Development of intranasal implantable devices for schizophrenia treatment.

In this work the preparation and characterisation of intranasal implants for the delivery of risperidone (RIS) is described. The aim of this work is to develop better therapies to treat chronic conditions affecting the brain such as schizophrenia. This type of systems combines the advantages of intranasal drug delivery with sustained drug release. The resulting implants were prepared using biodegradable materials, including poly(caprolactone) (PCL) and poly(lactic-co-glycolic acid) (PLGA). These polymers were combined with water-soluble compounds, such as poly(ethylene glycol) (PEG) 600, PEG 3000, and Tween® 80 using a solvent-casting method. The resulting implants contained RIS loadings ranging between 25 and 50 %. The obtained implants were characterised using a range of techniques including thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), attenuated total reflectance-Fourier transform infrared (ATR-FTIR), X-ray diffraction (XRD), and Scanning Electron Microscopy (SEM). Moreover, in vitro RIS release was evaluated showing that the addition of water-soluble compounds exhibited significant faster release profiles compared to pristine PCL and PLGA-based implants. Interestingly, PCL-based implants containing 25 % of RIS and PLGA-based implants loaded with 50 % of RIS showed sustained drug release profiles up to 90 days. The former showed faster release rates over the first 28 days but after this period PLGA implants presented higher release rates. The permeability of RIS released from the implants through a model membrane simulating nasal mucosa was subsequently evaluated showing desirable permeation rate of around 2 mg/day. Finally, following in vitro biocompatibility studies, PCL and PLGA-based implants showed acceptable biocompatibility. These results suggested that the resulting implants displayed potential of providing prolonged drug release for brain-targeting drugs.

Development and validation of a high-performance liquid chromatography method for levothyroxine sodium quantification in plasma for pre-clinical evaluation of long-acting drug delivery systems.

Levothyroxine (LEVO) sodium is an FDA-approved drug that is used to treat underactive thyroid (hypothyroidism) and other conditions. It is generally used as a thyroid-stimulating hormone administered orally. However, this approach has some drawbacks such as this drug should be taken every day 30 min to 1 h prior to breakfast with an empty stomach, moreover, some food interactions must be monitored. Thus, alternative innovative approaches capable of providing sustained LEVO release should be developed. Our research was designed to establish a simple quantitative determination method for LEVO in rat plasma for pre-clinical evaluation of long acting formulations using a high-performance liquid chromatography method, to validate the analytical method according to ICH guidelines and to characterise its pharmacokinetic behavior in rats. After simple protein precipitation with acetonitrile, LEVO was eluted on a Xselect CSH™ C18 column (Waters, 3.0 × 150 mm) with a particle size of 3.5 µm using a mobile phase of water and acetonitrile at a ratio of 65 : 35% v/v, including 0.1% v/v of trifluoracetic acid. The calibration standards used for plasma ranged between 0.5-1000 ng mL-1 with a correlation coefficient (r2) of ≥0.998. The limit of detection was 0.44 ng mL-1 and the lower limit of quantitation was 1.33 ng mL-1. The extraction recovery of LEVO in rat plasma samples by this method was between 80 and 85%. The method was selective, sensitive, accurate and precise for detecting and quantifying LEVO in a pharmacokinetic study carried out in rats for pre-clinical evaluation of long acting formulations. The validated HPLC method meets the ICH established requirements and therefore offers a wide range of potential applications in pre-clinical therapeutic drug monitoring, pharmacokinetics and toxicology.

Poly(caprolactone)-based subcutaneous implant for sustained delivery of levothyroxine.

