Evaluation of an on-farm culture system for the detection of subclinical mastitis pathogens in dairy cattle.

The purpose of this observational study was to compare the performance of a novel on-farm culture (OFC) test with the reference method (RM) in identifying pathogens, and in particular Staphylococcus aureus, associated with subclinical mastitis (SCM) in dairy cattle. The OFC test (Mastatest HiSCC; Mastaplex Limited) for SCM uses a cartridge with 2 × 12 wells allowing 1 sample to be analyzed in duplicate (24 wells) or 2 samples analyzed simultaneously, each in 12 wells. Results of the milk analyses are reported hierarchically (Staph. aureus → coagulase-negative staphylococci (CNS) → other gram positive or coliform/gram negative → no bacteria present) and emailed within 24 h. Milk samples (617 quarter level from 158 cows and 70 cow level) were collected from 288 cows [individual cow somatic cell count (ICSCC) ≥150,000 cells/mL] on 9 purposefully selected farms known to have a high prevalence of clinical and subclinical Staph. aureus mastitis in Southland New Zealand. Quarter samples were analyzed individually (617 samples) and after animal-level pooling, providing 228 (158 + 70) cow-level samples. Samples were analyzed by the OFC test (in duplicate) and the RM (culture agar medium and latex test based on the recommendation by the National Mastitis Council) and identifications confirmed with MALDI-TOF mass spectrometry. Sensitivity (Se), specificity (Sp), positive predictive value (PPV), and negative predictive value (NPV) for detection of Staph. aureus were all ∼90% with tight 95% confidence limits, and Cohen's kappa (κ) for agreement between the OFC test and RM was 0.81. Kappa for agreement between the OFC test duplicates was 0.93. About 35% of cows had only one quarter infected with Staph. aureus and all these animals could still be identified when pooled cow-level milk was analyzed. Although the high prevalence of Staph. aureus in the herds used in this study does not affect the Se and Sp values, it does elevate the PPV value (and decrease the NPV) and therefore use of PPV to extrapolate to a population with lower prevalence is not appropriate. For CNS, Sp, PPV, and NPV were all >0.8, κ was ≥0.6, and Se was >0.7. Kappa for agreement between the OFC test duplicates was 0.83. A result of "no bacteria detected" was reported in 13% of the cows with 93% agreement between OFC test and RM. We conclude that the OFC test provides a reliable method for detecting Staph. aureus in pooled cow-level milk even if only one quarter is infected; in the absence of Staph. aureus in the milk, it reliably identified CNS in pooled cow-level milk; it reliably identified cows with <10 cfu/10 µL of their milk. Compared with the RM, the method was rapid with results returned in 24 h of loading the cartridge.

In vitro Dissolution Testing of Rifampicin Powder Formulations For Prediction of Plasma Concentration-Time Profiles After Inhaled Delivery.

PURPOSE: The purpose of this study was to evaluate the in vitro lung dissolution of amorphous and crystalline powder formulations of rifampicin in polyethylene oxide (PEO) and 1,2-dipalmitoyl-sn-glycero-3-phosphatidylcholine (DPPC), and to predict the in vivo plasma concentration-time profiles using the in vitro data. METHODS: The in vitro dissolution and permeation profiles of respirable rifampicin particles were studied using a custom-made dissolution apparatus. Data from the in vitro dissolution test were used to estimate the parameters to be used as the input for the simulation of in vivo plasma concentration-time profiles using STELLA® software. For prediction of in vivo profiles, a one-compartment model either with a first order elimination or with a Michaelis-Menten kinetics-based elimination was used. RESULTS: Compared to the crystalline formulation, the amorphous formulation showed rapid in vitro dissolution suggesting their possible faster in vivo absorption and higher plasma concentrations of rifampicin following lung delivery. However, the simulations suggested that both powder formulations would result in similar plasma-concentration time profiles of rifampicin. CONCLUSIONS: Use of an in vitro dissolution test coupled with a simulation model for prediction of plasma-concentration time profiles of an inhaled drug was demonstrated in this work. These models can also be used in the design of inhaled formulations by controlling their release and dissolution properties to achieve desired lung retention or systemic absorption following delivery to the lungs.

