Hydrolyzable Ester Research Articles

Characterization of Novel Biodegradable Segmented Polyurethanes Prepared from Amino‐Acid Based Diisocyanate

AbstractSegmented polyurethanes (SPUs) which were expected to yield non‐toxic degradation products were synthesized from lysine‐based diisocyanate (LDI), 1,3‐propanediol (PDO), and polycaprolactone diol (PCL). SPUs were synthesized via a standard two‐step prepolymer method. The hard segment fraction was changed in order to tune the mechanical properties and the degradability. The aggregation structures of the SPUs were characterized by infrared spectroscopy and differential scanning calorimetry (DSC), temperature dependence of dynamic viscoelasticity, and small‐angle X‐ray scattering (SAXS). DSC and dynamic viscoelastic measurements revealed that the glass transition temperature (Tg) of the soft segment increased with an increase in the hard segment fraction. SAXS of SPUs revealed the aggregation states of hard and soft segments. Furthermore, the degradation of SPUs was investigated by exposing the polymers to a buffer solution at 310 K (pH=7.6). The degradation rate of SPUs increased with an increase in the soft segment fraction. This is because the soft segment has the hydrolyzable ester linkages and the ester linkages are susceptible to hydrolysis compared with the urethane linkages. Finally, an electrospray deposition method was used to fabricate biodegradable SPU micro‐fibers. FE‐SEM images showed that higher concentration of solution favored the formation of uniform biodegradable micro‐fibers without beads‐like structure.

Extended release of a novel antidepressant, venlafaxine, based on anionic polyamidoamine dendrimers and poly(ethylene glycol)‐containing semi‐interpenetrating networks

The multiple daily administration of venlafaxine, a novel third-generation antidepressant, was reduced based on polyamidoamine and polyethylene glycol (PEG)-containing semi-interpenetrating network (IPN), respectively. Venlafaxine was covalently linked to water-soluble G2.5 anionic polyamidoamine dendrimer via a hydrolyzable ester bond. Semi-IPN hydrogels were prepared by crosslinking acrylamide in the presence of PEG, and venlafaxine with predetermined amounts was loaded in situ. Dendrimer-venlafaxine conjugate and semi-IPNs were characterized by proton nuclear magnetic resonance and Fourier transform infrared, respectively. The effect of PEG concentration and molecular weight was studied and discussed for an optimal controlled release.

PH control for dyebath reuse in dyeing of polyamide with binary mixtures of acid dyes

Hydrolyzable organic esters were compared with ammonium sulphate as an acid donor for the pH control in dye-bath-reuse system of acid dyes mixtures. The ability of pH control, levelness, dyeing properties and reproducibility in dye-bath-reuse system were investigated comparatively. Hydrolyzable organic esters showed higher exhaustion and color yield than ammonium sulphate. In addition, hydrolyzable organic ester exhibited very low conductivity less than 0.5 mS, while ammonium sulphate give high conductivity. However, we could not observe any difference in levelness of dyed samples between two kinds of acid donors in laboratory scale dyeing. Over 10 cycles of reuse, hydrolyzable organic esters showed higher reproducibility than ammonium sulphate. No deterioration of the color fastness and levelness occurs over 10 cycles of reuse.

The dose-dependent antihypertensive actions of olmesartan medoxomil, an angiotensin II (AT-II) receptor blocker

P-242 Key Words: Olmesartan, Angiotensin Receptor Blocker, Antihypertensive

Novel biodegradable cholesterol-modified polyrotaxane hydrogels for cartilage regeneration

Cholesterol was introduced to a hydrolyzable polyrotaxane (PRx), not only to improve cell proliferation and glycosaminoglycan (GAG) production, but also to control the degradation rate of the hydrogels. The cholesterol was introduced to hydrolyzable PRx species by threading many α-cyclodextrins (α-CDs) on a poly(ethylene glycol) (PEG) chain having hydrolyzable ester linkages at the terminals; the PRx species were then cross-linked with other PEGs to prepare cholesterolmodified PRx hydrogels. The degree of cholesterol substitution was varied in the range of 1–25%. These hydrogels were examined to clarify the effect of cholesterol groups on mechanical properties, erosion time and chondrocyte proliferation. Highly porous biodegradable cholesterol-modified PRx hydrogels were fabricated using a combination of potassium hydrogen carbonate (as an effervescent salt) and citric acid. This fabrication process enabled the homogeneous expansion of pores within the polymer matrices, leading to well-interconnected macroporous hydrogels with a mean pore size of around 200–400 μm, ideal for high-density chondrocyte seeding. Time to complete degradation of the hydrogels was shortened by increasing the degree of substitution due to the aggregation of α-CDs through hydrophobic interaction of cholesterol groups. The presence of approx. 10% cholesterol improved the chondrocyte proliferation and GAG production. The modification of cholesterols to PRx is a good approach for creating new biodegradable hydrogels in terms of chondrocyte culture and controlling degradation time of the hydrogels.

