Development of acetazolamide-loaded,pH-triggered polymeric nanoparticulate in situ gel for sustained ocular delivery: in vitro. ex vivo evaluation and pharmacodynamic study

The objective of research was to develop a novel pH-triggered polymeric nanoparticulate in situ gel (NP-ISG) for ophthalmic delivery of acetazolamide (ACZ) to enhance the conjunctival permeation and precorneal residence time of the formulation by overcoming the limitations of protective ocular barriers. Nanoparticles (NP1--NP12) were developed by nanoprecipitation method and evaluated for pharmacotechnical characteristics including transmission electron microscopy. The optimized formulation, NP10 was dispersed in carbopol 934?P to form nanoparticulate in situ gels (NP-ISG1--NP-ISG5). NP-ISG5 was selected as optimized formulation on the basis of gelation ability and residence time. Ex vivo transcorneal permeation study exhibited significantly higher ACZ permeation from NP-ISG5 (74.50?±?2.20?mg/cm²) and NP10 (93.5?±?2.25?mg/cm²) than eye drops (20.08?±?3.12?mg/cm²) and ACZ suspension (16.03?±?2.14). Modified Draize test with zero score indicated nonirritant property of NP-ISG5. Corneal toxicity study revealed no visual signs of tissue damage. Further, NP-ISG5 when tested for hypotensive effect on intraocular pressure (IOP) in rabbits revealed that NP-ISG5 caused significant decrease in IOP (p?in vitro efficacy, safety and patient compliance.     

Multipronged,strategic delivery of paclitaxel-topotecan using engineered liposomes to ovarian cancer

Context: Synergistically active combinations have been used to enhance therapeutic efficacy for ovarian cancer chemotherapy.

Objective: The synergistically active combination of paclitaxel-topotecan (Pac-Top; 20:1, w/w) were loaded into folate-anchored PEGylated liposomes (FPL-Pac-Top) for safe and effective treatment of ovarian cancer.

Materials and methods: Coupling reactions were carried out using carbodiimide chemistry and confirmed by infrared spectral analysis. These liposomes were studied for shape and physical interaction (and integrity), in vitro drug release kinetics, hemolytic toxicity, ex vivo pharmacodynamics in OVCAR-3 cell lines, and pharmacokinetics in ovarian tumor-bearing mice.

Results: The differential scanning calorimeter studies exhibited melting of liposomes (～150?nm) at ～41?°C. The drug(s) release from liposomes followed Fickian diffusion model. The hematological studies revealed insignificant toxicity to blood cells. In vivo studies showed long circulatory behavior (increased AUC_0–t and AUMC_0–t and MRT) and selective accumulation of FPL-Pac-Top in the ovaries. FPL-Pac-Top showed less necrosis and more apoptosis in flow cytometry. Kaplan–Meier survival analysis revealed the doubling of the survival time with FPL-Pac-Top in comparison to Pac-Top solution.

Discussion and conclusion: Potentiated anti-cancer activity of FPL-Pac-Top was attributed to multiple features viz. thermosensitivity, long circulatory nature and targetability. Such approach could be a paradigm chemotherapeutic approach for safe and effective targeting of cancer.     

7-Ethyl-10-hydroxycamptothecin proliposomes with a novel preparation method: optimized formulation,characterization and in-vivo evaluation

Context:?The proliposomes were used to solve the stability of the ordinary liposomes. Objective: 7-ethyl-10-hydroxycamptothecin (SN-38) proliposomes for intravenous (i.v.) administration were prepared successfully by a new method.

Materials and methods:?SN-38 liposomes solution was reconstituting automatically from proliposomes on contact with the acetic acid buffer solution (0.2 M, pH 2.6). The formulation was optimized by the Box–Behnken design. The physicochemical characteristics of the SN-38 proliposomes were studied by scanning electron microscopy (SEM), transmission electron microscopy (TEM), differential scanning calorimetry (DSC) and X-ray diffraction (XRD). The stability studies were also carried on. The FLU–HPLC system was served to study the concentration of SN-38 in the plasma of Sprague Dawley (SD) rats.

