Alamdar Hussain

Cyclodextrins in Nasal Delivery of Low-Molecular-Weight Heparins: In Vivo and in Vitro Studies

AbstractPurpose. To test the hypothesis that cyclodextrins reversibly enhance nasal absorption of low-molecular-weight heparins (LMWHs) and to investigate the mechanisms by which cyclodextrins enhance LMWH absorption via the nose. Methods. Absorption of LMWHs was studied by measuring plasma anti-factor Xa activity after nasal administration of various LMWH formulations to anesthetized rats. In vivo reversibility studies were performed to investigate if the effects of cyclodextrins are reversible and diminish with time. The absorption-enhancing mechanisms of cyclodextrins were investigated in cell culture model. The transport of enoxaparin and mannitol, changes in transepithelial electrical resistance (TEER), and distribution of tight junction protein ZO-1 were investigated. Results. Formulations containing 5% dimethyl-β-cyclodextrin (DMβCD) produced the highest increase in the bioavailability of LMWH preparations tested. In vivo reversibility studies with 5% DMβCD showed that the effect of the absorption enhancer at the site of administration diminished with time. Transport studies using 16HBE14o− cells demonstrated that the increase in the permeability of enoxaparin and mannitol, reduction in TEER, and the changes in the tight junction protein ZO-1 distribution produced by 5% DMβCD were much greater than those produced by β-cyclodextrin (βCD) or hydroxyl-propyl-β-cyclodextrin (HPβCD). Conclusions. Of the cyclodextrins tested, DMβCD was the most efficacious in enhancing absorption of LMWHs both in vivo and in vitro. The study also suggests that cyclodextrins enhance nasal drug absorption by opening of cell-cell tight junctions.

Evaluation of sucrose esters as alternative surfactants in microencapsulation of proteins by the solvent evaporation method

Sucrose esters (SE) are surfactants with potential pharmaceutical applications because of their low toxicity, biocompatibility, and excellent biodegradability. The objective of the study was to investigate SE as alternative surfactants in stabilizing emulsions for the preparation of protein-loaded microparticles. To achieve this goal, using bovine serum albumin as model protein and 75/25 poly(d,1-lactide-co-glycolide) as polymer carrier, we have investigated the influence of the following formulation variables on particle characteristics: (1) SE concentration from 0.01% to 1% (wt/vol), (2) hydrophile-lipophile balance (HLB) value of SE from 6 to 15, and (3) the nature of emulsion stabilizer. The formulations were characterized using ATR-FTIR spectroscopy, bicinchoninic acid protein assay, optical microscopy and SDS-PAGE. Results showed that at 0.05% (wt/vol) surfactant concentration, SE with HLB of 6 to 15 provided discrete and spherical microparticles with the highest encapsulation efficiency compared with controls polyvinyl alcohol (PVA) and poloxamer 188. These results may be explained by the difference in critical micelle concentration, diffusion, and partition coefficient among the tested surfactants. HLB values were consistent with SE spectral data. The protein molecular weight was preserved after the encapsulation process. The effective SE concentration was far less (20-to 200-fold) than that is usually required for PVA in microencapsulation of proteins. However, the encapsulation efficiency was relatively lower (∼13.5%). These preliminary results suggest that it may be desirable to optimize such formulations in vitro and in vivo for SE to be eventually used as altermative surfactants in the development of microparticulate systems for parenteral delivery of protein and gene medicines.

Pulmonary absorption of insulin mediated by tetradecyl-β-maltoside and dimethyl-β-cyclodextrin

AbstractPurpose. To determine if tetradecyl-β-maltoside (TDM) and dimethyl-β-cyclodextrin (DMβCD) enhance pulmonary absorption of insulin and to investigate if they do so by a reversible action on respiratory epithelium. Methods. Insulin formulated with saline, TDM, or DMβCD was administered intratracheally, after laryngoscopic visualization, as a spray to anesthetized rats. Reversibility studies were conducted in intact rats by administering insulin at different time points after administration of TDM or DMβCD. The pharmacodynamics and pharmacokinetics of insulin formulations were assessed by measuring plasma glucose and plasma insulin concentrations. Results. When insulin formulated with increasing concentrations (0.06-0.25%) of TDM or DMβCD were administered to anesthetized rats, there was a concentration-dependent decrease in plasma glucose and increase in plasma insulin concentrations. The relative bioavailability of insulin formulations containing TDM was higher (0.34-0.84%) than that of formulations containing DMβCD (0.19-0.48%). When insulin was administered 120 min after an agent was administered, in the reversibility study, no significant change in plasma glucose and insulin levels occurred compared to control. Conclusions. Both TDM and DMBCD enhance pulmonary absorption of insulin, with TDM being more efficacious than DMβCD in enhancing insulin absorption via pulmonary administration. The effects of TDM and DMβCD on respiratory epithelium are reversible, and the epithelium reestablishes its normal physiologic barrier 120 min after exposure to these agents.