Delivery of antimicrobials into parasites.

To eliminate apicomplexan parasites, inhibitory compounds must cross host cell, parasitophorous vacuole, and parasite membranes and cyst walls, making delivery challenging. Here, we show that short oligomers of arginine enter Toxoplasma gondii tachyzoites and encysted bradyzoites. Triclosan, which inhibits enoyl-ACP reductase (ENR), conjugated to arginine oligomers enters extracellular tachyzoites, host cells, tachyzoites inside parasitophorous vacuoles within host cells, extracellular bradyzoites, and bradyzoites within cysts. We identify, clone, and sequence T. gondii enr and produce and characterize enzymatically active, recombinant ENR. This enzyme has the requisite amino acids to bind triclosan. Triclosan released after conjugation to octaarginine via a readily hydrolyzable ester linkage inhibits ENR activity, tachyzoites in vitro, and tachyzoites in mice. Delivery of an inhibitor to a microorganism via conjugation to octaarginine provides an approach to transporting antimicrobials and other small molecules to sequestered parasites, a model system to characterize transport across multiple membrane barriers and structures, a widely applicable paradigm for treatment of active and encysted apicomplexan and other infections, and a generic proof of principle for a mechanism of medicine delivery.

Controlled‐release of drugs from cross‐linked polymers containing cubane as a crosslinking agent

AbstractTwo anti‐inflammatory drugs, indomethacin and aspirin together with cubane‐1, 4‐dicarboxylic acid (CDA) were covalently linked with 2‐hydroxyethyl methacrylate (HEMA). The drug‐linked HEMA (indomethacin‐linked HEMA) is abbreviated as HI, aspirin‐linked HEMA as HA and cubane‐1, 4‐dicarboxylic acid (CDA) linked to two HEMA group is the cross‐linking agent (CA). A difunctional spacer group was introduced between the drugs and acrylic moiety of the monomer through a hydrolyzable ester linkage. Free radical cross‐linking polymerization of the monomers with drug effect was carried out in dioxane solution at various CA ratios, using AIBN as initiator in the temperature range 60‐70°C. The compositions of the cross‐linked three‐dimensional polymers were determined by FT‐IR spectroscopy. The glass transition temperature (Tg) of the network polymers was determined calorimetrically. The hydrolysis of drug‐polymer conjugates was carried out in cellophane membrane dialysis bags containing an aqueous buffer solution (pH 8 and pH 1) at 37°C. Detection of the hydrolysis product by UV spectroscopy shows that the drugs were released by hydrolysis of the ester bond located between the drug and spacer group.

The Preparation and Properties of Some α-Acyloxy- and α-Carbamoyloxy-Phosphonothionates

New f -acyloxy and f -carbamoyloxy derivatives of dimethyl 2,2,2-trichloroethyl-phosphonothionate have been prepared, characterized, and screened for activity against free-living soil nematodes. Several of the more easily hydrolysable esters, and also the N -methylcarbamoyl derivative, were as active as the parent pesticide, dimethyl f -hydroxy-2,2,2-trichloroethylphosphonothionate, after an induction period during which the active species is assumed to be released in vivo . It is concluded that the 2,2,2-trichloroethyl group is essential for activity in compounds of these types and that the presence of the N -methylcarbamoyl group does not in itself confer activity.

