Synthesis and in vitro characterization of entirely S-protected thiolated pectin for drug delivery

The study was aimed to synthesize a thiolated polymer (thiomer) that is resistant to oxidation in solutions above pH 5. In order to protect a pectin–cysteine conjugate against premature oxidation, the thiomer was S-protected by a disulfide connected leaving group. Therefore, 2-mercaptonicotinic acid was first coupled to l-cysteine by a disulfide exchange reaction and the purified product was subsequently attached to pectin by a carbodiimide mediated amid bond formation. The obtained fully S-protected thiolated pectin was in vitro characterized with respect to co- and mucoadhesive properties and stability toward oxidation. The results indicated a 1.8-fold and 2.3-fold enhanced disintegration time at pH 6.8 of the S-protected thiolated pectin (Pec–Cys–MNA) compared to thiolated pectin (Pec–Cys) and unmodified pectin (Pec). Moreover, rheological measurements of polymer/mucus mixtures showed a 1.6-fold (compared to Pec–Cys) and 6.7-fold (compared to Pec) increased dynamic viscosity of Pec–Cys–MNA. On the other hand, in the presence of a strong oxidizing agent such as H₂O₂ (0.3% v/v), no increase in viscosity of Pec–Cys–MNA could be observed. A 6-month experiment also demonstrated the long-term stability of a liquid formulation based on Pec–Cys–MNA. Further investigations proved that the first time all thiol groups on a thiolated polymer could be protected owing to the novel synthesis. Accordingly, these features may help to develop thiomer based liquid or gel formulations targeting mucosal surfaces such as nasal, ocular or vaginal drug delivery systems.     

Synthesis of zinc-crosslinked thiolated alginic acid beads and their in vitro evaluation as potential enteric delivery system with folic acid as model drug

The aim of this study is to explore the potential of synthetic modifications of alginic acid as a method to enhance the stability of its complexes with divalent cations under physiological conditions. A fraction of algin's carboxylic acid moieties was substituted with thiol groups to different substitution degrees through conjugating alginate to cysteine to produce alginate-cysteine (AC) conjugates. Infrared spectrophotometry and iodometry were used to characterize the resulting polymeric conjugates in terms of structure and degree of substitution. Moreover, zinc ions were used to crosslink the resulting AC polymers. Folic acid loaded beads were prepared from Zinc-crosslinked AC polymers (AC-Zn) of different cysteine substitution degrees. The generated beads were then investigated in vitro for their capacity to modify folic acid release. AC-Zn polymeric beads resisted drug release under acidic conditions (pH 1.0). However, upon transfer to a phosphate buffer solution (pH 7.0) they released most of their contents almost immediately. This change in drug release behavior is most probably due to the sequestering of zinc cations by phosphate ions within the buffer solution to form insoluble chelates and, to a lesser extent, the ionization of the carboxylic acid and thiol moieties. Removal of zinc ions from the polymeric matrix seems to promote polymeric disintegration and subsequent drug release. A similar behavior is expected in vivo due to the presence of natural zinc sequestering agents in the intestinal fluids. AC-Zn polymers provided a novel approach for enteric drug delivery as drug release from these matrices complied with the USP specifications for enteric dosage forms.     

A novel poloxamers/hyaluronic acid in situ forming hydrogel for drug delivery: Rheological, mucoadhesive and in vitro release properties

The influence of hyaluronic acid (HA) on the gelation properties of poloxamers blends has been studied with the aim of engineering thermosensitive and mucoadhesive polymeric platforms for drug delivery. The gelation temperature (T(gel)), viscoelastic properties and mucoadhesive force of the systems were investigated and optimised by means of rheological analyses. Poloxamers micellar diameter was evaluated by Photon Correlation Spectroscopy (PCS). Moreover in order to explore the feasibility of these platforms for drug delivery, the optimised systems were loaded with acyclovir and its release properties studied in vitro. By formulating poloxamers/HA platforms, at specific concentrations, it was possible to obtain a thermoreversible gel with a T(gel) close to body temperature. The addition of HA did not hamper the self assembling process of poloxamers just delaying the gelation temperature of few Celsius degrees. Furthermore, HA presence led to a strong increase of the poloxamer rheological properties thus indicating possible HA interactions with micelles through secondary bonds, such as hydrogen ones, which reinforce the gel structure. These interactions could also explain PCS results which show, in systems containing HA, aggregates with hydrodynamic diameters much higher than those of poloxamer micelles. Mucoadhesion experiments showed a rheological synergism between poloxamers/HA gels and mucin dispersion which led to a change of the flow behaviour from a quite Newtonian one of the separate solutions to a pseudoplastic one of their mixture. In vitro release experiments indicated that the optimised platform was able to prolong and control acyclovir release for more than 6h.     

