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The aim of this study was to evaluate the viscoelastic properties of chitosan-thioglycolic acid conjugates with different amounts of thiol groups immobilized on the polymer. The modification of chitosan was achieved via the covalent attachment of thioglycolic acid mediated by a carbodiimide. Chitosan-thioglycolic acid conjugates displaying 120, 209 and 439 microM thiol groups per gram of polymer were synthesized. The rheological properties of the three different conjugates were investigated. The elastic properties of the gels were found to increase significantly at pH 5.5. After 6 h the elastic modulus G' of chitosan-TGA 120, chitosan-TGA 209 and chitosan-TGA 439 gels increased 7-, 32- and 168-fold, respectively. In parallel the formation of disulfide bonds was observed. Accordingly, proof of principle that chitosan modified by the introduction of thiol groups has in situ gelling properties due to the formation of inter- and intramolecular disulfide bonds at physiological pH values is provided. Based on their in situ gelling properties, chitosan-thioglycolic acid conjugates seem to be very promising new excipients for liquid or semisolid formulations which should stabilize themselves once applied on the site of drug delivery.

Purpose. It was the purpose of this study to develop a new oral drug delivery system for low molecular weight heparin (LMWH) providing an improved bioavailability and a prolonged therapeutic effect.

Purpose. To verify or refute the mechanism of permeation enhancement with thiolated polymers via GSH by the use of NaFlu as marker for the paracellular permeation.

The aim of this study was to improve mucoadhesive properties of chitosan by the covalent attachment of thiol moieties to this cationic polymer. Mediated by a carbodiimide, thioglycolic acid (TGA) was covalently attached to chitosan. This was achieved by the formation of amide bonds between the primary amino groups of the polymer and the carboxylic acid group of TGA. Dependent on the pH-value and the weight ratio of polymer to TGA during the coupling reaction the resulting thiolated polymers, the so-called thiomers, displayed 6.58, 9.88, 27.44, and 38.23 micromole thiol groups per gram polymer. Tensile studies carried out with these chitosan-TGA conjugates on freshly excised porcine intestinal mucosa demonstrated a 6.3-, 8.6-, 8.9-, and 10.3-fold increase in the total work of adhesion (TWA) compared to the unmodified polymer, respectively. In contrast, the combination of chitosan and free unconjugated TGA showed almost no mucoadhesion. These data were in good correlation with further results obtained by another mucoadhesion test demonstrating a prolonged residence time of thiolated chitosan on porcine mucosa. The swelling behavior of all conjugates was thereby exactly in the same range as for an unmodified polymer pretreated in the same way. Furthermore, it could be shown that chitosan-TGA conjugates are still biodegradable by the glycosidase lysozyme. According to these results. chitosan-TGA conjugates represent a promising tool for the development of mucoadhesive drug delivery systems.

Purpose. To develop and evaluate an oral delivery system for salmon calcitonin. Methods. 2-Iminothiolane was covalently bound to chitosan in order to improve the mucoadhesive and cohesive properties of the polymer. The resulting chitosan-TBA conjugate (chitosan-4-thiobutylamidine conjugate) was homogenized with salmon calcitonin, mannitol, and a chitosan-Bowman-Birk inhibitor conjugate and a chitosan-elastatinal conjugate (6.75 + 0.25 + 1 + 1 + 1). Optionally 0.5% (m/m) reduced glutathione, used as permeation mediator, was added. Each mixture was compressed to 2 mg microtablets and enteric coated with a polymethacrylate. Biofeedback studies were performed in rats by oral administration of the delivery system and determination of the decrease in plasma calcium level as a function of time. Results. Test formulations led to a significant (p < 0.005) decrease in the plasma calcium level of the dosed animals in comparison to control tablets being based on unmodified chitosan. The addition of glutathione in the tablets led to a further improvement in the oral bioavailability of salmon calcitonin with an earlier onset of action and a decrease in the calcium level of about 10% for at least 10 h. Conclusions. The combination of mucoadhesive thiolated chitosan, chitosan-enzyme-inhibitor conjugates and the permeation mediator glutathione seems to represent a promising strategy for the oral delivery of salmon calcitonin.

Thiolated polymers (= thiomers) in combination with reduced glutathione (GSH) were shown to improve the uptake of hydrophilic macromolecules from the GI tract. The mechanism responsible for this permeation enhancing effect seems to be based on the thiol groups of the polymer. These groups inhibit protein tyrosine phosphatase, being involved in the closing process of tight junctions, via a GSH-mediated mechanism. The strong permeation enhancing effect of various thiomer/GSH systems such as poly(acrylic acid)-cysteine/GSH or chitosan-4-thio-butylamidine (chitosan-TBA)/GSH could be shown via permeation studies on freshly excised intestinal mucosa in Ussing-type chambers. Furthermore, the efficacy of the system was also shown in vivo. By utilizing poly(acrylic acid)-cysteine/GSH as carrier matrix, an absolute oral bioavailability for low molecular weight heparin of 19.9 +/- 9.3% and a pharmacological efficacy--calculated on the basis of the areas under the reduction in serum glucose levels of the oral formulation versus subcutaneous (s.c.) injection-for orally given insulin of 7% could be achieved. The incorporation of salmon calcitonin in chitosan-TBA/GSH led on the other hand to a pharmacological efficacy based on the areas under the reduction in plasma calcium levels of the oral thiomer formulation versus intravenous (i.v.) injection of 1.3%. Because of this high efficacy (i), the possibility to combine thiomer/GSH systems with additional low molecular weight permeation enhancers acting in other ways (ii) and minimal toxicological risks as these polymers are not absorbed from the GI tract (iii), thiolated polymers represent a promising novel tool for the oral administration of hydrophilic macromolecules.