This work aimed to develop a subcutaneous implant for prolonged delivery of LEVO to treat hypothyroidism. This could overcome challenges with patient compliance and co-administration and could improve treatment of this condition. For this purpose, implants were produced by solvent casting mixtures of poly(caprolactone) (PCL), poly(ethylene glycol) (PEG) and LEVO sodium. These implants contained mixtures of PCL of differing molecular weight, PEG and different LEVO sodium loadings (20% or 40% w/w). SEM images confirmed that the drug was evenly dispersed throughout the implant. In vitro release rates ranging from 28.37 ± 1.19 - 78.21 ± 19.93 µg/day and 47.39 ± 8.76 - 98.92 ± 4.27 µg/day were achieved for formulations containing 20% and 40% w/w drug loading, respectively. Implants containing higher amounts of low molecular weight PCL and 40% w/w of LEVO showed release profiles governed by zero order kinetics. On the other hand, implants containing higher amounts of high molecular weight PCL showed a release mechanism governed by Fickian diffusion. Finally, two representative formulations were tested in vivo. These implants were capable of providing detectable LEVO levels in plasma during the entire duration of the experiments (4 weeks) with LEVO plasma levels ranging between 5 and 20 ng/mL.

HPLC method for levothyroxine quantification in long-acting drug delivery systems. Validation and evaluation of bovine serum albumin as levothyroxine stabilizer.

Deficiency of thyroid hormones (hypothyroidism) is treated with oral levothyroxine (LEVO). However, the effectiveness of oral administration is highly dependent on the co-administration of food and other drugs. This factor, in combination with the chronic nature of this condition, mean that there are concerns with patient compliance. Development of long acting formulations to treat hypothyroidism could potentially solve this problem. However, LEVO instability in solution could be problematic. In order to develop long acting LEVO delivery systems in vitro drug release experiments should be carried out. However, short term LEVO stability in aqueous solution will prevent this. BSA was used as a stabiliser for LEVO; extending the stability of the drug in aqueous solutions from a few hours to 2 weeks. In order to achieve this, the required concentration of the protein was 0.1% w/v. Subsequently, an HPLC method capable of separating LEVO from the protein was developed and validated following ICH guidelines. The analysis was carried out using a reverse phase HPLC method on an Agilent 1220 Infinity II LC system. The column used to achieve separation was a Zorbax Eclipse plus C18 (95 Špore size, 250 mm length x 4.6 mm internal diameter; 5 µm particle size). The mobile phase used was composed of acetonitrile and 0.1% trifluoroacetic acid at a ratio of 50:50% v/v. UV detection of LEVO sodium was carried out at 225 nm. The retention time for the drug was 6.6 minutes. The method showed a limit of detection of 0.03 µg/mL and a limit of quantification of 0.09 µg/mL. Finally, this method was used to evaluate the release from implants containing 20% w/w of LEVO. These devices were prepared using a solvent casting method with poly(caprolactone) and LEVO. These devices showed an initial burst release over the first 3 days. Subsequently, they were capable of providing a linear release rate over the following 25 days.

Fused deposition modelling for the development of drug loaded cardiovascular prosthesis.

Cardiovascular diseases constitute a number of conditions which are the leading cause of death globally. To combat these diseases and improve the quality and duration of life, several cardiac implants have been developed, including stents, vascular grafts and valvular prostheses. The implantation of these vascular prosthesis has associated risks such as infection or blood clot formation. In order to overcome these limitations medicated vascular prosthesis have been previously used. The present paper describes a 3D printing method to develop medicated vascular prosthesis using fused deposition modelling (FDM) technology. For this purpose, rifampicin (RIF) was selected as a model molecule as it can be used to prevent vascular graft prosthesis infection. Thermoplastic polyurethane (TPU) and RIF were combined using hot melt extrusion (HME) to obtain filaments containing RIF concentrations ranging between 0 and 1% (w/w). These materials are capable of providing RIF release for periods ranging between 30 and 80 days. Moreover, TPU-based materials containing RIF were capable of inhibiting the growth of Staphylococcus aureus. This behaviour was observed even for TPU-based materials containing RIF concentrations of 0.1% (w/w). TPU containing 1% (w/w) of RIF showed antimicrobial properties even after 30 days of RIF release. Alternatively, these methods were used to prepare dipyridamole containing TPU filaments. Finally, using a dual extrusion 3D printer vascular grafts containing both drugs were prepared.

Poly(caprolactone)-Based Coatings on 3D-Printed Biodegradable Implants: A Novel Strategy to Prolong Delivery of Hydrophilic Drugs.