Synthesis and characterisation of mucoadhesive thiolated polymers

This study examined various factors influencing the mucoadhesive properties of thiolated polymers (thiomers), which are capable of forming covalent bonds with thiol sub-structures of the mucus glycoprotein. Mediated by a carbodiimide, L-cysteine was therefore covalently bound to polycarbophil (PCP) and to carboxymethylcellulose (CMC). The resulting polymer conjugates displayed 12.3 and 22.3 micromol thiol groups per gram, respectively. Whereas the swelling behaviour of tablets based on CMC was not markedly influenced by the immobilisation of cysteine, it was improved significantly (P<0.05) in case of PCP. Tensile studies carried out with the unmodified and thiolated polymers of pH 3, 5 and 7, respectively, revealed that only if the polymer displays a pH-value of 5, the total work of adhesion can be improved significantly due to the covalent attachment of thiol groups. These results were in good agreement with a new mucoadhesion test system described here taking also the cohesiveness of the delivery system into account. The results represent helpful basic information in order to improve the mucoadhesive properties of thiolated polymers.     

Thiomers: preparation and in vitro evaluation of a mucoadhesive nanoparticulate drug delivery system

It was the aim of this study to develop a mucoadhesive nanoparticulate delivery system. Nanoparticles were generated by in situ gellation of the thiomer chitosan-4-thiobutylamidine (chitosan-TBA) with tripolyphosphate (TPP) followed by stabilization via the formation of inter- and intrachain disulfide bonds by oxidation with H(2)O(2) in various concentrations. Afterwards TPP was removed by exhaustive dialysis at pH 1-2. Incorporation of the model compound fluorescein diacetate (FDA) was achieved by incubation of this fluorescence marker, dissolved in acetonitrile, with aqueous particle suspensions for 1h at room temperature. Mucoadhesion studies were performed on porcine intestinal mucosa. Results showed that the preparation method described above leads to nanoparticles of a mean diameter of 268+/-15 nm and a FDA load of 2%. Due to the removal of the anionic crosslinker TPP, the zeta potential of the nanoparticles was raised from 4+/-1 up to 19+/-2 mV without loosing stability of the nanoparticles. The more H(2)O(2) was added to the particles, the more inter- and intrachain disulfide bonds were formed. The more thiol groups were oxidized within the particles, however, the lower was the improvement in mucoadhesive properties. Nevertheless, even when 91% of all thiol groups on the nanoparticles were oxidized, their mucoadhesive properties were still twice as high as the mucoadhesive properties of unmodified nanoparticles. Thiolated chitosan nanoparticles show a two-fold higher zeta potential (I), improved stability (II) and more than doubled mucoadhesive properties (III) than corresponding unmodified chitosan nanoparticles. Therefore, they seem to be advantageous over ionically crosslinked chitosan nanoparticles.     

Mucoadhesive thiolated chitosans as platforms for oral controlled drug delivery: synthesis and in vitro evaluation.

The aim of the present study was to evaluate the influence of the degree of modification and the polymer chain length on the mucoadhesive properties and the swelling behavior of thiolated chitosan derivatives obtained via a simple one-step reaction between the polymer and 2-iminothiolane. The conjugates differing in molecular mass of the polymer backbone and in the amount of immobilized thiol groups were compressed into tablets. They were investigated for their mucoadhesive properties on freshly excised porcine mucosa via tensile studies and the rotating cylinder method. Moreover, the swelling behavior of these tablets in aqueous solutions was studied by a simple gravimetric method. The obtained results demonstrated that the total work of adhesion of chitosan-TBA (=4-thio-butyl-amidine) conjugates can be improved by an increasing number of covalently attached thiol groups; a 100-fold increase compared to unmodified chitosan was observed for a medium molecular mass chitosan-TBA conjugate exhibiting 264 microM thiol groups per gram polymer. Also, the polymer chain length had an influence on the mucoadhesive properties of the polymer. The medium molecular mass polymer displayed a fourfold improved adhesion on the rotating cylinder compared to the derivative of low molecular mass. These results contribute to the development of new delivery systems exhibiting improved mucoadhesive properties.     