Hydrogels having tubular α-cyclodextrin structure: effect of nano-tube structure on long alkyl chain partitions

New hydrogels having the tubular structure of a-cyclodextrins (a-CDs) crosslinked by poly(ethylene glycol) (PEG) (MT-PEG hydrogels) were prepared by using a hydrolyzable polyrotaxane. The hydrolyzable polyrotaxane, in which many a-CDs are threaded onto a PEG chain capped by a hydrolyzable ester moiety, was used to form a tubular structure in the hydrogel. After crosslinking with another PEG chain, the ester linkage in the polyrotaxane was hydrolyzed in 1N NaOH. This led to exposing hydrophobic cavity of a-CDs. The tubular-structured hydrogel incorporated sodium dodecyl sulfate (SDS) much faster than normally crosslinked a-CDs hydrogel (a-CD-PEG hydrogel). Furthermore, partition coefficient (K) of SDS to the MT-PEG hydrogel was two times larger than the a-CD-PEG hydrogel. These results suggest that the tubular structure of a-CDs, made from a template of the polyrotaxane, is much more attractive to include SDS in the a-CD cavities. Unsaturated fatty acids (palmitoleic acid (C16:1) and oleic acid (C18:1)) were also effectively incorporated into the tubular-structured hydrogel during 6 days. The K value of C16:1 was as the same order in magnitude as C18:1. Thus, the tubular structure of a-CDs was advantageous to incorporate long alkyl chains into the hydrophobic cavity of a-CDs in aqueous conditions.

Well‐defined star polylactides and their behavior in two‐dimensional chromatography

Let us consider an attempt to determine the actual structure and size of star-shaped macromolecules that are assumed, on the basis of the known chemistry, to have n arms. Such assumptions are usually made but seldom proved. Indeed, it is difficult if not impossible on the basis of the published methods to find out whether within starlike macromolecules prepared with the intention of having n arms there are macromolecules with n x (where x 1, 2, 3, . . . , n 1) arms. Methods such as size exclusion chromatography (SEC) would merely show a broader “molar masses” distribution for such structural imperfections. In certain specific instances, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry could help and shows the corresponding populations. Another approach is possible when arms can be detached from the core. Then measurements of the absolute molar masses of the original starshaped polymer and arms, after their detachment, could provide unambiguous information. This method was used, for instance, to analyze star-shaped polytetrahydrofuran, attached with easily hydrolyzable ester bonds to the aromatic core. Star-shaped polymers having a different number of arms also have a different number of end groups. Because liquid chromatography at the critical point of adsorption (LC-CC) separates macromolecules according to the structure of the end groups, we decided to use two-dimensional (2D) chromatography with LC-CC as the first dimension. We expected to suppress the influence of the differences in molar masses and to separate macromolecules according to the number of end groups. In the second dimension, SEC provides information on molar masses. The method applying critical conditions is based on the use of solvent (usually a mixture of solvents) at a specific temperature when enthalpic and entropic factors of macromolecules stationary-phase solvent interactions cancel each other. 4 Macromolecules at these conditions are separated exclusively with regard to the end groups and independently of the molar mass. Interaction of the end groups with the column packing is responsible for discrimination between macromolecules bearing different end groups. This method was confirmed to be successful for a number of systems and allowed, for instance, the determination of 13 different combinations of end groups in the bisphenol A epoxy resins. In a short overview on 2D chromatography one can find a monograph published by one of us. Correspondence to: S. Penczek (E-mail: spenczek@bilbo. cbmm.lodz.pl)

Cationic lipids derived from glycine betaine promote efficient and non‐toxic gene transfection in cultured hepatocytes

The low efficiency and toxicity of transfection in a primary culture of hepatocytes using cationic lipids remains a limiting step to the study of gene function and the setting up of non-viral gene therapy. A novel class of cationic lipids (GBs) derived from natural glycine betaine compounds covalently linked to acyl chains by enzymatically hydrolysable peptide and ester bonds, a structure designed to reduce cytotoxicity, was used to improve transfection efficiency in a primary culture of rat hepatocytes. The relationship between lipid structure, lipoplex formulation and transfection efficiency was studied using six GBs (12-14-16, 22-24-26) varying in their spacer and acyl chains. GB12, characterized by short [(CH(2))(10)] acyl chains and spacer, allowed plasmid uptake in all cells and reporter gene expression in up to 40% of hepatocytes with a low cytotoxicity, a much higher efficiency compared with transfections using other reagents including Fugene6 and Lipofectin. We also showed that numerous cells accumulated high amounts of plasmids demonstrating that GB12 promoted a very efficient DNA transfer through plasma membrane leading to an increase in nuclear plasmid translocation, allowing a much higher gene expression. Moreover, GB12-transfected hepatocytes survived to injection in normal livers and were found to express the LacZ reporter gene. The non-toxic GB12 formulation is a powerful vehicle for plasmid delivery in cultured hepatocytes with relevance in liver gene therapy.

