Combining spontaneous polymerization and click reactions for the synthesis of polymer brushes: a "grafting onto" approach.

Two novel benzofulvene monomers bearing propargyl or allyl groups have been synthesized by means of readily accessible reactions, and were found to polymerize spontaneously by solvent removal, in the apparent absence of catalysts or initiators, to give the corresponding polybenzofulvene derivatives bearing clickable propargyl or allyl moieties. The clickable propargyl and allyl groups were exploited in appropriate click reactions to develop a powerful and versatile "grafting onto" synthetic methodology for obtaining tailored polymer brushes.

Free-radical degradation of high-molar-mass hyaluronan induced by Weissberger's oxidative system: potential antioxidative effect of bucillamine.

OBJECTIVES: Hyaluronan (HA), one of the main components of extracellular matrix, is a glycosaminoglycan composed of repeating disaccharide units of N-acetyl-d-glucosamine and d-glucuronic acid linked by ß-(1→4) and ß-(1→3) glycoside bonds. High-molar-mass HA was used as a model for studying its oxidative degradation. In the present paper protective effects of bucillamine against the free-radical degradation of HA were investigated. The HA fragments generated were characterized as well. METHODS: To induce free-radical-mediated degradation of high-molar-mass HA under aerobic conditions, we applied Weissberger's oxidative system, comprising biogenic compounds in relevant pathophysiological concentrations, i.e. 100 µM ascorbate plus 1 µM Cu(II). Time-dependent decreases of dynamic viscosity of the HA solutions were recorded by rotational viscometry. Electron donor behaviors of bucillamine were studied by a standard ABTS test method and a chemiluminescence (CL) assay. Ability of incorporation of generated bucillamine thiyl radicals into the biopolymer was verified by Fourier-transform infrared spectroscopy (FT-IR) and size exclusion chromatography with a multi-angle light scattering photometer (SEC-MALS). RESULTS: Decrease of HA viscosity reflected HA degradation. The drug tested was applied in two arrangements: to prevent •OH radical generation (1) and ROO• type radicals propagation (2). Bucillamine, which acted as an efficient •H donor, is also a proper electron donor, as proved by ABTS and CL assays. FT-IR and SEC-MALS methods showed that the drug tested did not incorporate into the biopolymer chains. CONCLUSION: Bucillamine significantly protected high-molar-mass HA against free-radical degradation in vitro, and supposedly this positive action of the drug may be involved in its beneficial effect observed in clinical practice.

Biocompatibility of intraocular liquid tamponade agents: an update.

Intraocular liquids tamponade agents, such as perfluorocarbon liquids (PFCLs), semifluorinated alkanes (SFAs), silicone oils (SOs) and heavy silicone oils (HSOs), are a crucial intraoperative and/or postoperative tool in vitreoretinal surgery, in particular for the management of complex vitreoretinal diseases. However, their use is not without complications, which are potentially severe. Consequently, a growing interest has been devoted to the biocompatibility of these compounds and the adequacy of current regulations that should guarantee their safety. Obviously, an updated knowledge on research findings and potential risks associated to the use of intraocular liquid compounds is essential, not only for vitreoretinal surgeons, but also for any ophthalmologist involved in the management of patients receiving intraocular liquid tamponades. In light of this, the review provides a comprehensive characterisation of intraocular liquid tamponades, in terms of physical and chemical properties, current clinical use and possible complications. Moreover, this review focuses on the safety profile of these compounds, summarising the existing regulation and the available evidence on their biocompatibility.

Behavior and properties in aqueous solution of biopolymers isolated from urban refuse.

