Ming Thau Sheu

Characterization of collagen gel solutions and collagen matrices for cell culture

The influence of glutaraldehyde as a crosslinking agent to increase the strength of collagen matrices for cell culture was examined in this study. Collagen solutions of 1% were treated with different concentrations (0-0.2%) of glutaraldehyde for 24 h. The viscoelasticity of the resulting collagen gel solution was measured using dynamic mechanical analysis (DMA), which demonstrated that all collagen gel solutions examined followed the same model pattern. The creep compliance model of Voigt-Kelvin satisfactorily described the change of viscoelasticity expressed by these collagen gel solutions. These crosslinked collagen gel solutions were freeze-dried to form a matrix with a thickness of about 0.2-0.3 mm. The break modulus of these collagen matrices measured by DMA revealed that the higher the degree of crosslinking. the higher the break modulus. The compatibility of fibroblasts isolated from nude mouse skin with these collagen matrices was found to be acceptable at a cell density of 3 x 10(5) cells/cm2 with no contraction, even when using a concentration of glutaraldehyde of up to 0.2%.

Fungal mycelia as the source of chitin and polysaccharides and their applications as skin substitutes.

A wovenable skin substitute (Sacchachitin) made from the residue of the fruiting body of Ganoderma tsugae was developed in this study. Chemical analysis revealed that the treated residue was a copolymer of beta-1,3-glucan (ca 60%) and N-acetylglucosamine (ca 40%) with a filamental structure of mycelia form, as demonstrated by both optical and scanning electron microscopy. The pulp-like white residue was then woven into thin, porous sheets 7.0 cm in diameter and 0.1-0.2 mm in thickness by filtration and lyophilized for use as a skin substitute. The wound area produced by dissecting rat skin of full thickness was found to almost completely heal on the side covered with Sacchachitin, whereas the control side covered with cotton gauge was around 6.0 cm2 on the 28th day. Furthermore, the wound healing effects of the chitin sheet from crab shell (Beschitin) and Sacchachitin were not found to be significantly different.

The Influence of Cosolvents on the In‐vitro Percutaneous Penetration of Diclofenac Sodium From a Gel System

Abstract— The influence of cosolvents on the in‐vitro percutaneous penetration of diclofenac sodium from a gel system was studied using a simplex lattice experimental design. Gel formulations were prepared by gelling the vehicle mixture of water, alcohol and propylene glycol with Carbomer 940. The synthetic membrane Durapore and hairless mouse skin were employed as barriers in a Franz‐type diffusion cell. It was found that the penetration through the synthetic membrane was well described by the Higuchi model. There existed a better inverse relationship between the penetration rate and the drug solubility in the respective vehicle. It appeared to be a membrane‐controlled mechanism when using hairless mouse skin as the barrier. The penetration rates in steady‐state for nine formulations were fitted to a polynomial equation based on this simplex lattice method. A three‐dimensional plot was constructed in this simplex surface studied. The maximal penetration rate was found to be from the vehicle containing water and ethanol in an exact volume ratio of 3:1 and the minimal penetration rate was observed from the vehicle containing water only.

Dissolution of diclofenac sodium from matrix tablets

Abstract Several parameters were studied for their effect on the dissolution of diclofenac sodium from Voltaren SR and hydroxypropylmethylcellulose (HPMC) based matrix tablets. The results indicate that addition of sodium or potassium chloride to the dissolution medium decreases the solubility of the drug and slows the dissolution rate, with the effect of sodium chloride being greater. The dissolution of the drug was studied in a medium which simulates the changing pH of the pathway followed by the drug as it passes from the stomach to the intestine. Dissolution was found to be inversely related to the rate at which the pH was changed. This may be caused by the deposition of an insoluble drug layer when contact is made with an acid medium. When higher viscosity grades of HPMC are used, slower release rates result. Drug release from Voltaren SR is best described as non-Fickian in an aqueous medium irrespective of whether salt is added; however, a zero-order dependence became evident in pH-changing media. The release of diclofenac sodium from the hydrophilic HPMC matrices follows a non-Fickian transport in all media.