Results:?The optimized formulation was SN-38: 0.03 g; Soybean phospholipid (SP): 0.6 g; dextrose: 3.00 g. The entrapment efficiency of the optimized formulation was >85% and the mean particle size was about 231 nm. The stability studies showed that SN-38 proliposomes were stable in dark at 20–25°C for 6 months at least. The pharmacokinetic parameters of i.v. administration demonstrated that the half-life of SN-38 loaded in the liposomes was prolonged in vivo.

Discussion and conclusion:?The SN-38 proliposomes was prepared successful by the analysis of TEM, SEM, DSC and XRD, and SN-38 liposomes could be reconstituted on contact with the hydration medium. SN-38 liposomes circulated for a longer time in the blood circulating system than SN-38 solution, which contributed to maintaining the drug action.     

In vitro and in vivo investigation of taste-masking effectiveness of Eudragit E PO as drug particle coating agent in orally disintegrating tablets

Abstract

Context: Considering that bitter taste of drugs incorporated in orally disintegrating tablets (ODTs) can be the main reason for avoiding drug therapy, it is of the utmost importance to achieve successful taste-masking. The evaluation of taste-masking effectiveness is still a major challenge.

Objective: The objective of this study was to mask bitter taste of the selected model drugs by drug particle coating with Eudragit^® E PO, as well as to evaluate taste-masking effectiveness of prepared ODTs using compendial dissolution testing, dissolution in the small-volume shake-flask assembly and trained human taste panel.

Materials and methods: Model drugs were coated in fluidized bed. Disintequik? ODT was used as a novel co-processed excipient for ODT preparation. Selected formulations were investigated in vitro and in vivo using techniques for taste-masking assessment.

Results and discussion: Significantly slower drug dissolution was observed from tablets with coated drug particles during the first 3?min of investigation. Results of in vivo taste-masking assessment demonstrated significant improvement in drug bitterness suppression in formulations with coated drug. Strong correlation between the results of drug dissolution in the small-volume shake-flask assembly and in vivo evaluation data was established (R?≥?0.970).

Conclusion: Drug particle coating with Eudragit^® E PO can be a suitable approach for bitter taste-masking. Strong correlation between in vivo and in vitro results implicate that small-volume dissolution method may be used as surrogate for human panel taste-masking assessment, in the case of physical taste-masking approach application.     

Fusogenic liposomal formulation of sirolimus: improvement of drug anti-proliferative effect on human T-cells

Context: Fusogenic liposomes are unique delivery vehicles capable of introducing their contents directly and efficiently into the cytoplasm.

Objective: The objective of this study was to evaluate the potential of fusogenic liposomes containing Sirolimus to improve its anti-proliferative effect on T-lymphocyte cells.

Materials and methods: Conventional liposomes containing Sirolimus were prepared from Dipalmitoylphosphatidylcholine (DPPC) and cholesterol using the modified ethanol injection method. To prepare fusogenic liposomes, dioleoylphosphatidylethanolamine (DOPE) was added to the conventional liposome formulation. The liposomes were characterized by their size, zeta potential, encapsulation efficiency percent (EE%) and chemical stability during 6 months. The in vitro release of liposomes, anti-proliferative effect and liposome uptake of both types of liposomes with optimized formulations were studied on human T-lymphocyte cells employing the MTT assay and fluorescein isothiocyanate-loaded liposomes.

Results and discussion: The particle size of the liposomes was evaluated between 138 and 650?nm and mean zeta potential was in the range of ?32.95 to ?45.60?mV. The average EE% of the prepared conventional and fusogenic liposomes were 76.9% and 80.5%, respectively. Liposomal formulations released only 10–20% of encapsulated drug without any burst effect. In vitro immunosuppressive evaluation on T-cells showed that fusogenic liposomes have the best anti-proliferative effects and uptake on T-lymphocyte cell compared to the conventional liposomes.

Conclusion: Our results indicated that fusogenic liposomes can be useful carriers for improving the inhibition of T-cell proliferation.     