PH control for dyeing polyamide in a reused acid dyebath

The pH control that can be achieved in a reused dyebath was investigated. The study involved a phosphate buffer system and four pH sliding systems, including ammonium sulphate and three hydrolysable organic esters. Instead of discharging the dyebath after each dyeing cycle, the residual dyebath was analysed using a UV‐Vis spectrophotometer and reconstituted to the required concentration of dyes, auxiliaries and acid donors. The dyebath was reused for 10 cycles and colour reproducibility, levelness and fastness of the dyed samples were measured after each recycling. In comparison with ammonium sulphate and sodium dihydrogen phosphate, hydrolysable organic esters gave a stable and effective pH shifting in the dyebath reuse system. In addition, hydrolysable organic esters resulted in a very low amount of salt in residual dyebath. No deterioration in colour fastness of the dyed fabrics over 10 cycles of dyebath reuse was evident.

Model prodrugs designed for the intestinal peptide transporter. A synthetic approach for coupling of hydroxy-containing compounds to dipeptides

The human peptide transporter, hPepT1, situated in the small intestine, may be exploited to increase absorption of drugs or model drugs by attaching them to a dipeptide, which is recognised by hPepT1. A synthetic protocol for this kind of model prodrugs was developed, in which model drugs containing a hydroxy group were attached to enzymatically stable dipeptides by hydrolysable ester linkages. Furthermore, a number of benzyl alcohols with various substituents in the 4-position of the phenyl ring were coupled to d-Asp–Ala and d-Glu–Ala. Ideally, a prodrug should be stable in the upper small intestine and be converted to the parent drug during or after transport into the blood circulation. Therefore, we investigated the influence of the electronegativity of the substituent in the 4-position of the phenyl ring on stability in aqueous solution at pH 6.0 and 7.4, corresponding to pH in jejunum and blood, respectively. In addition, the influence of the electronegativity of the substituent on stability upon storage was examined. Model prodrugs containing electron donating substituents in the 4-position of the phenyl ring decomposed upon storage, while model prodrugs containing no substituents or electron withdrawing substituents in the 4-position were stable. In aqueous solution (pH 6.0 and 7.4), electron withdrawing substituents in the 4-position decreased the half-life of the model prodrug. These data provide important information on stability of this kind of model prodrugs upon storage and under aqueous conditions. The results may be applied in the rational design of oligopeptide ester prodrugs to obtain prodrugs, which are stable upon storage and have an optimal release profile of the drug.

PH control in the dyeing of polyamide with acid dyes

In this study, we report a number of pH control systems for the dyeing of polyamide fibres in a closed dyeing procedure. These include a phosphate buffer system and four pH sliding systems (ammonium sulphate and three hydrolysable organic esters). The dyeing properties of these systems are compared with their ability to control the pH. In comparison with sodium dihydrogen phosphate and ammonium sulphate, hydrolysable organic esters showed a much more effective pH sliding and, consequently, showed higher dyebath exhaustion and colour yields. They also showed very low conductivity, typically less than 0.5 mS. However, no difference in the levelness of the dyed samples among the five acid donors studied was observed and no difference in colour fastness of the dyed fabrics under various pH control systems was detected.