Acid soluble biopolymeric substances (SBP) were isolated from different urban biowastes comprised of a range of materials available from metropolitan areas. These biowastes provided products with a chemical nature and solubility properties changing over a wide range and, thus, allowed to assess the effect of the variability of the chemical nature on molecular conformation and surface activity in water solution. For this scope, the SBP were characterized for chemical composition and molecular weight (MW) by microanalysis, potentiometric titration, (13)C NMR spectroscopy, and size exclusion chromatography (SEC) coupled with an online multiangle light scattering (MALS) detector. These materials were found to have 67-463 kg mol(-1) MW and 6-53 polydispersity index and to contain carboxylic acid and phenol groups bonded to aromatic and aliphatic C chains. An empirical parameter (LH) was calculated for use as an index of the lipophilic/hydrophilic C atoms ratio. The products solubility properties in solvents of different polarity, surface activity, power to enhance the water solubility of hydrophobic compounds, and particle size in water solution were also investigated by measurements of the products partition coefficient between polyethylene glycol and water (KPEGW) and of air-water surface tension (γ), water-hexane interfacial tension (IFT), disperse red orange dye solubility (DS), and dynamic light scattering (DLS) versus added SBP concentration (Cs). The results indicate that LH correlates well with KPEGW and with the products surface activity properties. Both γ and DS are shown to depend on Cs, although in opposite ways, that is, higher Cs values yield lower γ and higher DS values. Both DS-Cs and γ-Cs plots showed a significant slope change at approximately the same 1.8-2.5 g L(-1) Cs value. This suggested a change of molecular conformation taking place at the above Cs values. Hydrodynamic diameter values for SBP in solution at Cs ≤ 10 g L(-1) were found to range from 130 to 300 nm, consistent with their macromolecular nature. The DLS coupled to the γ data were consistent with molecules at the water-air interphase and in the bulk water phase having different conformations, but not significantly different molecular sizes. Molecular aggregates more likely form at 50-100 g L(-1) Cs. The results confirm that urban biowastes are a sustainable source of biobased products that may have real commercial perspectives.

Role of Organo-Modifier and Metal Impurities of Commercial Nanoclays in the Photo- and Thermo-Oxidation of Polyamide 11 Nanocomposites.

The photo-oxidative degradation processes of bio-based PA11 nanocomposites containing montmorillonite (MMT) and the organo-modified Cloisite®30B were investigated to discriminate the influence of organo-modified components on the polymer durability. Indeed, despite the extensive studies reported, there are still ambiguous points to be clarified from the chemical point of view. To this aim, UV-aged materials were analyzed by Size Exclusion Chromatography (SEC), Inductively Coupled Plasma-Mass Spectrometry (ICP-MS) and Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS). This enabled determining changes in both chemical structure and Molar Masses (MMs) induced by light, heat and oxygen exposure. The addition of organo-modified nanoclays strongly affected the PA11 light durability, triggering the macromolecular chains scission due to the typical αH, Norrish I and II mechanisms. However, the main contribution in boosting the photo-oxidative degradation is induced by iron impurities contained into the clays. Conversely, thermo-oxidation process performed at 215 °C was unambiguously affected by the presence of the organo-modifiers, whose presence determined an enhancement of crosslinking reactions.

TEMPO-Nanocellulose/Ca2+ Hydrogels: Ibuprofen Drug Diffusion and In Vitro Cytocompatibility.

Stable hydrogels with tunable rheological properties were prepared by adding Ca2+ ions to aqueous dispersions of 2,2,6,6-tetramethylpiperidine 1-oxyl (TEMPO)-oxidized and ultra-sonicated cellulose nanofibers (TOUS-CNFs). The gelation occurred by interaction among polyvalent cations and the carboxylic units introduced on TOUS-CNFs during the oxidation process. Both dynamic viscosity values and pseudoplastic rheological behaviour increased by increasing the Ca2+ concentration, confirming the cross-linking action of the bivalent cation. The hydrogels were proved to be suitable controlled release systems by measuring the diffusion coefficient of a drug model (ibuprofen, IB) by high-resolution magic angle spinning (HR-MAS) nuclear magnetic resonance (NMR) spectroscopy. IB was used both as free molecule and as a 1:1 pre-formed complex with ß-cyclodextrin (IB/ß-CD), showing in this latter case a lower diffusion coefficient. Finally, the cytocompatibility of the TOUS-CNFs/Ca2+ hydrogels was demonstrated in vitro by indirect and direct tests conducted on a L929 murine fibroblast cell line, achieving a percentage number of viable cells after 7 days higher than 70%.

Comparative Study of Chemical Composition, Molecular and Rheological Properties of Silicone Oil Medical Devices.