Development of swelling/floating gastroretentive drug delivery system based on a combination of hydroxyethyl cellulose and sodium carboxymethyl cellulose for Losartan and its clinical relevance in healthy volunteers with CYP2C9 polymorphism.

The aim of this study was to develop an optimal gastroretentive drug delivery system (GRDDS) for administering Losartan. Additionally, the influence of optimized GRDDS on the bioavailability of Losartan and the formation extent of active metabolite E3174 by CYP2C9 polymorphism was investigated. Swellable and floatable GRDDS tablets combining hydroxyethyl cellulose (HEC), sodium carboxymethyl cellulose (NaCMC), and sodium bicarbonate were prepared at various compression pressures for evaluating swelling characteristics and floating capacity. Then Losartan was incorporated into optimized formulations for in vitro and in vivo characterizations. An appropriate ratio of HEC to NaCMC, addition of sodium bicarbonate, and compression at lower pressures resulted in the tablets floating over SGF for more than 16 h and swelling to 2 cm in diameter within 3h. The release patterns of Losartan from these tablets were pH-dependent. Results of the clinical trials showed that the mean bioavailability from GRD-A (HEC 91.67%, sodium bicarbonate 3.33% and Losartan 8.33%) was approximately 164%, relative to the immediate-release product (Cozaar). MRT and t(max) values were greater and C(max) values were lower for the GRDDS tablets compared with Cozaa. The lower bioavailability of Losartan in the CYP2C9*1/*1 subjects than CYP2C9*1/*3 subjects was found and could be due to the variety of enzymatic activity.

Influence of micelle solubilization by tocopheryl polyethylene glycol succinate (TPGS) on solubility enhancement and percutaneous penetration of estradiol

The effect of micellar solubilization on the enhancement of the solubility and percutaneous penetration of estradiol by the surface-active agent, tocopheryl polyethylene glycol succinate (TPGS) was characterized in this study. Results show that the solubility of estradiol was improved in the presence of TPGS through micellar solubilization. The critical micelle concentration (CMC) of TPGS increased with increasing ethanol concentration in the medium. With the flux corrected to the saturated level (J(corrected)) of the free form of estradiol, an increase in the alcohol content of the medium resulted in an increase in J(corrected) for all levels of TPGS examined. For the same level of alcohol content, an increase in the TPGS concentration mostly led to a small extent of decrease in J(corrected). However, the extent of decrease was more obvious in media containing more than 60% alcohol. We also confirmed that only an insignificant amount of TPGS was transported across the skin (below the detection limit of 2 microg/ml). Permeabilities (P(eff)), which describe the overall effects (DK/H) on the stratum corneum (SC), decreased with increasing TPGS concentration for media containing 0, 40, 60, and 80% alcohol, whereas they increased then decreased with increasing TPGS concentration for media containing 10 and 20% alcohol. The enhancement ratios based on P(eff) assuming that the medium contained 0% TPGS and alcohol as unity did not increase accordingly with increases in TPGS concentration at the same level as alcohol. Likewise, the enhancement ratios for the same level of TPGS increased with low alcohol content, but then decreased with increasing alcohol content. We concluded that micellar solubilization by TPGS was able to improve the solubility of estradiol, but it only had an insignificant influence on the skin. Interfacial coverage of TPGS with increasing TPGS concentration and hindrance of the partitioning of estradiol by the increasing alcohol content might play a role in influencing the permeability of estradiol.

The preparation and characterization of solid dispersions on pellets using a fluidized-bed system

In this study, solid dispersions of a poorly water-soluble drug, nifedipine, were prepared in hydroxypropylmethyl-cellulose (HPMC) on sugar spheres using a fluidized-bed coating system and characterized by differential scanning calorimetry (DSC) and dissolution measurements. A mixture of acetone and water (7:3) was found to be suitable as a spraying solution for simultaneous application of nifedipine and HPMC. DSC studies showed that the peak corresponding to the melting point of nifedipine became broadened when nifedipine was incorporated in a solid dispersion with HPMC at both ratios of 1:1 and 1:3. The results demonstrated that dissolution rates were fastest at the lowest nifedipine loading. Furthermore, the dissolution rate of nifedipine increased as more HPMC was added to the solid dispersions. The enhancement in the dissolution rate of nifedipine upon addition of Tween 80 in simulated gastric acid was attributed to the solubilizing effect of Tween 80 on nifedipine. Tween 80 had less influence when nifedipine was incorporated in solid dispersions containing higher fractions of HPMC.