Regulation of Controlled Release of Ibuprofen from Crosslinked Polymers Containing Cubane as a New Crosslinking Agent

The nonsteroidal anti-inflammatory drug 2-(4-isobutylphenyl)-propionic acid (ibuprofen) and cubane-1,4-dicarboxylic acid (CDA) were covalently linked with 2-hydroxyethyl methacrylate (HEMA). The drug-linked HEMA (ibuprofen-linked HEMA) abbreviated as HI and cubane-1,4-dicarboxylic acid (CDA) linked to two HEMA groups as CA. A difunctional spacer group was introduced between the drug and acrylic moiety of the monomer through a hydrolyzable ester linkage. Free radical crosslinking polymerization of the monomer HI with the various ratios CA as crosslinking agent were carried out in DMF, DMSO or dioxane solution, using AIBN as initiator at the temperature range of 60-70°C. The structure of the monomers was confirmed by FTIR, 1 H NMR and 13 C NMR spectroscopy. The compositions of the crosslinked three-dimensional polymers were determined by FTIR spectroscopy. Glass transition temperature (T g ) of the network polymers was determined calorimetrically. The hydrolysis of drug-polymer conjugates was carried out in cellophane membrane dialysis bags containing an aqueous buffer solution (pH 8 and pH 1) at 37°C. Detection of the hydrolysis product by UV spectroscopy shows that the drug (ibuprofen) was released by the hydrolysis of the ester bond located between the drug and spacer group.

The role of intraorganellar Ca(2+) in late endosome-lysosome heterotypic fusion and in the reformation of lysosomes from hybrid organelles.

We have investigated the requirement for Ca(2+) in the fusion and content mixing of rat hepatocyte late endosomes and lysosomes in a cell-free system. Fusion to form hybrid organelles was inhibited by 1,2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid (BAPTA), but not by EGTA, and this inhibition was reversed by adding additional Ca(2+). Fusion was also inhibited by methyl ester of EGTA (EGTA-AM), a membrane permeable, hydrolyzable ester of EGTA, and pretreatment of organelles with EGTA-AM showed that the chelation of lumenal Ca(2+) reduced the amount of fusion. The requirement for Ca(2+) for fusion was a later event than the requirement for a rab protein since the system became resistant to inhibition by GDP dissociation inhibitor at earlier times than it became resistant to BAPTA. We have developed a cell-free assay to study the reformation of lysosomes from late endosome-lysosome hybrid organelles that were isolated from the rat liver. The recovery of electron dense lysosomes was shown to require ATP and was inhibited by bafilomycin and EGTA-AM. The data support a model in which endocytosed Ca(2+) plays a role in the fusion of late endosomes and lysosomes, the reformation of lysosomes, and the dynamic equilibrium of organelles in the late endocytic pathway.

Formation of bound residues during microbial degradation of [14C]anthracene in soil.

Carbon partitioning and residue formation during microbial degradation of polycyclic aromatic hydrocarbons (PAH) in soil and soil-compost mixtures were examined by using [14C]anthracenes labeled at different positions. In native soil 43.8% of [9-14C]anthracene was mineralized by the autochthonous microflora and 45.4% was transformed into bound residues within 176 days. Addition of compost increased the metabolism (67.2% of the anthracene was mineralized) and decreased the residue formation (20. 7% of the anthracene was transformed). Thus, the higher organic carbon content after compost was added did not increase the level of residue formation. [14C]anthracene labeled at position 1,2,3,4,4a,5a was metabolized more rapidly and resulted in formation of higher levels of residues (28.5%) by the soil-compost mixture than [14C]anthracene radiolabeled at position C-9 (20.7%). Two phases of residue formation were observed in the experiments. In the first phase the original compound was sequestered in the soil, as indicated by its limited extractability. In the second phase metabolites were incorporated into humic substances after microbial degradation of the PAH (biogenic residue formation). PAH metabolites undergo oxidative coupling to phenolic compounds to form nonhydrolyzable humic substance-like macromolecules. We found indications that monomeric educts are coupled by C-C- or either bonds. Hydrolyzable ester bonds or sorption of the parent compounds plays a minor role in residue formation. Moreover, experiments performed with 14CO2 revealed that residues may arise from CO2 in the soil in amounts typical for anthracene biodegradation. The extent of residue formation depends on the metabolic capacity of the soil microflora and the characteristics of the soil. The position of the 14C label is another important factor which controls mineralization and residue formation from metabolized compounds.

Prodrugs for systemic bioreductive activation-independent delivery of phyllohydroquinone, an active form of phylloquinone (vitamin K1). 1: Preparation and in vitro evaluation.