PURPOSE: We evaluated chemical composition, and molecular and rheological properties in 10 commercially available silicone oils (SilOils), focusing on siloxane chains of low molecular weight (LMW components, LMWC) that are known to be "impurities" produced during the SilOil synthesis process. METHODS: We assessed the type of SilOil polymer and molecular weight distribution (MWD) by spectroscopy and conventional size exclusion chromatography, respectively. From the Cumulative MWD, we calculated the fractions of LMWC with molecular weight (M): ≤2000, ≤5000, and ≤10,000 g/mol. Due to the low MW, the content of LMWC with M ≤1000 g/mol was determined by gas chromatography-mass spectrometry. The dynamic viscosity (η) was assessed by rotational rheometry. RESULTS: For all SilOils, the polymer was polydimethylsiloxane. The samples differed significantly in terms of MWD and relative LMWC fractions. Specifically, the relative fraction of all LMWC (M ≤10,000 g/mol) ranged from 2.31% to 9.40% and the content of LMWC with M ≤1000 g/mol also varied significantly (range, 51-1151 ppm). The η values were different between the SilOils, and, for many of them, from the declared viscosity. CONCLUSIONS: Commercially available SilOils differ significantly in molecular and rheologic features. These compounds contain a significant amount of LMWC, "impurities" generated during the synthesis process, acting as emulsifier, potentially inducing ocular inflammation and toxicity. TRANSLATIONAL RELEVANCE: The amount of impurities in different SilOils may influence significantly their biocompatibility.

Physicochemical Properties of A New PEGylated Polybenzofulvene Brush for Drug Encapsulation.

A new polymer brush was synthesized by spontaneous polymerization of benzofulvene macromonomer 6-MOEG-9-T-BF3k bearing a nona(ethylene glycol) side chain linked to the 3-phenylindene scaffold by means of a triazole heterocycle. The polymer structure was studied by SEC-MALS, NMR spectroscopy, and MALDI-TOF MS techniques, and the results supported the role of oligomeric initiatory species in the spontaneous polymerization of polybenzofulvene derivatives. The aggregation features of high molecular weight poly-6-MOEG-9-T-BF3k-FE were investigated by pyrene fluorescence analysis, dynamic light scattering studies, and transmission electron microscopy, which suggested a tendency towards the formation of spherical objects showing dimensions in the range of 20-200 nm. Moreover, poly-6-MOEG-9-T-BF3k-FE showed an interesting cytocompatibility in the whole concentration range tested that, besides its aggregation features, makes this polybenzofulvene brush a good polymer candidate for nanoencapsulation and delivery of drug molecules. Finally, the photo-physical features of poly-6-MOEG-9-T-BF3k-FE could allow the biodistribution of the resulting drug delivery systems to be monitored by fluorescence microscopy techniques.

Self-Structuring in Water of Polyamidoamino Acids with Hydrophobic Side Chains Deriving from Natural α-Amino Acids.

This paper reports on synthesis, acid-base properties and self-structuring in water of chiral polyamidoamino acids (PAACs) obtained by polyaddition of N,N'-methylenebisacrylamide with l-alanine, l-valine and l-leucine (M-l-Ala, M-l-Val, M-l-Leu) with potential for selective interactions with biomolecules. The polymers maintained the acid-base properties of amino acids. In water, the circular dichroism spectra of PAACs revealed pH-dependent structuring in the range 3⁻11 and in the wavelength interval 200⁻280 nm. Taking as reference the values at pH 3, the differential molar ellipticities were plotted in the pH interval 3⁻11. Sigmoidal curves were obtained presenting inflection points at pH 8.1, 6.8 and 7.3 for M-l-Ala, M-l-Val and M-l-Leu, respectively, corresponding to the amine half-ionization. Theoretical modeling showed that PAACs assumed stable folded conformations. Intramolecular interactions led to transoid arrangements of the main chain reminiscent of protein hairpin motif. Oligomers with ten repeat units had simulated gyration radii consistent with the hydrodynamic radii obtained by dynamic light scattering.

Structural Manipulation of the Conjugated Phenyl Moiety in 3-Phenylbenzofulvene Monomers: Effects on Spontaneous Polymerization.

Spontaneous polymerization is an intriguing phenomenon in which pure monomers begin their polymerization without initiators or catalysts. Previously, 3-phenylbenzofulvene monomers were found to polymerize spontaneously after solvent removal. Here, eight new 3-substituted benzofulvene monomers 1a⁻h were synthesized in order to investigate the effects of differently substituted aromatic rings in position 3 of the benzofulvene scaffold on spontaneous polymerization. The newly synthesized monomers maintained the tendency toward spontaneous polymerization. However, monomer 1a, bearing an ortho-methoxy substituted phenyl, polymerized hardly, thus producing low polymerization yields, inhomogeneous structure, and low molecular weight of the obtained polymeric material. This result suggested the importance of the presence of hydrogen atoms in the 2'-position to achieve productive interactions among the monomers in the recognition step preluding the spontaneous polymerization and among the monomeric units in the polybenzofulvene backbones. Moreover, this study paves the way to modify the pendant rings in position 3 of the indene scaffold to synthesize new polybenzofulvene derivatives variously decorated.