Preparation and IVIVC evaluation of salvianolic acid B micro-porous osmotic pump pellets

Purpose: Salvianolic acid B micro-porous osmotic pump controlled release pellets (SalB-CRPs) with suitable in vitro release profiles and good in vitro and in vivo correlation (IVIVC) were developed.

Method: Extrusion-spheronization was used to prepare the starter cores containing SalB/MCC/Kollidon®CL-SF/Flowlac®100 of 30:40:15:15 [w/w, The formulation composition of SalB immediate-release pellets (SalB-IRPs)] and complexed with lactose. The pellets were subsequently coated with Surelease aqueous dispersion to achieve controlled-release properties. Furthermore, a single-dose pharmacokinetics study was carried out in New Zealand White (NZW) rabbits.

Results: In the starter cores, the lactose content was 25% based on the SalB-IRPs constituent. The optimal coating polymer ratio of Surelease aqueous dispersion and polyvinyl alcohol–polyethylene glycol (PVA–PEG) graft copolymer (EC/PVA–PEG) was found to be 70:30 (w/w, %) with a coating weight of 5%. The prepared SalB-CRPs had similar in vitro release under three different pH release mediums. A good IVIVC was characterized by a high coefficient of determination (r?=?0.9801). The in vivo study indicated that the maximum plasma concentration (C_max) of SalB-CRPs was decreased, peak concentration time (T_max) and mean residence time (MRT) were all prolonged, as that of SalB-IRPs. In addition, the area under concentration–time curve from 0 to 24?h (AUC_0–24?h) and 0 to infinity (AUC_0–∞) were significantly higher, compared with those of SalB-IRPs.

Conclusion: Collectively, these results manifested that SalB-CRPs were likely to be a more suitable formulation in treating cardiovascular disease with improved in vivo retention, decreased plasma drug concentration fluctuation.     

Development of delayed-release proliposomes tablets for oral protein drug delivery

Context: One among many attempts to improve oral protein drug delivery was utilizing the colloidal drug carriers particularly liposomes.

Objective: The purpose was to develop proliposomes of bovine serum albumin (BSA) in the form of granules and delayed-release tablets by using simple tablet manufacturing process.

Materials and methods: BSA proliposomes granules were prepared by spraying 7:3 (w/w) – lecithin:cholesterol solution mixture onto BSA-mannitol granules rotating in a glass coating pan. BSA proliposomes granules were directly compressed into tablets and subsequently coated with Eudragit^® L100 film. The physical properties and stability in gastrointestinal fluids of delayed-release BSA proliposomes tablets as well as reconstituted liposomes were assessed.

Results: The BSA proliposomes tablets disintegrated readily and the obtained reconstituted BSA liposomes exhibited multilamellar vesicles, the size and entrapment efficiency of which were around 2–3 µm and 10–14%, respectively. The delayed-release BSA proliposomes tablets were found to be relatively stable in United States Pharmacopoeia (USP) simulated gastric and intestinal fluids. Increase in amount of BSA in granules resulted in the increase in entrapment efficiency and loading capacity.

Discussion: The Fourier transform infrared spectroscopy (FTIR) results indicated increase in α-helix structure of BSA entrapped in liposomes. ³¹P phosphorous nuclear magnetic resonance spectroscopy (³¹P-NMR) spectrum indicated interaction between BSA molecules and phosphoric acid polar groups of bilayers membrane.

Conclusion: The delayed-release BSA proliposomes tablets developed could completely be reconstituted into liposomes with sufficient resistance to the hostile environment in gastrointestinal tract.     

Enhanced antitumor efficacy of gemcitabine-loaded temperature-sensitive liposome by hyperthermia in tumor-bearing mice

Introduction: Gemcitabine-loaded TSL (Gem-TSL) was used in combination with hyperthermia (HT) to treat the colon adenocarcinoma-bearing BALB/c mice for improved anticancer effect following intravenous administration.

Methods: A new temperature-sensitive liposome (TSL), composed of DPPC:DMPC:DSPC (4:1:1 molar ratio) releasing the encapsulated gemcitabine (Gem) at 41?°C, was developed and evaluated for enhanced antitumor efficacy both in vitro and in vivo.