Polymeric microspheres for drug delivery to the oral cavity: an in vitro evaluation of mucoadhesive potential

Polymeric microparticles were fabricated from Carbopol, polycarbophil, chitosan, or Gantrez using a "water-in-oil emulsification" solvent evaporation method. Mean particle sizes, as determined by laser diffraction, were in the range 23-38 microm. Electron microscopy revealed that all microparticles were spherical and of smooth surface morphology. In pH 7.0 phosphate buffered saline, the microspheres exhibited significantly increased swelling ratios and longer half-times of swelling than the corresponding powdered polymers. The relative merits of the potential usefulness of these microspheres as formulation tools for the enhanced retention of a therapeutic entity within the oral mucosa were evaluated by in vitro mucoadhesion tests. Tensile tests showed that all microspheres under consideration were capable of adhering to porcine esophageal mucosa, with particles prepared from the poly(acrylic acid)s exhibiting greater mucoadhesive strength than those constructed from chitosan or Gantrez. However, in elution experiments involving a challenge with artificial saliva, particles of chitosan or Gantrez were retained onto mucosal tissue for longer time periods than those assembled from the poly(acrylic acid)s.     

一种可用于黑色素瘤靶向递药的透明质酸纳米凝胶

黑色素瘤恶性程度高,且发病率逐年上升。本研究制备了一种能特异性靶向黑色素瘤的透明质酸纳米凝胶,将巯基化的透明质酸修饰于表面功能化的普朗尼克F127-TPGS混合胶束制备共价交联的纳米凝胶。通过测定粒径考察其体外稳定性;细胞毒性实验考察该载体材料对细胞的毒性作用;细胞摄取实验定量和定性考察B16F10黑色素瘤细胞对该纳米凝胶的摄取情况。结果显示,本研究制备了一种30 nm左右的小粒径纳米凝胶,该纳米凝胶对小鼠3T3成纤维细胞与小鼠黑色素瘤B16F10细胞均无明显细胞毒性作用,与低表达CD44受体的3T3细胞相比,高表达CD44受体的B16F10细胞的摄取效率显著增加(P<0.05)。     

Development and in vitro evaluation of a mucoadhesive vaginal delivery system for nystatin

The purpose of the present study was to design and evaluate a novel vaginal delivery system for nystatin based on mucoadhesive polymers. L-Cysteine and cysteamine, respectively, were covalently attached to poly(acrylic acid), and the two different thiolated polymers were evaluated in vitro regarding their swelling behavior, mucoadhesive properties and release behavior. Tablets comprising these thiolated polymers and nystatin demonstrated a high stability in vaginal fluid simulant pH 4.2 and an increase in weight by swelling whereas control tablets comprising unmodified poly(acrylic acid) disintegrated and dissolved. The mucoadhesion time of tablets on freshly excised bovine vaginal mucosa on a rotating cylinder and the total work of adhesion of gels and tablets increased significantly due to the formation of disulfide bonds between the thiolated polymer and cysteine rich subdomaines of the mucus layer. The drug nystatin was released more slowly out of thiomer tablets and gels than out of PAA control tablets and gels. Therefore these thiolated polymers are promising delivery systems for nystatin providing a prolonged residence time and a sustained drug release in vitro under physiological relevant conditions.     

Hyaluronic acid L-cysteine conjugate exhibits controlled-release potential for mucoadhesive drug delivery

Hyaluronic acid-L-cysteine conjugate, a novel thiolated polymer, was synthesized and characterized for mucoadhensive drug delivery. L-Cysteine was covalently attached to hyaluronic acid via the formation of an amide bond. Adhesion studies on the mucosa indicated a 4.82-fold increase in the adhesion force of the obtained conjugate (containing 210.58 micromol thiol groups per gram polymer) versus unmodified hyaluronic acid. The results of a peptidase inhibition study revealed that the inhibitory effect of hyaluronic acid toward trypsin and chymotrypsin was significantly improved compared to hyaluronic acid. Permeation studies utilizing a MDCK cell monolayer system demonstrated a sustained drug release. Based on these features the novel thiolated polymer might represent a promising multifunctional excipient for various drug delivery systems.     

