Potential of nonoral α-lipoic acid aqueous formulations to reduce ocular microvascular complications in a streptozotocin-induced diabetic rat model.

PURPOSE: α-Lipoic acid (LA) aqueous formulations were studied for nonoral administration, including intravitreal and intraperitoneal preparations and topical eyedrops. The potential retinoprotective effects of these LA preparations were also evaluated in streptozotocin (STZ)-induced diabetic rats for screening better delivery systems of LA. METHODS: Four LA liquid preparations were prepared and investigated. The short-term accelerated stabilities of LA preparations were investigated at 3 temperatures: 50°C, 70°C, and 90°C. The time courses of LA degradation in the preparations were determined by high-performance liquid chromatography. Furthermore, the potential therapeutic effects of LA preparations in a STZ-induced diabetic rat model were assessed by vitreous fluorophotometry to evaluate the fluorescein leakage from ocular vascular vessels into the vitreous. Capillary lesion in the retina was also examined using hematoxylin-eosin-stained microsections. RESULTS: LA in an aqueous solution was rapidly degraded with the activation energy of 10.4 kcal/mol. The 3 LA preparations had shelf lives of ∼1 month at 25°C. These formulations significantly reduced the vitreous fluorescein level in STZ-induced diabetic rats as evaluated by the fluorescein leakage after tail vein injection. Capillary lesions in the retina of the diabetic rats were remarkably reduced by nonoral administration, particularly the intraperitoneal injection (30 mg/kg/day). CONCLUSIONS: LA could be developed as aqueous preparations with suitable stability for short-term use in nonoral administration. LA preparations could be administered intravitreally or intraperitoneally to reduce ocular microvascular complications, such as retinopathy, in diabetic patients.

The preparation of sustained release erythropoietin microparticle.

PURPOSE: Protein microencapsulation in biodegradable polymers is a promising route to provide for sustained release. The erythropoietin (EPO) microparticles are using human serum albumin (HSA) and poly-L-lysine (PK) as the protection complex to increased EPO integrity, entrapped efficiency and active EPO release by w/o/w solvent evaporation techniques. The optimum formulation development process was also reported by using FITC-OVA as a model protein. METHODS: The model protein FITC-ovalbumin and EPO are protected by human serum albumin and poly-L-lysine complex and encapsulated in 50:50 poly(DL-lactide-co-glycolide) by a w/o/w solvent evaporation method. Protein active integrity and degradation compound is measured by size-exclusion chromatography. Protein-loaded microparticle physical properties and in vitro active and degradation compounds release profile are characterized. RESULTS: High active integrity protein loading efficiency and particle yield of EPO or OVA-HSA/PK-loaded PLG microparticles are successfully produced by a w/o/w solvent evaporation method. Varied protection protein complex formulations and encapsulation processes are investigated. The high OVA model protein loading efficiency (80.2%), FITC-OVA content (0.24 microg mg(-1)) and yield (72.4%) are obtained by adding 100 microg mL(-1) FITC-OVA complex with 10% HSA/0.05% PK (Mw 1.5-3 kD) in the initial solution to protect the model protein. In vitro release profiles show more active OVA release from HSA/PK OVA-loaded than OVA-loaded only microparticles and also the amount of degraded protein that comes out after 3 weeks incubated in the PBS medium for OVA-loaded only microparticles is observed. The same formulation and preparation process resulted in EPO loading efficiency (68.4%), EPO content (0.23 microg mg(-1)) and yield (76.1%) for HSA/PK EPO-loaded microparticles. In vitro release profiles show active EPO sustained release over 7 days. Using HSA/PK as carried in the primary emulsion of EPO-loaded microparticles resulted in less burst release% than EPO-loaded only microparticles.

Physicochemical properties and in vivo assessment of timolol-loaded poly(D,L-lactide-co-glycolide) films for long-term intraocular pressure lowering effects.

Timolol-loaded poly(D,L-lactide-co-glycolide; PLGA) films were prepared for achieving the long-term intraocular pressure (IOP) lowering effect on glaucoma treatment. The physicochemical properties and in vivo effects of films were determined and characterize the delivery system. PLGA, span 20, propylene glycol (PG), and timolol base were dissolved in dichloromethane (DCM) and kept in a stainless mold; timolol films were prepared after the evaporation of DCM. Timolol disc-shape film preparation (TDF), containing 1 mg of timolol in 0.37 cm(2) area, was fabricated by using a trephine and placed onto the cul de sac of alpha-chymotrypsin-induced ocular hypertension rabbits for assessing the IOP lowering effect. The prepared films characterized a Young's modulus ranged from 1.13 to approximately 2.49 MPa, and related to the content percentages of PLGA, PG, and the residual DCM. The timolol film could maintain drug release for 1 week. Following a single-dose application in ocular hypertension rabbits, the prepared TDF could achieve a long-term IOP lowering effect and maintain the IOP change (in comparison with baseline) of approximately 7 mmHg within 1 week. The aqueous humor levels of timolol were low within a range of 0.8 to approximately 0.24 microg/mL for the initial 24 h and less than 0.15 microg/mL for 4-7 days. The investigated film formulation might be potentially developed for the application of long-term ocular delivery systems.

Oral immunogenicity of the inactivated Vibrio cholerae whole-cell vaccine encapsulated in biodegradable microparticles.

Vibrio cholerae (VC)-loaded microparticles as an oral vaccine delivery system were prepared with 6% w/v poly(DL-lactide-co-glycolide)(PLG) in the oil phase as well as 10% w/v PVP and 5% w/v NaCl in the aqueous phase, by an water-in-oil-in-water emulsion/solvent extraction technique. VC was successfully entrapped in the microparticles with trapping efficiencies up to 97.8% and a loading level of 55.4+/-6.9 microg/mg. The microparticle delivery system with a particle size of 3.8 microm had different distribution of VC content in the core region (25.7+/-1.9 microg/mg) and surface (6.2+/-0.9 microg/mg). The immunogenic potential of VC-loaded microparticles in comparison with PLG microparticles or VC solution was evaluated in adult mice by oral immunization, in which mice received one dose of 20 mg VC-loaded microparticles or 20 mg VC-loaded microparticles physical mixed with amphotericin B. The control group received 20 mg PLG microparticle or VC solution. Serum samples were collected from all tested mice on the day of immunization and at 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10 weeks postimmunization. Sera were examined for vibriocidal antibodies by microtitration and Vibrio-specific serum IgG and IgM antibodies were assessed by the ELISA method. IgG and IgM antibodies to intact VC were detected in sera from all animals immunized with VC. The response was specific and of high magnitude. Significantly higher antibody responses were obtained when sera from both VC-loaded microparticles and VC-loaded microparticles physical mixed with amphotericin B immunized mice were titrated against VC. The immunogenicity of VC-loaded microparticles mixed with amphotericin B in evoking serum IgG and IgM responses was higher than that of VC-loaded microparticles only. These results demonstrate that VC-loaded microparticles physical mixed with amphotericin B and VC-loaded microparticles orally administered evoke Vibrio-specific serum IgG and IgM responses as well as vibriocidal antibody activity in mice. The VC incorporation, physicochemical characterization data, and the animal results obtained in this study may be relevant in optimizing the vaccine incorporation and delivery properties of these potential vaccine targeting carriers.