Results: Drug release from the TSL was sharply increased at 41?°C and in vitro cytotoxicity of Gem-TSL in colon adenocarcinoma cells (CT-26) was 10 times higher than the free drug (IC50?=?0.3?μM versus 3?μM). Apoptosis seemed to be the main mechanism of cell death as the treatment of the cells with Gem-TSL increased the caspse-3/7 activity by 1.5-fold and also caused the fragmentation of chromatin DNA. Gem-TSL suppressed the tumor growth in CT-26-bearing BALB/c mice more stronger than the free gemcitabine after intravenous administration. Moreover, this in vivo antitumor efficacy of Gem-TSL was further increased when HT was added.

Discussion: This study suggests that this new TSL-Gem formulation could serve as a new chemotherapy modality together with HT.     

Formulation and ex vivo–in vivo evaluation of pH-triggered brimonidine tartrate in situ gel for the glaucoma treatment using application of 32 factorial design

Context: Short residence time, poor bioavailability and poor permeability are the major problems for conventional eye drops treatment.

Objective: The aim of this article is to develop, optimize and ex vivo–in vivo investigation of brimonidine tartrate in situ gel as compared to marketed eye drops for the treatment of glaucoma.

Materials and methods: The effect of independent variables, namely concentrations of polymers, on various dependent variables like viscosity at physiological pH and in vitro drug release were studied by using 3² factorial design. Further the optimized formulation was characterized for ex vivo and in vivo study.

Results and discussion: Experimental data demonstrated that optimized in situ gel formulation (F8) showed in vitro–ex vivo sustained release profile with polymer composites carbopol 974P and HPMC K4M. After 5?h of ex vivo transcorneal permeation study, the amount recovered from the corneal surface on the donor chamber 12.40% (124 ug) and the amount collected from the receptor chamber 76.8% (760 ug) of the initial dose 1?mg. The total amount recovered from the permeation experiment was 89.2%. Bioadhesive carbopol 974P and viscosity HPMC K4M composites optimized formulation (F 8) produce greater influence on the duration of drug action and improved intraocular pressure reduction activity as compared to marketed eye drop solution in in vivo study.

Conclusion: The developed in situ gelling system as a promising ophthalmic formulation to prolong the drug lowering effect on the intraocular pressure.     

Nanosponge-based pediatric-controlled release dry suspension of Gabapentin for reconstitution

Abstract

Context: Gabapentin was selected to formulate oral controlled release dry suspension because of short biological half life of 5–7?h and low bioavailability (60%). Gabapentin is a bitter drug so an attempt was made to mask its taste.

Objective: To formulate and evaluate controlled release dry suspension for reconstitution to increase the bioavailability and to control bitter taste of drug.

Materials and methods: Cyclodextrin based nanosponges were synthesized by previously reported melt method. The nanosponge–drug complexes were characterized by FTIR, DSC and PXRD as well as evaluated for taste and saturation solubility. The complexes were coated on Espheres by a suspension layering technique followed by coating with ethyl cellulose and Eudragit RS-100. A dry powder suspension for reconstitution of the microspheres was formulated and evaluated for taste, redispersibility, in vitro dissolution, sedimentation volume, leaching and pharmacokinetics.

Results and discussion: The complexes showed partial entrapment of drug nanocavities. Significant decrease in solubility (25%) was observed in the complexes than pure drug in different media. The microspheres of nanosponge complexes showed desired controlled release profile for 12?h. Insignificant drug leaching was observed in reconstituted suspension during storage for 7 days at 45?°C/75% RH. Nanosponges effectively masked the taste of Gabapentin and the coating polymers provided controlled release of the drug and enhanced taste masking. The results of in vivo studies showed increase in bioavailability of controlled release suspension by 24.09% as compared to pure drug.

Conclusion: The dry powder suspension loaded with microspheres of nanosponges complexes can be proposed as a suitable controlled release drug delivery for Gabapentin.     

The preparation,characterization of Lupeol PEGylated liposome and its functional evaluation in vitro as well as pharmacokinetics in rats

Objective: The objective of this study was to enhance the solubility and bioavailability of Lupeol.