It was the purpose of this study to develop a new oral drug delivery system for low molecular weight heparin (LMWH) providing an improved bioavailability and a prolonged therapeutic effect.The permeation enhancing polycarbophil-cysteine conjugate (PCP-Cys) used in this study displayed 111.4 +/- 6.4 microM thiol groups per gram polymer. Permeation studies on freshly excised intestinal mucosa were performed in Ussing chambers demonstrating a 2-fold improved uptake of heparin as a result of the addition of 0.5% (w/v) PCP-Cys and the permeation mediator glutathione (GSH).Tablets containing PCP-Cys, GSH, and 279 IU of LMWH showed a sustained drug release over 4 h. To guarantee the swelling of the polymeric carrier matrix in the small intestine tablets were enteric coated. They were orally given to rats. For tablets being based on the thiomer/GSH system an absolute bioavailability of 19.9 +/- 9.3% (means +/- SD; n = 5) vs. intravenous injection could be achieved. whereas tablets comprising unmodified PCP did not lead to a significant (p0.01) heparin concentration in plasma. The permeation enhancing effect and subsequently a therapeutic heparin level was maintained for 24 h after a single dose.Because of the strong and prolonged lasting permeation enhancing effect of the thiomer/GSH system, the oral bioavailability of LMWH could be significantly improved. This new delivery system represents therefore a promising tool for the oral administration of heparin.

It was the aim of this study to evaluate chitosan-thioglycolic acid (chitosan-TGA) conjugate as scaffold material in tissue engineering. Chitosan was modified by the introduction of thiol groups. Briefly, TGA was introduced to chitosan via amide bond formation mediated by a carbodiimide. The properties of the resulting polymer were thereby altered in regard to water solubility, mucoadhesion, biodegradability and in situ gelling compared to the original polymer. Due to the immobilised thiol groups (240±30 μmol thiol groups per gram polymer), the viscosity of a 1.5% chitosan-TGA solution was improved 4.3-fold. This can be explained by the formation of disulphide bonds within this polymeric network. The conjugate was tested as scaffold material in form of a gel and sheets. Furthermore, the influence of the thiol groups on the viability of L-929 mouse fibroblasts was evaluated. It was shown that the L-929 mouse fibroblasts grew on both scaffolds despite the thiol groups, although the different surface conditions seemed to have an influence on the growing rate. Chitosan-TGA sheets seemed to be the more preferred layer. The improved in situ gelling may be important for ongoing developments. Direct injectable matrices at the site of tissue damage mimicking the tissue being restored may be a future trend on this topic. Hence, chitosan-TGA is a promising candidate as scaffold material in tissue engineering.

To verify or refute the mechanism of permeation enhancement with thiolated polymers via GSH by the use of NaFlu as marker for the paracellular permeation.The capability of 0.5% polycarbophil cysteine conjugate (PCP-Cys) to reduce 0.02% oxidized glutathione (GSSG) was evaluated via iodometric titration in aqueous solution. Glutathione in its reduced form (GSH; 0.1%-0.4%) and in combination with 0.5% PCP-Cys were tested for their permeation enhancement of sodium fluorescein (NaFlu) and fluorescence labeled bacitracin (bac-FITC) used as paracellular markers. Permeation studies across guinea pig duodenum were carried out in Ussing-type chambers. Opening of the tight junctions was additionally monitored by transepithelial electrical resistance (TEER) measurements.PCP-Cys (0.5%) was shown to reduce 22.0%+/-8.2% of GSSG (0.02%) to GSH in aqueous solution at pH 7.0 and 37 degrees C within 3 h. Permeation of NaFlu was shown to depend on the concentration of GSH. The apparent permeability coefficient (Papp) of NaFlu in buffer only was 4.98+/-0.5*10(-6), while in the presence of 0.4% GSH a Papp of 9.31+/-0.92*10(-6) was achieved, representing an enhancement ratio (R = Papp enhancer system/Papp control) of 1.86. The combination of GSH (0.4%) with PCP-Cys (0.5%) led to a significant (p0.001) improvement of R for NaFlu up to 2.93 accompanied by a decrease in TEER of 20.3%+/-1.4%. Incubation of bac-FITC with the same GSH/PCP-Cys combination led to an enhancement ratio of 2.06 within 3 h.GSH plays an important role in the opening of tight junctions of intestinal epithelia. It would appear that PCP-Cys is able to reduce GSSG, prolonging the concentration of GSH at the apical membrane, resulting in significantly enhanced paracellular transport.

