Thiolated Polymers of Chitosan

With the same aim as described in the last paragraph, chitosan has been chemically modified by covalent binding of sulfur containing moieties. To date, different thiolated chitosan have been synthesized: chitosan-thioglycolic acid conjugate (Fig. 6.3e), chitosan-cysteine conjugates (Bernkop-Schnurch and Hopf, 2001; Kast and Bernkop-Schnurch, 2001; Hornof et al., 2003), chitosan-cystein conjugates (Bernkop-Schnurch et al., 1999), and chitosan-4-thio-butyl-amide (chitosan-TBA) conjugates (Fig. 6.3f) (Bernkop-Schnurch et al., 2003). These thiolated chitosans have numerous advantageous features in comparison to unmodified chitosan, such as significantly improved mucoadhesive properties and permeation enhancing properties.

The strong cohesive properties of thiolated chitosans make them highly suitable excipients for controlled drug release dosage forms (Bernkop-Schnurch et al., 2003; Kast and Bernkop-Schnurch, 2001). Moreover, solutions of thiolated chitosans display in situ gelling properties at physiological pH values, which make them suitable for novel application systems to the eye (Bernkop-Schnurch et al., 2004).

The improved mucoadhesive properties of thiolated chitosans were explained by the formation of covalent bonds between thiol groups of the polymer and cysteine-rich subdomains of glycoporteins in the mucus layer (Leitner et al., 2003b). These covalent bonds are supposed to be stronger than noncovalent bonds, such as ionic interactions of chitosan with nonionic substructures as sialic acid moieties of the mucus layer. This theory was supported by the results of tensile studies with tablets of thiolated chitosan, which demonstrated a positive correlation between the degree of modification with thiol bearing moieties and the adhesive properties of the polymer (Kast and Bernkop-Schnurch, 2001; Roldo et al., 2004). These findings were confirmed by another in vitro mucoadhesion system, where the time of adhesion of tablets on intestinal mucosa was determined. The contact time of the thiolated chitosan derivatives increased with increasing amounts of immobilized thiol groups (Kast and Bernkop-Schnurch, 2001; Bernkop-Schnurch et al., 2003). With chitosan-thioglycolic acid conjugates a 5-to 10-fold increase in mucoadhesion in comparison to unmodified chitosan was achieved. Solid Dosage Form Design Based on TMC and Thiolated Polymers and Their In Vivo Evaluation

As in the sections described above about TMC, this chitosan derivative has only been administered as a gel formulation or solution. However, the impracticability of administering a polymer solution intraduodenally with the peptide dispersed or dissolved in it, as well as the fact that most peptides are unstable in the presence of an aqueous milieu, has led to the need for a solid oral dosage form in which TMC can be administered together with peptide drugs. To optimally make use of the absorption enhancing properties of TMC in a solid dosage form, the polymer should be able to dissolve rapidly and then be allowed to spread over a wide area of the epithelium in the small intestine. The opening of the tight junctions is a time-dependent process and it is therefore necessary that most of the TMC should be released from the dosage form prior to the release of the peptide drug. The site at which the peptide is released should coincide with the side where the TMC is opening the paracellular pathway for maximum paracellular absorption of the peptide drug (Van der Merwe et al., 2004a).

Minitablets with a diameter of 2-3 mm and granule formulation were developed as solid oral dosage form for the delivery of TMC and the peptide drug desmopressin (1-(3-mercaptopropionic acid)-8-D-arginine vasopressin monoacetate; DDAVP). Both the developed minitablet and granule formulations showed an initial burst release of TMC with a delayed release for DDAVP. Maximum release of TMC was in the order of 50% for all formulations, which is acceptable considering the high molecular weight of the polymer. Domestic pigs were used for the evaluation of the developed minitablet and granule formulations. However, the somatostatin analogue, octreotide, was used in this study and therefore the formulations were slightly adapted to give the same release profiles (Van der Merwe et al., 2004b). The delivery systems were filled in 000 capsules and enteric coated and applied with a special designed applicator into the stomach of the pigs.

Statistical analysis showed no significant difference between the absolute bioavailabilities for the different formulations administered via the peroral route. The average bioavailabilities for the negative control, minitablet formulation, granule formulation, and TMC/octreotide solution were, respectively, 0.9 ± 0.5%, 1.0 ± 1.5%, 1.4 ± 0.5%, and 0.5 ± 0.2%. The combination of the gelatin, the enteric coating, and the sticky properties of TMC might have resulted in a delay of release of both the octreotide and the TMC, resulting in the unsatisfactory absorption enhancement with the polymer.

A similar low bioavailability with the peptide drug antide has been the result of an in vivo study of Bernkop-Schniirch and coworkers (2005). Antide and the permeation mediator glutathione were embedded in the thiolated polymer chitosan-4-thio-butylamidine conjugate and compressed to tablets. Because it turned out that antide was strongly degraded in the small intestine by elastase, a stomach targeted delivery system was designed. The absolute and relative bioavailability after oral application of the tablet delivery system to pigs were 1.1% and 3.2%, respectively.