Methods: Utilizing a thin-film dispersion method, we prepared Lupeol-loaded PEGylated liposomes and Lupeol-loaded liposomes, which was characterized using SEM, mean diameter, PDI, zeta potential, and entrapment efficiency (EE). The EE, in vitro release, and stability of Lupeol-loaded PEGylated liposomes were detected using HPLC. In addition to the safety evaluation, the evaluation was carried out on HepG2 cells in vitro; the pharmacokinetics were carried out after i.v. in the rats.

Results: The size, PDI, zeta potential, and EE of Lupeol-loaded PEGylated liposomes and Lupeol-loaded liposomes were 126.9?nm, 0.246, ?1.97?mV, 87%; 97.23?nm, 0.25, 1.6?mV, 86.2%, respectively. Lupeol-loaded PEGylated liposomes showed the slow-release effect in vitro release experiments. Lupeol-loaded PEGylated liposomes offered significant advantages over other experimental groups in vitro studies, such as the highest inhibition rate and the highest apoptosis rate. We also found that Lupeol-loaded PEGylated liposomes blocked cells in the G₂M phase. The pharmacokinetics result showed that the AUC of Lupeol-loaded PEGylated liposomes group was 3.2 times higher than free Lupeol group after i.v., the MRT and t_1/2 values of Lupeol-loaded PEGylated liposomes (MRT = 6.09?h, t_1/2 =12.94?h) showed improvements of 2.5 and 4.1 times compared to free Lupeol (MRT = 2.43?h, t_1/2 = 3.16?h).

Conclusion: The Lupeol-loaded PEGylated liposomes have successfully solved its poor hydrophilicity, low bioavailability.     

Intradermal and follicular delivery of adapalene liposomes

Abstract

Background: Adapalene is a widely used topical anti-acne drug; however, it has many side effects. Liposomal drug delivery can play a major role by targeting delivery to pilosebaceous units, reducing side effects and offering better patient compliance.

Objective: To prepare and evaluate adapalene-encapsulated liposomes for their physiochemical and skin permeation properties.

Methods: A liposomal formulation of adapalene was prepared by the film hydration method and characterized for shape, size, polydispersity index (PDI), encapsulation efficiency and thermal behavior by techniques such as Zetasizer®, differential scanning calorimetry and transmission electron microscopy. Stability of the liposomes was evaluated for three months at different storage conditions. In vitro skin permeation studies and confocal laser microscopy were performed to evaluate adapalene permeation in pig ear skin and hair follicles.

Results: The optimized process and formulation parameters resulted in homogeneous population of liposomes with a diameter of 86.66?±?3.5?nm in diameter and encapsulation efficiency of 97.01?±?1.84% w/w. In vitro permeation studies indicated liposomal formulation delivered more drug (6.72?±?0.83?μg/cm²) in hair follicles than gel (3.33?±?0.26?μg/cm²) and drug solution (1.62?±?0.054?μg/cm²). Drug concentration delivered to the skin layers was also enhanced compared to other two formulations. Confocal microscopy images confirmed drug penetration in the hair follicles when delivered using the liposomal formulation.

Conclusion: Adapalene was efficiently encapsulated in liposomes and led to enhanced delivery in hair follicles, the desired target site for acne.     

Brucine-loaded liposomes composed of HSPC and DPPC at different ratios: in vitro and in vivo evaluation

Objective: The objective of this study is to test the hypothesis that the phase transition temperature (T_m), the main property of liposomes, can be easily controlled by changing the molar ratio of hydrogenated soy phosphatidylcholine (HSPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphacholine (DPPC) after drug encapsulation.

Materials and methods: Brucine, an antitumor alkaloid, was encapsulated into the liposomes with different HSPC/DPPC compositions. The T_ms of the brucine-loaded liposomes (BLs) were determined by differential scanning calorimetry (DSC). Then the physicochemical properties and pharmacokinetics of the BLs with different HSPC/DPPC compositions were investigated and compared.