Modeling the interaction of polymeric nanoparticles functionalized with cell penetrating peptides at the nano-bio interface.

The interaction of nanoparticles with Caco-2 monolayers in cell culture underpins our predictions of the uptake of nanoformulations in vivo for drug delivery. Cell-penetrating peptides (CPP), such as oligoarginine, are currently of interest to enhance cellular uptake of bioactives and nanoparticles. This paper assesses the cellular association of poly(ethyl-cyanoacrylate) nanoparticles functionalized with di-arginine-histidine (RRH) in a Caco-2 cell model. We applied a computational model of particokinetics, In vitro Sedimentation, Diffusion and Dosimetry (ISDD) to predict the accumulation of nanoparticles on the cell surface. An important finding is that the proportion of nanoparticles associated with cells was less than 5 %. This has important implications for interpreting nanoparticle uptake in vitro. RRH-decoration does not appear to alter nanoparticle deposition, but increases association of nanoparticles with Caco-2 cells. Immediate deposition of nanoparticles on the cell surface was apparent and similar between formulations, but underestimated by the ISDD model. Key to understanding the nano-bio interface for drug delivery, nanoparticles that reach the cells were not necessarily absorbed by them, but can become detached. This variable of nanoparticle release from cells was incorporated into a new mathematical model presented here.

Pharmacokinetics of rifampicin after repeated intra-tracheal administration of amorphous and crystalline powder formulations to Sprague Dawley rats.

Rifampicin is one of the key drugs used to treat tuberculosis and is currently used orally. The use of higher oral doses of rifampicin is desired for better therapeutic efficacy, but this is accompanied by increased risk of systemic toxicity thus limiting its recommended oral dose to 10 mg/kg per day. Inhaled delivery of rifampicin is a potential alternative mode of delivery, to achieve high drug concentrations in both the lung and potentially the systemic circulation. In addition, rifampicin exists either as amorphous or crystalline particles, which may show different pharmacokinetic behaviour. However, disposition behaviour of amorphous and crystalline rifampicin formulations after inhaled high-dose delivery is unknown. In this study, rifampicin pharmacokinetics after intra-tracheal administration of carrier-free, amorphous and crystalline powder formulations to Sprague Dawley rats were evaluated. The formulations were administered once daily for seven days by oral, intra-tracheal and oral plus intra-tracheal delivery, and the pharmacokinetics were studied on day 0 and day 6. Intra-tracheal administration of the amorphous formulation resulted in a higher area under the plasma concentration curve (AUC) compared to the crystalline formulation. For both formulations, the intra-tracheal delivery led to significantly higher AUC compared to the oral delivery at the same dose suggesting higher rifampicin bioavailability from the inhaled route. Increasing the intra-tracheal dose resulted in a more than dose proportional AUC suggesting non-linear pharmacokinetics of rifampicin from the inhaled route. Upon repeated administration for seven days, no significant decrease in the AUCs were observed suggesting the absence of rifampicin induced enzyme auto-induction in this study. The present study suggests an advantage of inhaled delivery of rifampicin in achieving higher drug bioavailability compared to the oral route.

Studies on the safety and the tissue distribution of inhaled high-dose amorphous and crystalline rifampicin in a rat model.

Inhaled delivery of rifampicin has the potential to achieve high drug concentrations in the lung and the blood for efficient treatment of tuberculosis (TB). Due to its existence as polymorphs, in vivo evaluation of the respiratory tract safety of inhalable amorphous and crystalline rifampicin particles, at clinically relevant high-dose, is necessary. This study investigates the lung and liver safety and the tissue distribution of rifampicin after intra-tracheal administration of high (≥25 mg/kg) doses of amorphous and crystalline powder formulations to Sprague Dawley rats. Powder formulations were administered by intra-tracheal insufflation to rats. Lung and liver safety were evaluated by histopathology. Serum alanine transaminase (ALT) and aspartate aminotransferase (AST) assays were performed to study the hepatic effects. Rifampicin was quantified in the tissues using LC-MS/MS. Intra-tracheal administration of rifampicin decreased the drug burden on the liver compared to oral administration based on its lower serum ALT activity. Repeated-dose intra-tracheal rifampicin was well tolerated by rats, confirmed by the absence of drug or delivery induced complexities. The histopathological evaluation of rat lungs, after both single and repeated drug administration for seven days, suggested the absence of drug-induced toxicity. Following single intra-tracheal delivery of 50 mg/kg doses, comparable rifampicin concentrations to that from same oral dose were observed in lung, liver, heart and brain. Inhaled delivery of high-dose rifampicin was safe to rat lungs and liver suggesting its potential for localized as well as systemic drug delivery without toxicity concerns.