In an earlier study, Guggi and coworkers (2002,2003) developed a solid dosage for the peroral delivery of salmon calcitonin to rats. Different drug carrier matrices, comprising chitosan-4-thio-butyl-amide (chitosan-TBA) conjugate as substantial polymeric excipient and containing equal amounts of salmon calcitonin and optionally the permeation mediator reduced glutathione (Bernkop-Schniirch and Scerbe-Saiko, 1998) were developed. In order to avoid an enzymatic degradation of the peptide drug in the gastrointestinal tract chitosan inhibitor conjugates were also added. All compounds were homogenized and directly compressed to tablets. To enteric-coated tablets targeted to the small intestine, a chitosan-BBI conjugate (Bowman-Birk inhibitor) (Guggi and Bernkop-Schnurch, 2003) and a chitosan-elastatinal conjugate were added (Bernkop-Schnurch and Scerbe-Saiko, 1998). The different tablets were given orally to rats and the plasma calcium levels were monitored as a function of time. Pharmacological efficacy was calculated on the basis of the area under the reduction in plasma calcium levels of the oral matrix tablets versus i.v. injection.

No significant effects were measured when calcitonin was given as a solution orally and also when chitosan was used as the main tablet ingredient due to its insolubility in pH values above 6.5. Only with tablets containing chitosan TBA conjugate as the main tablet excipient a moderate decrease of the calcium level of more than 5% for several hours have been reached. The increased absorption of the peptide, when embedded in a thiolated chitosan matrix, occurs due to the properties of this compound: the high stability and cohesiveness can provide a sustained release of the peptide, while the mucoadhesive features should lead to a prolonged residence time of the dosage form on the site of absorption (which still has to be demonstrated). Moreover, the combination of thiolated chitosan with the permeation mediator reduced glutothione, seems to have an impact on the bioresponse of orally given calcitonin. The significantly higher pharmacological efficacy of thiolated chitosan tablets containing glutathione in comparison to corresponding tablets without glutathione indicates that glutathione contributes additionally to the drug absorption enhancing process.

As pointed out before, it is extremely difficult to develop a peptide drug delivery system for peroral application intended to trigger paracellular uptake of the drug by locally widening of the tight junctions due to the deactivation of the mucoadhe-sive polymers by soluble mucins in the intestinal liquids before the delivery system reaches the mucosal epithelial tissue and could exert its polymer-mucus (and even preferred directly with the epithelial tissue sialic and sulfate groups) interactions. With small animals as mice and rats, some success with small particulate delivery systems has been achieved due to the short distances between delivery system and gut wall. However, translating the efficacy of such delivery systems to the gut of the pig or human being with much wider diameter of the gut lumen has not been successfully solved so far.

6.4 Conclusions

Most of the pharmaceutical excipients, which when used in appropriate amounts for the manufacturing of common drug delivery systems, are inert and do not show interactions with absorbing tissues. Some excipients such as low molecular weight surfactants may not be as neutral in their interactions with the mucosal membranes. in most cases, their intrinsic properties to form at a defined concentration micelles is used to solubilize poorly soluble drugs. When those solubilisates come in contact with the mucosal tissues of the gut they may interact with the phospholipid bilayer of this membrane and cause some perturbations of the phospholipid bilayer structure and form also micelles with those components or solubilize membrane proteins. Dependent on their concentration, this may lead locally to toxic effects. it is well known that the absorption enhancing effect is directly related to the surfactant concentration. STDHF seemed to be a promising absorption enhancer for the nasal application of insulin; however, chronic toxicity studies with ciliated chicken membranes showed that this compound was too toxic for chronic use and was therefore withdrawn from the market.

Cyclodextrins show basically the same effect as low molecular mass permeation enhancers: they are predominantly used for the solubility increase for poorly soluble drugs such as estrogens, progesterone, testosterone, and hydrocortisone (Duchene et al., 1999). There seem to be also some species differences: whereas, e.g., p-cyclodextrins showed very promising results in the nasal absorption of insulin of rats and sheep, no significant improvement could be obtained in the human nose. it can be concluded that until today low molecular weight surfactants and also cyclodextrins play a minor role as absorption enhancer for hydrophilic drugs.

With the advent of new biotechnological techniques endogenous compounds like insulin have become available at affordable prices. However, until, today the development of alternative dosage forms (for the nasal, buccal, peroral, rectal, vaginal, and ocular route) for the administration of those class iii drugs (high solubility/low permeability) according to the BCS (Amidon et al., 1995) could not keep pace with this development of endogenous peptides. Multifunctional high molecular weight polymers as polyacrylates and chitosan with its various derivatives show promising properties as specific penetration enhancers for the paracel-lular route of absorption of hydrophilic molecules with high enhancing potency of reversibly opening of the tight junctions and practically with no toxicity when used in normal doses. However, the physical properties (poor solubility and high viscosity and easy saturation of the mucoadhesive properties by soluble mucins in the intestinal liquids) make it very difficult to develop suitable dosage forms which are able to quickly swell in the intestinal gut fluids, to develop mucoadhe-sive properties, and finally reach the mucous linings of the (human) gut in a still mucoadhesive form. After adhesion to the gut mucus and widening of the tight junctions the peptide drug should be released in the desired controlled manner. Hence, the development of such dosage forms is still in its infancy, but there are promising perspectives that such delivery systems will be successfully developed further on and will see the light of the market.

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