Implantable devices are versatile and promising drug delivery systems, and their advantages are well established. Of these advantages, long-acting drug delivery is perhaps the most valuable. Hydrophilic compounds are particularly difficult to deliver for prolonged times. This work investigates the use of poly(caprolactone) (PCL)-based implant coatings as a novel strategy to prolong the delivery of hydrophilic compounds from implantable devices that have been prepared by additive manufacturing (AM). Hollow implants were prepared from poly(lactic acid) (PLA) and poly(vinyl alcohol) (PVA) using fused filament fabrication (FFF) AM and subsequently coated in a PCL-based coating. Coatings were prepared by solution-casting mixtures of differing molecular weights of PCL and poly(ethylene glycol) (PEG). Increasing the proportion of low-molecular-weight PCL up to 60% in the formulations decreased the crystallinity by over 20%, melting temperature by over 4 °C, and water contact angle by over 40°, resulting in an increased degradation rate when compared to pure high-molecular-weight PCL. Addition of 30% PEG to the formulation increased the porosity of the formulation by over 50% when compared to an equivalent PCL-only formulation. These implants demonstrated in vitro release rates for hydrophilic model compounds (methylene blue and ibuprofen sodium) ranging from 0.01 to 34.09 mg/day, depending on the drug used. The versatility of the devices produced in this work and the range of release rates achievable show great potential. Implants could be specifically developed in order to match the specific release rate required for a number of drugs for a wide range of conditions.

Multicenter retrospective evaluation of ileocecocolic perforations associated with diagnostic lower gastrointestinal endoscopy in dogs and cats.

BACKGROUND: Ileoscopy is increasingly performed in dogs and cats with gastrointestinal signs, but iatrogenic ileocecocolic (ICC) perforations have not been described. HYPOTHESIS/OBJECTIVES: To characterize endoscopic ICC perforations in dogs and cats. ANIMALS: Thirteen dogs and 2 cats. METHODS: This is a retrospective case series. Signalment, presentation, endoscopic equipment, colonic preparation, endoscopist's experience level, ileal intubation technique, method of diagnosis, perforation location, histopathology, management, and outcome data were collected and reviewed. RESULTS: Six ileal, 5 cecal, and 4 colonic perforations were identified between 2012 and 2019. Dogs weighed 2.4-26 kg (median, 10.3 kg) and cats 4.6-5.1 kg (median, 4.9 kg). Endoscopy was performed in dogs presented for vomiting (n = 4), as well as large (n = 5), mixed (n = 4), and small (n = 1) bowel diarrhea. Cats had large bowel diarrhea. Endoscopists included 1 supervised intern, 9 supervised internal medicine residents (2 first year, 6 second year, 1 third year), and 5 internal medicine diplomates. Diagnosis was delayed in 5 dogs, occurring 1-5 days after endoscopy (median, 3 days); dogs were presented again with inappetence (n = 4), lethargy (n = 4), abdominal pain (n = 3), retching (n = 2), and syncope (n = 1). All animals underwent surgical correction. Histopathology did not identify lesions at the perforation site in any animal. Two dogs required a second surgery; 1 died 12 hours after surgery. Survival to discharge was 93%, with 78% surviving ≥8 months. CONCLUSIONS AND CLINICAL IMPORTANCE: Iatrogenic endoscopic ICC perforation is not indicative of underlying disease and is associated with a good prognosis. Delayed diagnosis can occur. Therefore, perforation should be considered in the differential diagnosis for animals with clinical deterioration after endoscopy.

Development of a Biodegradable Subcutaneous Implant for Prolonged Drug Delivery Using 3D Printing.

Implantable drug delivery devices offer many advantages over other routes of drug delivery. Most significantly, the delivery of lower doses of drug, thus, potentially reducing side-effects and improving patient compliance. Three dimensional (3D) printing is a flexible technique, which has been subject to increasing interest in the past few years, especially in the area of medical devices. The present work focussed on the use of 3D printing as a tool to manufacture implantable drug delivery devices to deliver a range of model compounds (methylene blue, ibuprofen sodium and ibuprofen acid) in two in vitro models. Five implant designs were produced, and the release rate varied, depending on the implant design and the drug properties. Additionally, a rate controlling membrane was produced, which further prolonged the release from the produced implants, signalling the potential use of these devices for chronic conditions.