Results: The results of DSC revealed that HSPC and DPPC can combine into one phase. The findings of molecular modeling study suggested that HSPC interacts with DPPC via electrostatic interaction. The molar ratio of HSPC/DPPC influenced the sizes of BLs but had little effect on the entrapment efficiency (EE). The stability of BLs was improved with the increase of the HSPC ratios, especially with the presence of plasma. Following i.v. administration, it was found that AUC values of BLs in vivo were directly related to the HSPC/DPPC ratios of BLs, namely the T_ms of BLs.

Discussion: The behavior of liposomes, especially in vivo pharmacokinetic behavior, can be controlled by the modification of T_m.

Conclusion: The characterization of BLs in vitro and in vivo had demonstrated that the T_m could be flexibly modified for liposomes composed of both HSPC and DPPC. Using HSPC/DPPC composition may be an efficient strategy to control the T_m, thus control the in vivo pharmacokinetic behavio, of BLs.     

Bioavailability enhancement of baclofen by gastroretentive floating formulation: statistical optimization,in vitro and in vivo pharmacokinetic studies

Objectives: The study was aimed to improve bioavailability of baclofen by developing gastroretentive floating drug delivery system (GFDDS).

Methods: Preliminary optimization was done to select various release retardants to obtain minimum floating lag time, maximum floating duration and sustained release. Optimization by 3² factorial design was done using Polyox WSR 303 (X₁) and HPMC K₄M (X₂) as independent variables and cumulative percentage drug released at 6?h (Q6h) as dependent variable.

Results: Optimized formulation showed floating lag time of 4–5 s, floated for more than 12?h and released the drug in sustained manner. In vitro release followed zero ordered kinetics and when fitted to Korsemeyer Peppas model, indicated drug release by combination of diffusion as well as chain relaxation. In vivo floatability study confirmed floatation for more than 6?h. In vivo pharmacokinetic studies in rabbits showed C_max of 189.96?±?13.04?ng/mL and T_max of 4?±?0.35?h for GFDDS. The difference for AUC_(0–T) and AUC_(0–∞) between the test and reference formulation was statistically significant (p > 0.05). AUC_(0–T) and AUC_(0–∞) for GFDDS was 2.34 and 2.43 times greater than the marketed formulation respectively.

Conclusion: GFDDS provided prolonged gastric residence and showed significant increase in bi oavailability of baclofen.     

The effect of nanoliposomal formulations on Staphylococcus epidermidis biofilm

Objective: The aim of the present study was to assess the in vitro antimicrobial activities of nanoliposomal formulations loaded with vancomycin or/and rifampin against the biofilm formed by Staphylococcus epidermidis at 37?°C under aerobic condition.

Materials and methods: Liposomal formulations were prepared by dehydration-rehydration (DRV) method and characterized for size, zeta potential and encapsulation efficacy. The ability of different formulations on eradication of bacterial biofilm was assessed through optical density ratio (OD_r) and the results implicate higher survival rates of S. epidermidis on biofilm. Positive control was defined as an OD_r?=?1.0.

Results: The zeta potential of anionic, cationic and PEGylated liposomes was ?35?±?2, 35?±?1 and 27?±?2?mV whereas the mean sizes of these liposomal formulations were 145?±?4, 134?±?1 and 142?±?6?nm, respectively. Encapsulation efficacy of rifampin and vancomycin was more than 60% and about 25%, respectively. Cationic liposomal rifampin lowered the OD_r to 0.61 and was the most effective formulations against S. epidermidis biofilm (p?Conclusion: The results of this study showed that rifampin-loaded liposomes were effective against bacterial biofilm.     