Cell-based, animal and H1 receptor binding studies relative to the sedative effects of ketotifen and norketotifen atropisomers.

OBJECTIVES: Ketotifen (K) and its active metabolite norketotifen (N) exist as optically active atropisomers. They both have antihistaminic and anti-inflammatory properties but the S-atropisomer of N (SN) causes less sedation than K and RN in rodents. This study investigated whether this could be related to a lower concentration of SN in brain or a lower affinity of SN for rat brain H1 receptors. METHODS: Ketotifen and norketotifen atropisomers were quantified using a validated chiral HPLC assay. RBE4 and Caco-2 cell monolayers were used in uptake and permeability studies, respectively. Free and total brain-to-plasma (B/P) ratios were determined after injecting racemic K and N into rat tail veins. Affinity for rat brain H1 receptors (KI ) was determined using the [3 H]mepyramine binding assay. KEY FINDINGS: Uptake and permeation studies indicate no stereoselective transport for K or N. B/P ratios reveal the brain concentration of N is lower than K with no stereoselective transport into brain. Finally, the [3 H]mepyramine binding assay shows SN has the lowest affinity for rat brain H1 receptors. CONCLUSION: The lower sedative effect of SN in rodents is probably due to a combination of a lower uptake of N than K into the brain and less affinity of SN for CNS H1 receptors.

A STELLA simulation model for in vitro dissolution testing of respirable size particles.

In vitro dissolution testing is a useful quality control tool to discriminate the formulations and to approximate the in vivo drug release profiles. A dissolution apparatus has been custom-made for dissolution testing of dry powder formulations in a small volume of stationary medium (25 µL spread over 4.91 cm2 area i.e. ~50 µm thick). To understand the system and predict the key parameters which influence the dissolution of respirable size particles, a simulation model was constructed using STELLA modeling software. Using this model, the permeation (dissolution followed by diffusion through the membrane) of two anti-tubercular drugs of differing solubilities, moxifloxacin (17.68 ± 0.85 mg mL-1) and ethionamide (0.46 ± 0.02 mg mL-1), from the respirable size particles and their diffusion from a solution were simulated. The simulated permeation profiles of moxifloxacin from solution and respirable size particles were similar, indicating fast dissolution of the particles. However, the simulated permeation profile of ethionamide from respirable size particles showed slower permeation compared to the solution indicating the slow dissolution of the respirable size particles of ethionamide. The sensitivity analysis suggested that increased mucus volume and membrane thickness decreased the permeation of drug. While this model was useful in predicting and distinguishing the dissolution behaviours of respirable size moxifloxacin and ethionamide, further improvement could be made using appropriate initial parameter values obtained by experiments.

Crystalline adduct of moxifloxacin with trans-cinnamic acid to reduce the aqueous solubility and dissolution rate for improved residence time in the lungs.

A crystalline adduct of the anti-tubercular drug, moxifloxacin and trans-cinnamic acid (1:1 molar ratio (MCA1:1)) was prepared to prolong the residence time of the drug in the lungs by reducing its solubility and dissolution rate. Whether the adduct is a salt or cocrystal has not been unequivocally determined. Equilibrium solubility and intrinsic dissolution rate measurements for the adduct (MCA1:1) in phosphate buffered saline (PBS, pH 7.4) revealed a significant decrease in the solubility of moxifloxacin (from 17.68 ±â€¯0.85 mg mL-1 to 6.10 ±â€¯0.05 mg mL-1) and intrinsic dissolution rate (from 0.47 ±â€¯0.04 mg cm-2 min-1 to 0.14 ±â€¯0.03 mg cm-2 min-1) compared to the supplied moxifloxacin. The aerosolization behaviour of the adduct from an inhaler device, Aerolizer®, using a Next Generation Impactor showed a fine particle fraction of 30.4 ±â€¯1.2%. The dissolution behaviour of the fine particle dose of respirable particles collected was assessed in a small volume of stationary mucus fluid using a custom-made dissolution apparatus. The respirable adduct particles showed a lower dissolution (microscopic observation) and permeation compared to the supplied moxifloxacin. The crystalline adduct MCA1:1 has a lower solubility and dissolution rate than moxifloxacin and could improve the local residence time and therapeutic action of moxifloxacin in the lungs.