It was the purpose of this study to design and evaluate a new bioadhesive vaginal drug delivery system for clotrimazole. Chitosan, a cationic biopolymer derived by deacetylation of chitin, was modified by the introduction of thioglycolic acid (TGA). The modification was achieved by utilising a carbodiimide to link the carboxylic acid moieties of TGA covalently to the primary amino groups of chitosan. The amount of added carbodiimide was thereby varied, resulting in chitosan-TGA conjugates A and B with 160 microM (=micromol) and 280 microM thiol groups per gram polymer, respectively. In order to characterise the new polymers the water uptake, the disintegration behaviour, the bioadhesive properties utilising the rotating cylinder method, as well as the release of clotrimazole from tablets based on these derivatives were studied. The water uptake and cohesive properties of vaginal tablets consisting of these new conjugates could be significantly (p0.05) improved. By adding clotrimazole the disintegration time of the conjugates was prolonged 1.6-fold for conjugate A and even 100-fold for conjugate B. Furthermore, the adhesion on vaginal mucosal tissue could be significantly improved. The addition of clotrimazole had also an impact on the adhesion time of chitosan-TGA conjugate B, which remained 26-times longer on vaginal mucosa than the corresponding unmodified polymer. The immobilisation of thiol groups guarantees a controlled drug release. Results of this study demonstrate that these new chitosan-TGA conjugates are very promising vehicles for the vaginal application of clotrimazole in treatment of mycotic infections.

In the present study, the features of two new thiolated polymers--the so-called thiomers--were investigated. Mediated by a carbodiimide cysteamine was covalently attached to sodium carboxymethylcellulose (Na-CMC) and neutralised polycarbophil (Na-PCP). Depending on the weight-ratio polymer to cysteamine during the coupling reaction, the resulting CMC-cysteamine conjugate and PCP-cysteamine conjugate showed in maximum 43 +/- 15 and 138 +/- 22 micromole thiol groups per g polymer (mean +/- S.D.; n=3), respectively, which were used for further characterisation. Tensile studies carried out with the CMC-cysteamine conjugate on freshly excised porcine intestinal mucosa displayed no significantly (P0.01) improved mucoadhesion, whereas, the mucoadhesive properties of the PCP-cysteamine conjugate were increased 2.5-fold compared with the unmodified polymer. The swelling behaviour of the CMC-cysteamine conjugate was uninfluenced by the covalent attachment of the sulfhydryl compound. In contrast the swelling behaviour of the PCP-cysteamine conjugate was improved significantly (P0.01) versus unmodified PCP. Furthermore, in aqueous solutions the disintegration time of tablets based on the CMC- and PCP-cysteamine conjugates was prolonged 1.5 and 3.2-fold, respectively, in comparison to tablets containing the corresponding unmodified polymers. According to these results, especially the PCP-cysteamine conjugate represents a promising new pharmaceutical excipient for various drug delivery systems.

The purpose of the present study was to design a novel carrier system based on a mucoadhesive polymer exhibiting improved properties concerning drug delivery to the vaginal mucosa. This was reached by the covalent attachment of L-cysteine to commercially available polyacrylic acid (Carbopol 974P). Mediated by a carbodiimide, increasing amounts of L-cysteine were covalently linked to the polymer. The resulting thiolated polyacrylic acid conjugates (NaC974P-Cys) displayed between 24.8 and 45.8 micromol thiol groups per gram of polymer. Because of the formation of intra- and/or intermolecular disulfide bonds, the viscosity of an aqueous thiolated polymer gel (3%) increased about 50% at pH 7.0 within 1 h. In oscillatory rheological measurements, it was shown that this increase in viscosity is mainly due to the increase in elasticity. Tensile studies carried out on freshly excised cow vagina demonstrated a significant (P

Because of its permeation enhancing effect (I), mucoadhesive properties (II) and the capability to provide a controlled release of incorporated drugs (III), chitosan represents an advantageous excipient in non-invasive peptide delivery. The use of chitosan for such delivery systems, however, is limited by the lack of inhibitory properties towards secreted and membrane bound enzymes. Due to the covalent attachment of enzyme inhibitors and/or complexing agents at the 2-position of this poly(β1-4- d -glucosamine), chitosans can be transformed into polymers that exhibit inhibitory properties. The immobilization of inhibitors such as antipain, chymostatin, elastatinal and Bowman–Birk inhibitor provide a protective effect towards pancreatic serine proteases, whereas covalently attached complexing agents such as EDTA guarantee the inactivation of membrane bound Zn-dependent peptidases as well as carboxypeptidase A and B. As the inhibition of these enzymes strongly improves the bioavailability of non-invasively administered peptide drugs, chemically modified chitosans represent promising auxiliary polymers.