Cuboidal lipid polymer nanoparticles of rosuvastatin for oral delivery

The present work aimed to develop and characterize sustained release cuboidal lipid polymeric nanoparticles (LPN) of rosuvastatin calcium (ROS) by solvent emulsification-evaporation process. A three factor, two level (2³) full-factorial design was applied to study the effect of independent variables, i.e. amount of lipid, surfactant and polymer on dependent variables, i.e. percent entrapment efficiency and particle size. Optimized formulations were further studied for zeta potential, TEM, in vitro drug release and ex vivo intestinal permeability. Cuboidal nanoparticles exhibited average particle size 61.37?±?3.95?nm, entrapment efficiency 86.77?±?1.27% and zeta potential ?6.72?±?3.25?mV. Nanoparticles were lyophilized to improve physical stability and obtain free-flowing powder. Effect of type and concentration of cryoprotectant required to lyophilize nanoparticles was optimized using freeze-thaw cycles. Mannitol as cryoprotectant in concentration of 5-8% w/v was found to be optimal providing zeta potential ?20.4?±?4.63?mV. Lyophilized nanoparticles were characterized using FTIR, DSC, XRD and SEM. Absence of C=C and C–F aromatic stretch at 1548 and 1197?cm^?1, respectively, in LPN indicated coating of drug by lipid and polymer. In vitro diffusion of ROS using dialysis bag showed pH-independent sustained release of ROS from LPN in comparison to drug suspension. Intestinal permeability by non-everted gut sac model showed prolonged release of ROS from LPN owing to adhesion of polymer to mucus layer. In vivo absorption of ROS from LPN resulted in 3.95-fold increase in AUC_0–last and 7.87-fold increase in mean residence time compared to drug suspension. Furthermore modified tyloxapol-induced rat model demonstrated the potential of ROS-loaded LPN in reducing elevated lipid profile.     

Aerosolized liposomes with dipalmitoyl phosphatidylcholine enhance pulmonary absorption of encapsulated insulin compared with co-administered insulin

Objective: We have previously shown that aerosolized liposomes with dipalmitoyl phosphatidylcholine (DPPC) enhance the pulmonary absorption of encapsulated insulin. In this study, we aimed to compare insulin encapsulated into the liposomes versus co-administration of empty liposomes and unencapsulated free insulin, where the DPCC liposomes would serve as absorption enhancer.

Significance: The present study provides the useful information for development of noninvasive treatment of diabetes.

Methods: Co-administration of empty DPPC liposomes and unencapsulated free insulin was investigated in vivo to assess the potential enhancement in protein pulmonary absorption. Co-administration was compared to DPPC liposomes encapsulating insulin, and free insulin.

Results: DPPC liposomes enhanced the pulmonary absorption of unencapsulated free insulin; however, the enhancing effect was lower than that of the DPPC liposomes encapsulating insulin. The mechanism of the pulmonary absorption of unencapsulated free insulin by DPPC liposomes involved the opening of epithelial cell space in alveolar mucosa, and not mucosal cell damage, similar to that of the DPPC liposomes encapsulating insulin. In an in vitro stability test, insulin in the alveolar mucus layer that covers epithelial cells was stable. These findings suggest that, although unencapsulated free insulin spreads throughout the alveolar mucus layer, the concentration of insulin released near the absorption surface is increased by the encapsulation of insulin into DPPC liposomes and the absorption efficiency is also increased.

Conclusion: We revealed that the encapsulation of insulin into DPPC liposomes is more effective for pulmonary insulin absorption than co-administration of DPPC liposomes and unencapsulated free insulin.     

Cryocornea – toward enhancing the capacity and throughput of ex vivo corneal model

Abstract

Objective: The objective of this study was to investigate the effects of the concentration of two intracellular (i.e. propylene glycol and glycerol) and four extracellular (i.e. dextran, hydroxypropyl methylcellulose, polyvinylpyrolidone, trehalose) cryoprotective agents as well as the effects of freeze-thawing procedures on the corneal cryoprotection.

Significance: The corneal cryopreservation may possibly become the long-term storage technique of choice for collection of animal corneas suitable for ex vivo drug testing.

Methods: The integrity of corneal barrier was evaluated by measurements of transepithelial electrical resistance.

Results: Under the investigated experimental conditions the best result was obtained for slow freezing (2?h at ?20?°C followed by 46?h at ?70?°C) and rapid thawing (0.25?h at 34?°C) procedure where 20% (w/V) trehalose in Krebs Ringer buffer solution was used as extracellular cryoprotective agent.

Conclusions: The selection of corneal freeze-thawing protocol as well as the optimal type and concentration of a cryoprotective agent allows the cryostorage of porcine corneal tissues with suitable TEER properties (cryocornea).     