Aniracetam does not improve working memory in neurologically healthy pigeons.

Understanding the effects of cognitive enhancing drugs is an important area of research. Much of the research, however, has focused on restoring memory following some sort of disruption to the brain, such as damage or injections of scopolamine. Aniracetam is a positive AMPA-receptor modulator that has shown promise for improving memory under conditions when the brain has been damaged, but its effectiveness in improving memory in neurologically healthy subjects is unclear. The aim of the present study was to examine the effects of aniracetam (100mg/kg and 200 mg/kg) on short-term memory in "neurologically healthy" pigeons. Pigeons were administered aniracetam via either intramuscular injection or orally, either 30 or 60 minutes prior to testing on a delayed matching-to-sample task. Aniracetam had no effect on the pigeons' memory performance, nor did it affect response latency. These findings add to the growing evidence that, while effective at improving memory function in models of impaired memory, aniracetam has no effect in improving memory in healthy organisms.

The influence of storage relative humidity on aerosolization of co-spray dried powders of hygroscopic kanamycin with the hydrophobic drug rifampicin.

The purpose of this study was to investigate the influence of storage humidity on in vitro aerosolization and physicochemical properties of co-spray dried powders of kanamycin with rifampicin. The powders were stored for one-month in an open Petri dish at different relative humidities (RHs) (15%, 43%, and 75%) and 25 ± 2 °C. The in vitro aerosolization (fine particle fraction, FPF) of the powders was determined by a next generation impactor (NGI). The moisture content, particle morphology and crystallinity of the powders were determined by Karl Fischer titration, scanning electron microscopy, and X-ray powder diffractometry, respectively. At all RH, the FPF of hydrophobic rifampicin-only powder was unaffected but the FPF of hygroscopic kanamycin-only powder significantly decreased even at 43% RH. The kanamycin-only particles fused together, crystallized and formed hard cakes at 75% RH. The aerosolization of kanamycin and rifampicin in the combination powders remained unaffected at 15% and 43% RH, but aerosolization significantly decreased at 75% RH. Enrichment of the surface of the particles with hydrophobic rifampicin did not protect the combination powders from moisture uptake but it prevented particle agglomeration up to 43% RH. At 75% RH, the moisture uptake led to agglomeration of the particles of the combination powder particles and consequently an increase in aerodynamic diameter. Further studies are required to investigate how rifampicin enrichment prevents particle agglomeration, the possible mechanisms (e.g. particle interactions due to capillary forces or electrostatic forces) for the changes in the aerosolization and changes in surface composition during storage.

Carrier-free combination dry powder inhaler formulation of ethionamide and moxifloxacin for treating drug-resistant tuberculosis.

This study aimed to develop a combination dry powder formulation of ethionamide and moxifloxacin HCl as this combination is synergistic against drug-resistant Mycobacterium tuberculosis (Mtb). L-leucine (20% w/w) was added in the formulations to maximize the process yield. Moxifloxacin HCl and/or ethionamide powders with/without L-leucine were produced using a Buchi Mini Spray-dryer. A next generation impactor was used to determine the in vitro aerosolization efficiency. The powders were also characterized for other physicochemical properties and cytotoxicity. All the spray-dried powders were within the aerodynamic size range of <5.0 µm except ethionamide-only powder (6.0 µm). The combination powders with L-leucine aerosolized better (% fine particle fraction (FPF): 61.3 and 61.1 for ethionamide and moxifloxacin, respectively) than ethionamide-only (%FPF: 9.0) and moxifloxacin-only (%FPF: 30.8) powders. The combination powder particles were collapsed with wrinkled surfaces whereas moxifloxacin-only powders were spherical and smooth and ethionamide-only powders were angular-shaped flakes. The combination powders had low water content (<2.0%). All the powders were physically stable at 15% RH and 25 ± 2 °C during 1-month storage and tolerated by bronchial epithelial cell-lines up to 100 µg/ml. The improved aerosolization of the combination formulation may be helpful for the effective treatment of drug-resistant tuberculosis. Further studies are required to understand the mechanisms for improved aerosolization and test the synergistic activity of the combination powder.