Characterization of collagen isolation and application of collagen gel as a drug carrier

Abstract The effects of various purification conditions on the characteristics of telopepeptide-poor collagen fiber were compared. It was first confirmed that a 1:50 ratio of pepsin to skin was favorable for the digestion of porcine skin. The morphological characteristics observed by scanning electron microscopy (SEM) showed that fibril collagen, porous fibril membrane or dense membrane were all possibly formed depending on the digestion and freeze-drying media. Analysis by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and size-exclusion HPLC revealed that the collagen sample produced by digestion with pepsin in a pH 2.5 HCl solution to obtain telopeptide-poor collagen, and freeze-drying in 0.5 M acetic acid for the final step, resulted in less oligomers. Not only were its fibril characteristics qualitatively identical to those of standard collagen but also were easily hydrated suggesting that this procedure is an appropriate choice for both isolation and purification. Furthermore, it was demonstrated that gel formulations prepared by gelling the vehicle mixture of citric acid solution, ethanol and propylene glycol with 1% w/w of such a collagen sample was suitable for transdermal delivery.

DEVELOPMENT OF A LIQUID CHROMATOGRAPHIC METHOD FOR BIOANALYTICAL APPLICATIONS WITH SILDENAFIL

An improved HPLC method was developed for the determination of sildenafil concentrations in plasma. Analysis of sildenafil in plasma samples was simplified by utilizing a one-step liquid-liquid extraction after alkaline treatment of only 1 ml of plasma. The lower limit of quantitation was 10 ng/ml with a coefficient of variation of less than 20%. A linear range was found to exist from 10 to 1000 ng/ml. This HPLC method was validated with precisions (coefficient of variation, C.V.) for inter- and intra-day runs of 0.41-11.15% and 0.36-8.05%, respectively, and accuracies (the relative error of the mean, REM) for inter- and intra-day runs of -8.72-6.81% and 0.41-11.15%, respectively. A bioavailability study of sildenafil was performed on one normal healthy male volunteer by analyzing sildenafil plasma concentrations with this validated HPLC method. Results demonstrated that this HPLC method is appropriate for applications to bioavailability studies of sildenafil. In addition, an example of the influence of the co-administration of grapefruit juice on sildenafil pharmacokinetics in a healthy volunteer is presented.

Development of fungal mycelia as skin substitutes: Effects on wound healing and fibroblast

In this study, Sacchachitin membrane, prepared from the residue of the fruiting body of Ganoderma tsugae, was estimated for its effects on wound healing and the proliferation and migration of fibroblast cells. Two mirror-image wounds were made on the back of female guinea pigs by dissecting a 1.5 x 1.5 cm2 skin surface of full thickness. Sacchachitin membrane was placed randomly on one of the wounds and gauze or Beschitin on the other. Changes in the wound area were measured and photographed after a predetermined amount of time postoperatively. Histological examination of the wound and surrounding tissue was also performed to reveal any interaction of tissue with the dressing. The results showed that the wound area covered with Sacchachitin membrane was statistically smaller than that covering with gauze on day 10, whereas there was no significant difference in the wound size compared to that with Beschitin. Fibroblast cells from the dermis layer of guinea pigs were used. The number of fibroblast cells were counted on the predetermined days in the culture suspended with or without 0.01% w/v dressing materials. By layering on DMEM plates, the number of fibroblast cells migrating across the center line or outside of the central hole were counted after five days. All the results indicated that both 0.01% w/v of Sacchachitin and chitin significantly enhanced the proliferation and migration of fibroblast cells.