Inhaled Insulin is Better Absorbed When Administered as a Dry Powder Compared to Solution in the Presence or Absence of Alkylglycosides

PurposeThis study was performed to investigate the safety of alkylglycosides administered via the respiratory route and to compare the pulmonary absorption profiles of insulin administered as dry powder inhaler and inhaler solution.MethodsThe safety of a series of alkylglycosides with varying alkyl chain lengths was studied by measuring the enzymatic activities in the bronchoalveolar lavage (BAL) fluid of rat lungs. Pulmonary formulations of insulin plus octylmaltoside were administered either as solution or lyophilized dry powder to anesthetized rats, and absorption of insulin was assessed by measuring plasma insulin and glucose levels. The physical characterization of the dry powder formulation was performed using scanning electron microscope (SEM) and Fourier transform infrared spectrophotometer (FTIR).ResultsThe BAL analysis showed that there was a gradual increase in the amount of lung injury markers released with the increase in the hydrophobic chain length of alkylglycosides. The pulmonary administration of lyophilized dry powder of insulin plus octylmaltoside or its solution counterpart showed that the bioavailability of powder formulation was about 2-fold higher than that of the formulation administered as solution. The SEM studies showed a subtle difference in the surface morphologies of formulation particles after lyophilization. FTIR data showed minor interactions between the peptide and excipients upon lyophilization.ConclusionsOf the alkylglycosides tested, octylmaltoside was least toxic in releasing lung injury markers. Octylmaltoside-based dry powder insulin formulations were more efficacious in enhancing pulmonary insulin absorption and reducing plasma glucose levels compared with the formulations administered as a solution.

Development of a HPLC method for the determination of cyclosporin-A in rat blood and plasma using naproxen as an internal standard

An isocratic reversed phase high-performance liquid chromatographic (HPLC) method with ultraviolet detection at 205 nm has been developed for the determination of cyclosporin-A (CyA) in rat blood and plasma. Naproxen was successfully used as an internal standard. Blood or plasma samples were pretreated by liquid-liquid extraction with diethyl ether. The ether extract was evaporated and the residue was reconstituted in acetonitrile-0.04 M monobasic potassium phosphate buffer (pH 2.5) solvent mixture. After washing with n-hexane, 30 microl of the reconstituted solution was injected into HPLC system. Good chromatographic separation between CyA and internal standard peaks was achieved by using a stainless steel analytical column packed with 4 microm Nova-Pak Phenyl material. The system was operated at 75 degrees C using a mobile phase consisting of acetonitrile-0.04 M monobasic potassium phosphate (pH 2.5) (65:35 v/v) at a flow rate of 1 ml/min. The calibration curve for CyA in rat blood was linear over the tested concentration range of 0.0033-0.0166 M with a correlation coefficient of 0.989. For rat plasma, the range of the concentrations tested were between 0.002 and 0.0166 M and showed linearity with a correlation coefficient of 0.953. The intra- and inter-run precision and accuracy results were 1.24-21.87 and 3.1-12.23%, respectively. The low volume of blood or plasma needed (200 microl), simplicity of the extraction process, short run time (5 min) and low injection volume (30 microl) make this method suitable for quick and routine analysis.

Indication of transcytotic movement of insulin across human bronchial epithelial cells

This study was performed to evaluate insulin permeability across human bronchial epithelial cell lines and investigate if insulin is transported via the paracellular or transcellular pathway. The movement of insulin across two bronchial epithelial cells, 16HBE14o- and Calu-3, was studied in the presence or absence of octylmaltoside. Mannitol and propanolol have been used as paracellular and transcellular marker, respectively, and transepithelial electrical resistance (TEER) was determined to investigate the tight junctional integrity of the monolayers. The possible endocytotic mechanism of insulin across these two cell lines was studied by confocal laser scanning microscopy after incubating the cells with fluorescent-labeled insulin. The TEER values for both cell monolayers were >400 Ω cm2 at confluency. There was a decrease in the TEER values when octylmaltoside was added to the apical side of transwells. Similarly, the apparent permeability coefficient (Papp) values of insulin, mannitol and propanolol, showed an increase with the rise in the concentration of octylmaltoside. In the absence of octylmaltoside, the Papp values for insulin and the markers were in the following order: propanolol > mannitol > insulin. Confocal microscopic studies revealed that the uptake of insulin by the bronchial epithelial cells perhaps occurs via translocation across the cell. The data presented in this study demonstrate that insulin perhaps moves across the bronchial cells via both paracellular and transcellular pathways.