An insight on hyaluronic acid in drug targeting and drug delivery

Hyaluronic acid (HA) has recently been studied for its use in drug delivery applications. Medically, HA is used as a surgical aid in ophthalmology. It also possesses therapeutic potential in the treatment of arthritis and wound healing. HA-binding receptors, CD44 and receptor for hyaluronan-mediated motility have attracted much enthusiasm, mainly because they are believed to be involved in cancer metastasis. This review unravels the role of HA in drug delivery and targeting. Designing of various novel drug delivery systems using HA as a biopolymer will also be reviewed in the present article.     

An insight on hyaluronic acid in drug targeting and drug delivery

Hyaluronic acid (HA) has recently been studied for its use in drug delivery applications. Medically, HA is used as a surgical aid in ophthalmology. It also possesses therapeutic potential in the treatment of arthritis and wound healing. HA-binding receptors, CD44 and receptor for hyaluronan-mediated motility have attracted much enthusiasm, mainly because they are believed to be involved in cancer metastasis. This review unravels the role of HA in drug delivery and targeting. Designing of various novel drug delivery systems using HA as a biopolymer will also be reviewed in the present article.     

In vitro evaluation of the mucoadhesive properties of polysaccharide-based nanoparticulate oral drug delivery systems.

Impedance quartz crystal microbalance (QCM) and surface plasmon resonance (SPR) measurements were performed in order to assess the mucoadhesive properties of hydrophobically modified (HM) derivatives of dextran (DEX), with an average molecular weight of 10,000 Da, and of hydroxypropylcellulose (HPC), with an average molecular weight of 80,000 Da. The measurements involved (1) treatment of a hydrophobic surface with bovine submaxillary gland mucin (BSM) under various pH conditions (2.0-8.0) and (2) treatment of the BSM layer with buffer solutions of the amphiphilic polysaccharides (pH 3.0 and 7.0). Control measurements were carried out with DEX, HPC, and chitosan (CH) used as a model mucoadhesive polymer. All HM-polysaccharides were shown to adsorb onto a BSM layer, the extent of adsorption increasing with increasing hydrophobicity of the samples. Under the same conditions, HPC and CH interacted with the BSM layer, but DEX showed no affinity to BSM. All the results suggest that HM-polysaccharide micellar systems have the potential of enhancing the bioavailability of poorly adsorbed drugs in peroral delivery.     

Application of hyaluronic acid as carriers in drug delivery

Hyaluronic acid has good biocompatibility, biodegradability, and nonimmunogenicity. In addition, it has the ability to recognize specific receptors that are overexpressed on the surface of tumor cells, and cancer drugs can be targeted to the tumor cells to better kill them. Therefore, hyaluronic acid has attracted much attention as drug delivery vehicle. Herein, the application of hyaluronic acid as carrier in drug delivery was analyzed and summarized in detail. It showed that hyaluronic acid would have broad prospects for drug delivery.     

Development and optimization of thiolated dendrimer as a viable mucoadhesive excipient for the controlled drug delivery: An acyclovir model formulation

In the present study we report the development of novel thiolated dendrimers for mucoadhesive drug delivery. The thiolated dendrimers were synthesized by conjugating PAMAM dendrimer (G3.5)with cysteamine at two different molar ratios, i.e. 1:30 (DCys1) and 1:60 (DCys2). The thiolated dendrimers were further encapsulated with acyclovir (DCys1Ac and DCys2Ac) and the conjugates were characterized for thiol content, drug loading, drug release, and mucoadhesive behavior. The thiolated dendrimer conjugates showed thiol content of 10.56±0.34 and 68.21±1.84 μM/mg of the conjugate for DCys1 and DCys2, respectively. The acyclovir loading was observed to be highest in dendrimer drug conjugate (DAc) compared to other DCys1Ac and DCys2Ac conjugates. The thiolated dendrimers showed sustained release of acyclovir and showed higher mucoadhesion. The in vitro mucoadhesive activity of DCys2Ac was 1.53 and 2.89 fold higher mucoadhesion compared to DCys1Ac and DAc, respectively. These results demonstrated the usefulness of thiolated dendrimers as a mucoadhesive carrier and represent a novel platform for drug delivery.From the Clinical EditorThis study demonstrates the utility of thiolated dendrimers as mucoadhesive carriers as reported in an acyclovir delivery model system.     

