimed to investigate the short and long-term effect of early in life exposure to different feed sources on feed sorting and rumen fermentation of calves. Forty newborn female Holstein calves were randomly divided based on supplementation of concentrate (CON) or hay (HAY) only during the milk-feeding stage (d 1 to 56). After that, all calves were offered a total mixed ration (TMR) containing 43% of forage and 57% of concentrate until the end of the experiment (d 57 to 196). Calves were weighed immediately after birth and raised in individual hutches until d 70. The calves were then transferred to a heifer barn, housed within treatment in pairs (2 calves/pen). Rectal temperature, feed intake, body weight, body structural measurement, fecal and cough score were recorded ranges from day 1 to 196. Fresh and orts feed were sampled daily at the first two weeks (d 57-70) once TMR was offered and at the last week of the experiment (d 190 to 196) for analysis of feed sorting. Rumen fluid samples were collected at ranges from d 14-190 for detecting rumen pH and volatile fatty acids (VFA) concentrations. Our findings showed that early feed exposure did not influence calf health and growth performance but affected feed sorting in a short period. Upon transition to the same mixed diet, differences were found between treatments in the sorting of the different particle fractions. At week 9 and 10, calves early exposed to concentrates began to sort for fine particle fraction (primarily concentrate in this fraction), and against the long particle fraction (solely forage in this fraction), whereas calves early fed hay sorted for the long particle fraction, and against the fine particle fraction. However, no carryover effect was found, the differences between treatments fade away when detecting feed sorting at the last week, all calves sorted for short and fine particle fractions, and sort against long and medium particle fractions. Although the pattern of feed sorting initially differed between treatments, the rumen pH and VFA concentration were not altered once all calves transitioned to a TMR. These results suggested that the feed familiarity established early in life would not affect diet selection and rumen fermentation in heifer later in life, the dietary experience after transition to same TMR would eventually override the effect of early feed experience.

Inhalable Dry Powder of Bedaquiline for Pulmonary Tuberculosis: In Vitro Physicochemical Characterization, Antimicrobial Activity and Safety Studies

Bedaquiline is a newly developed anti-tuberculosis drug, conditionally approved by the United States Food and Drug Administration (USFDA) for treating drug-resistant tuberculosis in adults. Oral delivery of bedaquiline causes severe side effects such as increased hepatic aminotransferase levels and cardiac arrhythmias (prolongation of QT-interval). This study aimed to develop inhalable dry powder particles of bedaquiline with high aerosolization efficiency to reduce the side-effects of oral bedaquiline. Bedaquiline (with or without l-leucine) powders were prepared using a Buchi Mini Spray-dryer. The powders were characterized for physicochemical properties and for their in vitro aerosolization efficiency using a next-generation impactor (NGI). The formulation with maximum aerosolization efficiency was investigated for physicochemical and aerosolization stability after one-month storage at 20 ± 2 °C/30 ± 2% relative humidity (RH) and 25 ± 2 °C/75% RH in an open Petri dish. The cytotoxicity of the powders on A549 and Calu-3 cell-lines was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The powders were also evaluated for antimicrobial activity against Mycobacterium tuberculosis. The aerodynamic diameter of the l-leucine-containing powder was 2.4 µm, and the powder was amorphous in nature. The aerosolization efficiency (fine-particle fraction) of l-leucine-containing powder (fine-particle fraction (FPF): 74.4%) was higher than the bedaquiline-only powder (FPF: 31.3%). l-leucine containing powder particles were plate-shaped with rough surfaces, but the bedaquiline-only powder was spherical and smooth. The optimized powder was stable at both storage conditions during one-month storage and non-toxic (up to 50 µg/mL) to the respiratory cell-lines. Bedaquiline powders were effective against Mycobacterium tuberculosis and had a minimal inhibitory concentration (MIC) value of 0.1 µg/mL. Improved aerosolization may help to combat pulmonary tuberculosis by potentially reducing the side-effects of oral bedaquiline. Further research is required to understand the safety of the optimized inhalable powder in animal models.

Aerosolization Performance of Jet Nebulizers and Biopharmaceutical Aspects

In this work, 13 jet nebulizers, some of which in different configurations, were investigated in order to identify the biopharmaceutical constraints related to the quality attributes of the medicinal products, which affect their safety, efficiency, compliance, and effectiveness. The aerosolization parameters, including the aerosol output, aerosol output rate, mass median aerodynamic diameter, and fine particle fraction, were determined according to the European Standard EN 13544-1, using sodium fluoride as a reference formulation. A comparison between the aerosol output nebulization time and the fine particle fraction displayed a correlation between the aerosol quality and the nebulization rate. Indeed, the quality of the nebulization significantly increased when the rate of aerosol emission was reduced. Moreover, the performance of the nebulizers was analyzed in terms of respirable delivered dose and respirable dose delivery rate, which characterize nebulization as the rate and amount of respirable product that could be deposited into the lungs. Depending on which of these two latter parameters was used, the nebulizers showed different performances. The differences, in terms of the rate and amount of delivered aerosol, could provide relevant information for the appropriate choice of nebulizer as a function of drug product, therapy, and patient characteristics.