In vitro dissolution testing of respirable size anti-tubercular drug particles using a small volume dissolution apparatus.

A dissolution apparatus that uses a small volume of stationary medium (25 µL) has been developed for in vitro dissolution testing of respirable drug particles and used to evaluate the dissolution of two anti-tubercular drugs, moxifloxacin and ethionamide. Solubilities of moxifloxacin and ethionamide in phosphate buffered saline (PBS, pH 7.4) were 17.68 ±â€¯0.85 mg mL-1 and 0.46 ±â€¯0.02 mg mL-1 whereas in the presence of lung surfactant (0.4% w/v Curosurf® in PBS) solubilities were 20.76 ±â€¯0.35 mg mL-1 and 0.56 ±â€¯0.03 mg mL-1, respectively. A fine particle dose (∼50 µg) of aerodynamically separated moxifloxacin or ethionamide particles (<6.4 µm) was collected onto a glass coverslip using a modified Twin Stage Impinger. The dissolution behaviour of the fine particle dose was evaluated at various perfusate flow rates (0.2, 0.4 and 0.8 mL min-1 of PBS), mucus simulant concentrations (1.0, 1.5 and 2.0% w/v polyethylene oxide in PBS), and in the presence of lung surfactant. The dissolution behaviour of the respirable size particles was observed under an optical microscope and the dissolved drug that diffused into the perfusate was quantified by HPLC. The moxifloxacin particles disappeared quickly and showed faster permeation (<30 min) compared to the ethionamide particles at all the dissolution conditions evaluated. This study demonstrated the differences in the dissolution rates of moxifloxacin and ethionamide particles and may be useful to estimate the residence time of the inhaled dry powder particles in the lungs.

Preadmission predictors of academic performance in a pharmacy program: A longitudinal, multi-cohort study.

INTRODUCTION: This study examined the extent to which a preadmission health science program and demographic variables predicted academic performance throughout an undergraduate pharmacy degree (BPharm) program. METHODS: A longitudinal, multi-cohort study was undertaken of 557 students admitted to the University of Otago School of Pharmacy BPharm program between 2008 and 2012, from a preceding health science year (HSFY). Preadmission baseline data including health science grade point average (GPA), sex, age, ethnicity, residency status, and high school qualifications were matched against outputs of GPA performances in all three years of the BPharm program using regression analyses. RESULTS: Five hundred thirty-eight students (96.6%) completed their BPharm degree. The regression models were significantly predictive of performance in the BPharm program with 57%, 43% and 38% of variances explained for GPA performance across years two, three and four, respectively (p < 0.001). Demographic variables including being male, being from certain minority ethnic groups or not having a specific domestic high school qualification were associated with lower GPA performances across the BPharm program compared to reference groups. DISCUSSION AND CONCLUSIONS: Determining admission from performance rankings as the single selection tool holds reasonable predictive value early in progression, however additional measures may be warranted to better predict performances extending beyond the first year of the BPharm program.

High dose dry powder inhalers to overcome the challenges of tuberculosis treatment.

Tuberculosis (TB) is a major global health burden. The emergence of the human immunodeficiency virus (HIV) epidemic and drug resistance has complicated global TB control. Pulmonary delivery of drugs using dry powder inhalers (DPI) is an emerging approach to treat TB. In comparison with the conventional pulmonary delivery for asthma and chronic obstructive pulmonary disease (COPD), TB requires high dose delivery to the lung. However, high dose delivery depends on the successful design of the inhaler device and the formulation of highly aerosolizable powders. Particle engineering techniques play an important role in the development of high dose dry powder formulations. This review focuses on the development of high dose dry powder formulations for TB treatment with background information on the challenges of the current treatment of TB and the potential for pulmonary delivery. Particle engineering techniques with a particular focus on the spray drying and a summary of the developed dry powder formulations using different techniques are also discussed.