Methylprednisolone esters of hyaluronic acid in ophthalmic drug delivery: in vitro and in vivo release studies

Films and microspheres were prepared from various esters of hyaluronic acid. A model drug, methylprednisolone, was either physically incorporated into the polymer matrix or chemically bound to the polymer backbone through an ester linkage. In vitro release from films with covalently bound drug was much slower (t_50% = 71 h) than that for physically dispersed drug (t_50% = 2.5−17 h). Methylprednisolone concentrations in the tear fluid of New Zealand rabbits were measured after ocular application of drug (approx. 420 μg) in different dosage forms. When methylprenisolone was physically dispersed in the polymer matrix, in vivo drug release from matrices was slower than that observed in vitro. Compared with a suspension control, peak methylprednisolone concentrations in tear fluid were 9–14 times lower after administration of drug in polymer films and AUC_{0–8 h} values were 4–7 times higher. These results imply that hyaluronic acid ester preparations can increase the residence time of methylprednisolone in the tear fluid of rabbits.     

巯基化透明质酸对牛血清白蛋白体外释药机制的研究

目的 研究基于巯基化透明质酸骨架片的牛血清白蛋白的体外释放特性.方法 以牛血清白蛋白(BSA)为模型药物,通过测定不同巯基取代度、不同pH下骨架片体外释放度,并采用零级动力学方程、一级动力学方程、Higuchi方程、Ritger-Peppas方程、威布尔方程拟合,阐明BSA体外释放的机理.结果 巯基化透明质酸能延缓BSA的释放,并随着结合物巯基含量的增加,释放速率减慢;巯基化透明质酸骨架片在pH6.8磷酸盐缓冲液中的释放速率大于在pH1.7氯化钾-盐酸缓冲液中的释放速率.结论 基于巯基化透明质酸骨架片的BSA的体外释放机制为骨架溶蚀.     

Synthesis and evaluation of stearylated hyaluronic acid for the active delivery of liposomes to liver endothelial cells

Hyaluronic acid (HA) is a naturally-occurring ligand that can be useful for targeting liver endothelial cells. We describe herein the development of a new HA-lipid conjugate for the efficient delivery of liposomes to liver endothelial cells. When free HA coated cationic liposomes were injected into mice, their accumulation in the liver was significantly decreased depending on the content of free HA, while accumulation in the lung was not significantly changed. When cationic liposomes modified with HA-stearylamine (HA-SA conjugate) were injected in mice, liver accumulation was increased depending on the amount of HA-SA conjugate and accumulation in the lung was drastically reduced, compared to non-modified liposomes. Confocal imaging analyses showed that HA-SA modified liposomes were accumulated to a greater extent along with blood vessels than non-modified liposomes, suggesitng that HA-SA modified liposomes are distributed in endothelial cells in the liver. Collectively, these findings indicate that an HA-SA conjugate is a useful material that can be used to modify liposomes and for delivering bioactive liposomal cargoes to liver endothelial cells.     

Development, optimization and in vitro evaluation of alginate mucoadhesive microspheres of carvedilol for nasal delivery

The present research work was aimed at development and optimization of alginate mucoadhesive microspheres of carvedilol for nasal delivery to avoid first pass metabolism and to improve the therapeutic efficacy in the treatment of hypertension and angina pectoris. The microspheres were prepared by a water-in-oil (w/o) emulsification technique. A 2(3) factorial design was employed with drug : polymer ratio, calcium chloride concentration and cross-linking time as independent variables while particle size of the microspheres and in vitro mucoadhesion were the dependent variables. Regression analysis was performed to identify the best formulation conditions. Particle size was analysed by dynamic laser light diffraction technique and found to be in the range of 26.36-54.32 microm, which is favourable for intranasal absorption. The shape and surface characteristics were determined by scanning electron microscopy (SEM) which depicted the spherical nature and nearly smooth surfaces of the microspheres. The percentage encapsulation efficiency was found to be in the range between 36.62-56.18. In vitro mucoadhesion was performed by adhesion number using sheep nasal mucosa and was observed in a range from 69.25-85.28. Differential scanning calorimetry and X-ray diffraction results indicated a molecular level dispersion of carvedilol in the microspheres. In vitro release studies in pH 6.2 phosphate buffer indicated non-Fickian or anomalous type of transport for the release of carvedilol from the microspheres.