With the aim of overcoming the delivery problems (water-solubility and bioreductive activation problems) of phyllohydroquinone (PKH), an active form of phylloquinone (PK, vitamin K1), the N,N-dimethylglycine esters of phyllohydroquinone (1-mono, 1; 4-mono, 2; and 1,4-bis, 3) have been synthesized and assessed in vitro as a prodrug for the systemic bioreductive activation-independent delivery of PKH. The hydrochloride salts of the esters were found to be quite soluble in water. Hydrolysis of the esters in rat liver S9 fraction, rat plasma and phosphate buffer, pH 7.4, at 37 degrees C, was kinetically studied in the presence and absence of an esterase inhibitor. The hydrolysis was catalyzed by esterases located in rat liver and rat plasma and quantitatively yielded PKH. The enzymatic cleavage and the vitamin K-dependent carboxylation activity of the esters in the rat liver microsome preparation at pH 7.2 and 25 degrees C were studied. The regeneration of PKH from the esters was catalyzed by carboxylesterases located in the rat liver microsome, and the order was as follows: 1 > 3 > 2. The carboxylation was stimulated by selected ester 1 in the absence of dithiothreitol, an activator of the vitamin K cycle. The carboxylation activity of 1 was strongly inhibited in the presence of eserine, a carboxylesterase inhibitor. Compound 1 could also stimulate carboxylase under warfarin-poisoning conditions, where the vitamin K cycle was strongly inhibited. These results indicated that these highly water-soluble and liver-esterase hydrolyzable ester derivatives of PKH are potential candidates for parenteral prodrugs which can thus achieve the systemic bioreductive activation-independent delivery of PKH.

Chemical modification of hydroxyl functions: Introduction of hydrolyzable ester function and bactericidal quaternary ammonium groups

Polymers carrying a hydrolyzable ester function and bactericidal quaternary ammonium salts were successfully synthesized in 2 steps. The first one was the modification of hydroxyl functions of poly(vinyl alcohol) by chloroacetic anhydride. The structure of synthesized polymers was confirmed by infrared (IR), 1H-, and 13C- nuclear magnetic resonance (NMR). The kinetic results were consistent with a 1-order reaction, and the activation energy in the case of total modification was found to be 16.8 kJ mol−1. The second step was the quaternization of the pendant chlorine atom with a long alkyl chain or aromatic tertiary amines. The percentage of grafting was almost total, except with 3-diethylaminophenol, probably due to steric hindrance. The thermal degradation of polymers was studied. Side chains carrying quaternary ammonium salts begin to degrade at 168°C with the emission of chloride and amine compounds. At about 250°C, acetic and chloroacetic acid are evolved due to acetate and chloroacetate side chains. © 1998 John Wiley & Sons, Inc. J Appl Polym Sci 70: 2657–2666, 1998

D - myo -Inositol 1,4,5,6-tetrakisphosphate produced in human intestinal epithelial cells in response to Salmonella invasion inhibits phosphoinositide 3-kinase signaling pathways

Several inositol-containing compounds play key roles in receptor-mediated cell signaling events. Here, we describe a function for a specific inositol polyphosphate, D-myo-inositol 1,4,5,6-tetrakisphosphate [Ins(1,4,5,6)P4], that is produced acutely in response to a receptor-independent process. Thus, infection of intestinal epithelial cells with the enteric pathogen Salmonella, but not with other invasive bacteria, induced a multifold increase in Ins(1,4,5,6)P4 levels. To define a specific function of Ins(1,4,5,6)P4, a membrane-permeant, hydrolyzable ester was used to deliver it to the intracellular compartment, where it antagonized epidermal growth factor (EGF)-induced inhibition of calcium-mediated chloride (Cl-) secretion (CaMCS) in intestinal epithelia. This EGF function is likely mediated through a phosphoinositide 3-kinase (PtdIns3K)-dependent mechanism because the EGF effects are abolished by wortmannin, and three different membrane-permeant esters of the PtdIns3K product phosphatidylinositol 3,4,5-trisphosphate mimicked the EGF effect on CaMCS. We further demonstrate that Ins(1,4,5,6)P4 antagonized EGF signaling downstream of PtdIns3K because Ins(1,4,5, 6)P4 interfered with the PtdInsP3 effect on CaMCS without affecting PtdIns3K activity. Thus, elevation of Ins(1,4,5,6)P4 in Salmonella-infected epithelia may promote Cl- flux by antagonizing EGF inhibition mediated through PtdIns3K and PtdInsP3.