Manipulation of spray-drying conditions to develop dry powder particles with surfaces enriched in hydrophobic material to achieve high aerosolization of a hygroscopic drug.

This study aimed to develop dry powder particles with surfaces enriched in hydrophobic material by manipulation of spray-drying conditions and to investigate the effect of hydrophobic surface enrichment on aerosolization of hygroscopic drug. The composite dry powder formulations of kanamycin (hygroscopic drug) and rifampicin (hydrophobic drug) were produced by systematically (23 full factorial design) varying the drug ratio, co-solvent composition and inlet temperature using Buchi B-290 Mini Spray-Dryer. All the composite powder particles were inhalable in size (3.1-3.9 µm), wrinkled, flake-shaped and amorphous. X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry showed that hydrophobic surface enrichment was significantly affected by co-solvent composition. Complete hydrophobic surface enrichment was achieved in one formulation (F7). The aerosolization efficiency by next generation impactor (NGI) showed that the composite formulations had higher fine particle fraction (FPF: >48.0%) than kanamycin-only formulation (FPF: 27.6%). Increase in hydrophobic surface enrichment (from 80.8 to 100%) decreased the powder density and increased FPF (from 48.0 to 77.2%). This is the first systematic study reporting the manipulation of spray-drying conditions for hydrophobic surface enrichment in composite dry powder particles and its effect on aerosolization. The high aerosolization efficiency of the combination formulations may be useful to deliver high doses of these drugs to treat lung infections.

Co-spray drying of hygroscopic kanamycin with the hydrophobic drug rifampicin to improve the aerosolization of kanamycin powder for treating respiratory infections.

High dose delivery of drugs to the lung using a dry powder inhaler (DPI) is an emerging approach to combat drug-resistant local infections. To achieve this, highly aerosolizable powders are required. We hypothesized that co-spray-drying kanamycin, a hydrophilic hygroscopic antibiotic, with rifampicin, a hydrophobic antibiotic, would produce inhalable particles with surfaces enriched in rifampicin. Such particles would have higher aerosolization than kanamycin alone, and minimise the mass of powder for inhalation avoiding use of non-active excipients. Kanamycin was co-spray-dried with rifampicin using a Buchi Mini Spray-dryer. All powders were inhalable in size (1.1-5.9 µm) and noncrystalline. X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (ToF-SIMS) showed the surface of the combination powder was enriched with rifampicin. In vitro aerosolization (fine particle fraction) determined by next generation impactor (NGI), dramatically improved from 29.5 ±â€¯0.2% (kanamycin-only) to 78.2 ±â€¯1.3% (kanamycin-rifampicin combination). The combination powder was flake-shaped in morphology, stable at 15% and 53% RH and 25 ±â€¯2 °C during one-month storage in an open Petri dish, and non-toxic (up to 50 µg/mL) to human alveolar and bronchial cell-lines. Surface enrichment of kanamycin by hydrophobic rifampicin improves aerosolization, which may help to combat drug-resistant local infections by facilitating high dose delivery to deep lung.

A real-time in vitro assay to evaluate the release of macromolecules from liposomes.

The availability of a real-time assay to experimentally investigate the release of encapsulated proteins would be beneficial given the interest in the use of liposomes as a drug delivery vehicle. Although simple assays for small molecular weight substances exist, assays to evaluate macromolecules do not. Here we describe a method that detects the release of model macromolecules from liposomes in real time. The assay employs the intermolecular distance-dependent phenomenon of fluorescence resonance energy transfer (FRET) between the fluorophore donor, fluorescein (FITC), and fluorescent quencher, QSY® 9. The macromolecular species were conjugated to the markers fluorescein (44kDa dextran) and QSY® 9 (67 kDa bovine serum albumin, BSA). Following confirmation of quenching between FITC-Dex and QSY® 9-BSA, liposomes were loaded with the macromolecular markers and subjected to various treatments (high-pressure extrusion and Triton X solubilisation) to cause release from liposomes. An increase in FITC fluorescence was observed when liposomes were subjected to extrusion cycles. Surprisingly, the addition of Triton X did not cause an increase in fluorescence probably because the FRET pair became associated with mixed micelles. This assay method should be useful in studies to investigate the mechanisms by which macromolecules are released from liposomes, particularly when liposomes are exposed to release-triggers (eg, temperature change, pH change, ultrasound). Such understanding will underpin the formulation of triggered liposomal delivery systems for macromolecules.