Lignin and Cellulose Blends as Pharmaceutical Excipient for Tablet Manufacturing via Direct Compression.

Extensive efforts are being made to find alternative uses for lignin (LIG). In the present work the use of this biopolymer as excipient to prepare tablets was studied. For this purpose, LIG was combined with microcrystalline cellulose (MCC) and used as excipients to prepare directly compressed tablets containing a model drug, tetracycline (TC). The excipients contained different concentrations of LIG: 100%, 75%, 50%, 25% and 0% (w/w). Two different compression forces were used (two and five tonnes). When formulations were prepared using LIG as the only excipient, tablets were formed, but they showed lower densities and crushing strength than the ones obtained with only MCC or LIG/MCC blends. Moreover, tablets prepared using five tonnes of compression force showed TC releases ranging from 40% to 70% of the drug loading. On the other hand, the tablets prepared using two tonnes of compression force showed a faster and more efficient TC release, between 60% and 90%. The presence of LIG in the tablets modified significantly the release profile and the maximum amount of TC released. Finally, a DPPH (2,2-diphenyl-1-picrylhydrozyl) assay was performed to confirm that the presence of LIG provided antioxidant properties to the formulations. Accordingly, LIG has potential as a pharmaceutical excipient.

Antioxidant PLA Composites Containing Lignin for 3D Printing Applications: A Potential Material for Healthcare Applications.

Lignin (LIG) is a natural biopolymer with well-known antioxidant capabilities. Accordingly, in the present work, a method to combine LIG with poly(lactic acid) (PLA) for fused filament fabrication applications (FFF) is proposed. For this purpose, PLA pellets were successfully coated with LIG powder and a biocompatible oil (castor oil). The resulting pellets were placed into an extruder at 200 °C. The resulting PLA filaments contained LIG loadings ranging from 0% to 3% (w/w). The obtained filaments were successfully used for FFF applications. The LIG content affected the mechanical and surface properties of the overall material. The inclusion of LIG yielded materials with lower resistance to fracture and higher wettabilities. Moreover, the resulting 3D printed materials showed antioxidant capabilities. By using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) method, the materials were capable of reducing the concentration of this compound up to ca. 80% in 5 h. This radical scavenging activity could be potentially beneficial for healthcare applications, especially for wound care. Accordingly, PLA/LIG were used to design meshes with different designs for wound dressing purposes. A wound healing model compound, curcumin (CUR), was applied in the surface of the mesh and its diffusion was studied. It was observed that the dimensions of the meshes affected the permeation rate of CUR. Accordingly, the design of the mesh could be modified according to the patient's needs.

Fused Deposition Modeling as an Effective Tool for Anti-Infective Dialysis Catheter Fabrication.

Catheter-associated infections are a common complication that occurs in dialysis patients. Current strategies to prevent infection include catheter coatings containing heparin, pyrogallol, or silver nanoparticles, which all have an increased risk of causing resistance in bacteria. Therefore, a novel approach for manufacture, such as the use of additive manufacturing (AM), also known as three-dimensional (3D) printing, is required. Filaments were produced by extrusion using thermoplastic polyurethane (TPU) and tetracycline hydrochloride (TC) in various concentrations (e.g., 0, 0.25, 0.5, and 1%). The extruded filaments were used in a fused deposition modeling (FDM) 3D printer to print catheter constructs at varying concentrations. Release studies in phosphate-buffered saline, microbiology studies, thermal analysis, contact angle, attenuated total reflection-Fourier transform infrared, scanning electron microscopy, and X-ray microcomputer tomography (µCT) analysis were conducted on the printed catheters. The results suggested that TC was uniformly distributed within the TPU matrix. The microbiology testing of the catheters showed that devices containing TC had an inhibitory effect on the growth of Staphylococcus aureus NCTC 10788 bacteria. Catheters containing 1% TC maintained inhibitory effect after 10 day release studies. After an initial burst release in the first 24 h, there was a steady release of TC in all concentrations of catheters. 3D-printed antibiotic catheters were successfully printed with inhibitory effect on S. aureus bacteria. Finally, TC containing catheters showed resistance to S. aureus adherence to their surfaces when compared with catheters containing no TC. Catheters containing 1% of TC showed a bacterial adherence reduction of up to 99.97%. Accordingly, the incorporation of TC to TPU materials can be effectively used to prepare anti-infective catheters using FDM. This study highlights the potential for drug-impregnated medical devices to be created through AM.