Densely PEGylated Polybenzofulvene Brushes for Potential Applications in Drug Encapsulation.

The technique of grafting side chains onto a linear polymeric backbone is commonly used to confer to the new polymeric material with desired properties, such as tunable solubility, ionic charge, biocompatibility, or specific interactions with biological systems. In this paper, two new polybenzofulvene backbones were assembled by spontaneous polymerization of the appropriate benzofulvene monomers (4,6-PO-BF3k and 4',6-PO-BF3k) bearing two clickable propargyloxy groups in different positions of the 3-phenylindene scaffold. Poly-4,6-PO-BF3k and poly-4',6-PO-BF3k were grafted with monomethyl oligo(ethylene glycol) (MOEG) to prepare two new polybenzofulvene brushes (i.e., poly-4,6-MOEG-9-TM-BF3k and poly-4',6-MOEG-9-TM-BF3k) by means of a "grafting onto" approach, that were characterized from the point of view of their macromolecular features, aggregation liability, and in a preliminary evaluation of biocompatibility. The obtained results make these PEGylated polybenzofulvene brushes (PPBFB) derivatives potentially useful as nanocarriers for nanoencapsulation and delivery of drug molecules.

Hyaluronan-based graft copolymers bearing aggregation-induced emission fluorogens.

In order to develop a technology platform based on two natural compounds from biorenewable resources, a short series of hyaluronan (HA) copolymers grafted with propargylated ferulic acid (HA-FA-Pg) were designed and synthesized to show different grafting degree values and their optical properties were characterized in comparison with reference compounds containing the same ferulate fluorophore. Interestingly, these studies revealed that the ferulate fluorophore was quite sensitive to the restriction of intramolecular motion and its introduction into the rigid HA backbone, as in HA-FA-Pg graft copolymers, led to higher photoluminescence quantum yield values than those obtained with the isolated fluorophore. Thus, the propargyl groups of HA-FA-Pg derivatives were exploited in the coupling with oleic acid through a biocompatible nona(ethylene glycol) spacer as an example of the possible applications of this technology platform. The resulting HA-FA-NEG-OA materials showed self-assembling capabilities in aqueous environment. Furthermore, HA-FA-NEG-OA derivatives have been shown to interact with phospholipid bilayers both in liposomes and living cells, retaining their fluorogenic properties and showing a high degree of cytocompatibility and for this reason they were proposed as potential biocompatible self-assembled aggregates forming new materials for biomedical applications.

Solution properties of high-molar-mass hyaluronans: the biopolymer degradation by ascorbate.

An accurate molecular characterization, molar mass and size distributions, of 10 hyaluronan (HA) samples was performed by using a multi-angle light scattering detector connected on-line to a size exclusion chromatographic system. The dynamic viscosity eta of the HA solutions was investigated using a rotational viscometer. On monitoring the sample dynamic viscosity for up to 5h, a small however constant increase of the eta value was observed, indicating rheopectic behavior of all 10 HA solutions. Addition of ascorbic acid to the HA solutions caused significant changes in the rheological properties of the samples investigated. The change of eta values in the course of time was explained by the redox reactions (caused by the added ascorbate) that occur during the dynamic viscosity monitoring.

Composite films prepared from agricultural by-products.

In our study we used holocelluloses from sugar beet and bagasse for film preparation. Films from sugar beet holocellulose have better mechanical properties than from bagasse holocellulose. By subsequent carboxymethylation of bagasse holocellulose, films with better properties were produced. Specimens prepared from combined sugar beet and bagasse carboxymethylated holocellulose had the best mechanical properties. The results could be explained by the ratios of cellulose, arabinan, polygalacturonan and xylan content in individual films, based on the elemental analysis data. The use of microwaves to prepare holocellulose film speed up the process, but negatively affected the mechanical properties. Lignin content of the sugar beet holocellulose and bagasse samples was low and did not affect the mechanical properties. Both types of agricultural by-products could be used for preparation of composite film with high strength and stiffness suitable for broad range of applications.

Grafting of Hindered Phenol Groups onto Ethylene/α-Olefin Copolymer by Nitroxide Radical Coupling.