Dry powder formulation of kanamycin with enhanced aerosolization efficiency for drug-resistant tuberculosis.

BACKGROUND: Kanamycin, an injectable agent, is currently used to treat drug-resistant tuberculosis (TB). Parenteral kanamycin causes high systemic toxicity which could be avoided by direct delivery to the lungs. This study focused on producing a highly aerosolizable dry-powder of hygroscopic kanamycin by spray-drying with l-leucine. METHODS: Kanamycin powders were prepared with different concentrations (0, 5, 10, 15 and 20% w/w) of l-leucine using the Buchi B-290 Mini Spray-Dryer. In vitro aerosolization efficiency, particle size, morphology, crystallinity, surface composition, drug-excipient interaction and moisture content of the powders were characterized by a Next Generation Impactor (NGI), laser diffraction, scanning electron microscopy, X-ray diffractometry, XPS, ATR-FTIR and thermogravimetric analysis. The physicochemical and aerosolization stability of the powders were investigated after one-month storage at 25±2°C/15% RH and 25±2°C/75% RH. The cytotoxicity on Calu-3 and A549 cells of the kanamycin powders was evaluated by MTT assay. RESULTS: The spray-dried powder particles were in the inhalable size range (<6.1µm). The powders with l-leucine were wrinkled in shape, amorphous in nature and had low moisture content (<5.0%). Kanamycin with 5% (w/w) of l-leucine showed the best aerosolization efficiency of 73.0±2.5%. The powders remained stable during storage at 25±2°C/15% RH and tolerated by respiratory cell lines. CONCLUSION: l-leucine improved the aerosolization of kanamycin by surface modification, which may be helpful for the effective treatment of drug-resistant tuberculosis.

Pharmacokinetics of Abamectin/Levamisole Combination in a Medium Chain Mono and Diglyceride-Based Vehicle and an In Vitro Release and In Vitro In Vivo Correlation Study for Levamisole.

A combination of lipophilic and hydrophilic drugs in a single solution is a challenge due to their different physicochemical properties. In vitro and in vivo release studies are useful to optimize this solution. The in vitro (Franz diffusion cell) release rate of levamisole phosphate from an isotropic vehicle of medium chain mono and diglycerides (MCMDG) was significantly slower than the release from water. The injectable solution of the isotropic MCMDG-based system was prepared with 13.65% of levamisole phosphate and 0.5% of abamectin. Two milliliters/50 kg (0.04 ml/kg) was injected subcutaneously into five healthy adult sheep. None of the animals showed the signs of inflammation at injection site. Both drugs were assayed using validated HPLC methods. The absorption rates for levamisole (0.71 ± 0.32 h-1) and abamectin (0.24 ± 0.08 day-1) from the MCMDG-based formulation were considerably slower than those of other studies conducted on the commercial products. The tmax was delayed for levamisole (2.20 ± 0.45 h) and abamectin (4.20 ± 1.64 days) compared with those in published studies. Longer MRT values for levamisole (6.14 ± 1.14 h) and abamectin (8.80 ± 1.39 days) were found in this study compared to those reported. A correlation was observed between in vivo fraction absorbed and in vitro fraction released for levamisole phosphate in the MCMDG-based formulation. The injection vehicle of isotropic MCMDG-based system delayed the subcutaneous absorption of levamisole phosphate and abamectin compared to the commercial subcutaneous injection products for levamisole and abamectin. Notably, this isotropic MCMDG-based vehicle system is prepared with a combination of two drugs with different physicochemical properties.