Implantable Polymeric Drug Delivery Devices: Classification, Manufacture, Materials, and Clinical Applications.

The oral route is a popular and convenient means of drug delivery. However, despite its advantages, it also has challenges. Many drugs are not suitable for oral delivery due to: first pass metabolism; less than ideal properties; and side-effects of treatment. Additionally, oral delivery relies heavily on patient compliance. Implantable drug delivery devices are an alternative system that can achieve effective delivery with lower drug concentrations, and as a result, minimise side-effects whilst increasing patient compliance. This article gives an overview of classification of these drug delivery devices; the mechanism of drug release; the materials used for manufacture; the various methods of manufacture; and examples of clinical applications of implantable drug delivery devices.

Nocturama gen. nov., Nothocladus s. lat. and other taxonomic novelties resulting from the further resolution of paraphyly in Australasian members of Batrachospermum (Batrachospermales, Rhodophyta).

The informal "Australasica Group" was established in 2009 to include several Australasian endemic Batrachospermum species, a few species of the cosmopolitan Batrachospermum section Setacea, and the South American endemic Petrohua bernabei. Although useful for communication purposes, no formal taxonomic designation was proposed due to weakly supported basal nodes. The present research took a two-pronged approach of adding more taxa (29 additional specimens) as well as more sequence data (LSU, cox1, psaA, and psbA markers added to rbcL data) to provide better resolution. The resulting tree showed improved statistical support values (Bayesian posterior probability and maximum likelihood bootstrap) for most nodes providing a framework for taxonomic revision. Based on our well-resolved phylogeny, a new genus, Nocturama, is proposed for a clade of Batrachospermum antipodites specimens. The circumscription of Nothocladus is expanded to include Batrachospermum section Setacea and four additional sections composed of at least 10 species, mostly from Australia and New Zealand. One new species added to the data set, N. diatyches, did not form a clade with the other species of section Setaceus, where it was classified previously, rendering that section paraphyletic. To resolve this, N. diatyches and the morphologically similar species N. latericius are included with N. theaquus, in the new section Theaquus within Nothocladus s. lat. A specimen from Australia unaligned to these clades was sister to the Australia-New Zealand genus Psilosiphon and the cosmopolitan B. cayennense, but lacked statistical support. This specimen has the gross morphology of Batrachospermum s. lat. and is here provisionally assigned to that genus, as B. serendipidum sp. nov.

Methylene Blue-Loaded Dissolving Microneedles: Potential Use in Photodynamic Antimicrobial Chemotherapy of Infected Wounds.