Lyophilized insulin nanoparticles prepared from quaternized N-aryl derivatives of chitosan as a new strategy for oral delivery of insulin: in vitro,ex vivo and in vivo characterizations

Objective: The purpose of this research was the development, in vitro, ex vivo and in vivo characterization of lyophilized insulin nanoparticles prepared from quaternized N-aryl derivatives of chitosan.

Methods: Insulin nanoparticles were prepared from methylated N-(4-N,N-dimethylaminobenzyl), methylated N-(4 pyridinyl) and methylated N-(benzyl). Insulin nanoparticles containing non-modified chitosan and also trimethyl chiotsan (TMC) were also prepared as control. The effects of the freeze-drying process on physico-chemical properties of nanoparticles were investigated. The release of insulin from the nanoparticles was studied in vitro. The mechanism of the release of insulin from different types of nanoparticles was determined using curve fitting. The secondary structure of the insulin released from the nanoparticles was analyzed using circular dichroism and the cell cytotoxicity of nanoparticles on a Caco-2 cell line was determined. Ex vivo studies were performed on excised rat jejunum using Frantz diffusion cells. In vivo studies were performed on diabetic male Wistar rats and blood glucose level and insulin serum concentration were determined.

Results: Optimized nanoparticles with proper physico-chemical properties were obtained. The lyophilization process was found to cause a decrease in zeta potential and an increase in PdI as well as and a decrease in entrapment efficiency (EE%) and loading efficiency (LE%) but conservation in size of nanoparticles. Atomic force microscopy (AFM) images showed non-aggregated, stable and spherical to sub-spherical nanoparticles. The in vitro release study revealed higher release rates for lyophilized compared to non-lyophilized nanoparticles. Cytotoxicity studies on Caco-2 cells revealed no significant cytotoxicity for prepared nanoparticles after 3-h post-incubation but did show the concentration-dependent cytotoxicity after 24?h. The percentage of cumulative insulin determined from ex vivo studies was significantly higher in nanoparticles prepared from quaternized aromatic derivatives of chitosan. In vivo data showed significantly higher insulin intestinal absorption in nanoparticles prepared from methylated N-(4-N, N-dimethylaminobenzyl) chitosan nanoparticles compared to trimethyl chitosan.

Conclusion: These data obtained demonstrated that as the result of optimized physico-chemical properties, drug release rate, cytotoxicity profile, ex vivo permeation enhancement and increased in vivo absorption, nanoparticles prepared from N-aryl derivatives of chitosan can be considered as valuable method for the oral delivery of insulin.     

Benchtop MRI for pharmacokinetic evaluation of two aqueous-based nano-scaled formulations of oleic acid stabilized magnetite nanocrystals

Background: The interplay between numerous factors, including the size, shape, coating, surface charge and composition of particles is known to affect the pharmacokinetics and biodistribution of superparamagnetic iron oxides (SPIOs). This makes understanding the role of each factor independently quite challenging.

Methods: In the present study, the in vivo magnetic resonance imaging (MRI), biodistribution and hepatic clearance evaluations of two SPIOs Formulations A and B developed from ～13.5?nm hydrophobic oleic acid stabilized monodisperse magnetite nanocrystals core and lipid-based amphiphilic stabilizers were performed using a prototype benchtop MR imager (22?MHz) and pulsed nuclear magnetic resonance (NMR) system (20?MHz), respectively. Formulation A was composed of mPEG-2000-DSPE and Formulation B was composed of Phospholipon-100H, sucrose ester M-1695 and Cremophor RH-40.

Results: The in vivo MRI investigations showed that both formulations were safe and effective as potential liver MR contrast agents with sustained liver contrast for at least seven days. In addition, ex vivo relaxometric investigations revealed that the formulations predominantly distribute to the liver and spleen following I.V. injection. The hepatic clearance kinetics determined based on the relaxometric quantification method indicated that both formulations exhibited a biphasic clearance process with a slow terminal clearance half-life of 11.5 and 12.7 days, respectively, for Formulations A and B.

Conclusions: The results of this study showed the potential biomedical applications of the investigated magnetopharmaceutical formulations as MRI contrast agents.