Photodynamic therapy involves delivery of a photosensitising drug that is activated by light of a specific wavelength, resulting in generation of highly reactive radicals. This activated species can cause destruction of targeted cells. Application of this process for treatment of microbial infections has been termed "photodynamic antimicrobial chemotherapy" (PACT). In the treatment of chronic wounds, the delivery of photosensitising agents is often impeded by the presence of a thick hyperkeratotic/necrotic tissue layer, reducing their therapeutic efficacy. Microneedles (MNs) are an emerging drug delivery technology that have been demonstrated to successfully penetrate the outer layers of the skin, whilst minimising damage to skin barrier function. Delivering photosensitising drugs using this platform has been demonstrated to have several advantages over conventional photodynamic therapy, such as, painless application, reduced erythema, enhanced cosmetic results and improved intradermal delivery. The aim of this study was to physically characterise dissolving MNs loaded with the photosensitising agent, methylene blue and assess their photodynamic antimicrobial activity. Dissolving MNs were fabricated from aqueous blends of Gantrez(®) AN-139 co-polymer containing varying loadings of methylene blue. A height reduction of 29.8% was observed for MNs prepared from blends containing 0.5% w/w methylene blue following application of a total force of 70.56 N/array. A previously validated insertion test was used to assess the effect of drug loading on MN insertion into a wound model. Staphylococcus aureus, Escherichia coli and Candida albicans biofilms were incubated with various methylene blue concentrations within the range delivered by MNs in vitro (0.1-2.5 mg/mL) and either irradiated at 635 nm using a Paterson Lamp or subjected to a dark period. Microbial susceptibility to PACT was determined by assessing the total viable count. Kill rates of >96%, were achieved for S. aureus and >99% for E. coli and C. albicans with the combination of PACT and methylene blue concentrations between 0.1 and 2.5 mg/mL. A reduction in the colony count was also observed when incorporating the photosensitiser without irradiation, this reduction was more notable in S. aureus and E. coli strains than in C. albicans.

Accidental entrapment of cats in front-loading washing machines.

Two clinical cases of accidental entrapment of cats in front-loading washing machines are described. One cat died the day after presentation as a result of aspiration pneumonia and head trauma, despite supportive care. The second cat survived with supportive treatment, but developed dermatologic complications 10 d later.

Effect of energy substrates on protein degradation in isolated small intestinal enterocytes from rats.

BACKGROUND: Nutrients affect small intestinal protein mass and metabolism, but studies on the effect of nutrients on small intestinal protein degradation are very limited due to a lack of a proper method. The objectives of this study were to establish a method to directly estimate protein degradation in isolated enterocytes from rats and to test the effect of energy substrates on protein degradation. METHODS: Male Sprague-Dawley rats (150-200 g, n>or=8 per treatment) were used. Cell viability, tyrosine release as an indicator of protein degradation, and the effect of osmolarity, 50 mmol/L glucose, 20 mmol/L beta-hydroxybutyrate, 4.7 mmol/L butyrate, and 30 mmol/L glutamine on protein degradation were measured. RESULTS: The average viability of enterocytes at time 30 minutes was 85.8% (range, 81%-94%). Tyrosine release was linear over the course of experiments, indicating constant protein degradation (R2=0.9943; p<.05). Osmolarity, glucose, and glutamine had no effect on protein degradation in isolated enterocytes. Beta-hydroxybutyrate significantly decreased it (-16%; p<.05), whereas butyrate slightly increased it (+5%; p<.05). CONCLUSIONS: A high viability and constant protein degradation indicate a successful establishment of a method to estimate protein degradation in isolated small intestinal enterocytes from rats. The large effect of beta-hydroxybutyrate suggests a potential positive role for ketone bodies to limit the loss of small intestinal protein mass by decreasing protein degradation.

Liver protein synthesis stays elevated after chemotherapy in tumour-bearing mice.

We studied the effect of chemotherapy on liver protein synthesis in mice bearing colon 26 adenocarcinoma (C26). Liver protein mass decreased (-32%; P<0.05) in cachectic mice, but protein synthesis increased (20-35%; P<0.05) in cachectic mice, which is consistent with increased export protein synthesis. Increased protein synthesis in tumour-bearing mice was primarily mediated by increasing ( approximately 15%; P<0.05) the RNA concentration, i.e. the capacity for protein synthesis (Cs; mg RNA/g protein). Cystemustine, a nitrosourea chemotherapy that cures C26 with 100% efficacy, rapidly restored liver protein mass; protein synthesis however stayed higher than in healthy mice ( approximately 15%) throughout the initial and later stages of recovery. Chemotherapy had no significant effect on liver protein mass and synthesis in healthy mice. Reduced food intake was not a factor in this model. These data suggest a high priority for liver protein synthesis during cancer cachexia and recovery.