The covalent immobilization of hindered phenol groups, with potential antioxidant activity, onto an ethylene/α-olefin (EOC) copolymer was carried out by the nitroxide radical coupling (NRC) reaction performed in the melt with a peroxide and the 3,5-di-tert-butyl-4-hydroxybenzoyl-2,2,6,6-tetramethylpiperidine-1-oxyl radical (BHB-T). Functionalized EOC (EOC-g-(BHB-T)) was exposed to photo- and thermo-oxidation. By comparison with some model compounds bearing the (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO) moiety or the hindered phenol unit, it was observed that the grafted BHB-T could effectively help the stabilization of the polymer matrix both under photo- and thermo-oxidation. In addition, the immobilization of BHB-T can effectively increase the service life of the functionalized polymers when polymer films were put in contact with ethanol solution thus simulating a possible application of the modified polymer.

Hydrogen peroxide generation by the Weissberger biogenic oxidative system during hyaluronan degradation.

By applying the enzyme catalase, our study on hyaluronan degradation confirms the generation of hydrogen peroxide using the Weissberger biogenic oxidative system (WBOS), which is composed of ascorbate and cupric ions. Dynamic viscosities of hyaluronan (HA) solutions influenced by WBOS in the absence and presence of catalase were analysed by rotational viscometry. Molar masses of HAs were determined by size-exclusion chromatography with multi-angle laser-light scattering. Our results show that catalase dose-dependently inhibited the degradation of HA macromolecules, which presumably confirms the generation of H2O2 in the reaction system. This has implications in range of biomedical applications such as arthritic joint treatment, tissue engineering, ocular and cosmetic surgery.

Cationization of heparin for film applications.

Trimethylammonium-2-hydroxypropyl-(TMAHP) spacer was introduced into heparin (H) and the prepared films were characterized by elemental analysis, NMR, SEC-MALS, TG/DTG/DTA, AFM and mechanical tester. When quaternized at the ratio of H/NaOH/alkylating agent/H2O=0.1-1/0-2/0.1-1/50-500 mmol, H was substituted at A6 and A3 positions. The formation of double-substituted structures by substitution of free hydroxyl group of the previously introduced TMAHP substituent is evident. In the absence of NaOH (H/GTMAC/H2O=1:1:500) the most drastic decrease of M(n) to 8.639 kg/mol and M(w)/M(n) at 1.48 was observed in comparison to H (Mn=9532 g/mol with M(w)/M(n)=1.38). The film mechanical properties were better on H (E=4030 MPa; σ(b)=65 MPa; ɛ(b)=4.6%) than on quaternized specimens (E=2500-3340 MPa; σ(b)=25-40 MPa; ɛ(b)=1.7-1.8%). The AFM images did not prove relation between mechanical properties and surface shape.

Enzymatic and chemical oxidation of polygalactomannans from the seeds of a few species of leguminous plants and characterization of the oxidized products.

Plant polysaccharides are used in a growing number of applications, in their native or in chemically and/or biochemically modified forms. In the present work, we compare TEMPO-mediated oxidation with laccase of polygalactomannans (PGM) from different species of plant leguminous to chemical oxidation with NaClO/NaBr/TEMPO. We have investigated the gums from: locust bean (Ceratonia siliqua), tara (Caesalpinia spinosa), guar (Cyamopsis tetragonolobus), sesbania (Sesbania bispinosa) and fenugreek (Trigonella foenum-graecum). Upon laccase/TEMPO oxidation, PGM viscosity and concentration of reducing groups increased up to five-fold and structured, elastic, stable gels were formed, which could be degraded by hydrolysis with ß-mannanase. Conversely, chemical oxidation with NaClO/NaBr/TEMPO caused a rapid, intermediate transition of the gum solutions to compact gels, that immediately reverted to liquid, with a lower viscosity than at the start and an increased concentration of reducing groups, similar to the reaction with laccase. We interpret the above as due to, in the case of laccase, oxidation of primary hydroxyl groups to aldehydes, able to form stable hemiacetalic bonds with free hydroxyl groups. While upon chemical oxidation, primary OH's are only transiently oxidized to aldehydes, followed by rapid oxidation of all carbonyl groups to carboxylates. In either cases, TEMPO appeared to cause a limited splitting of glycosidic bonds of PGM. Native and oxidized PGM were further characterized by 1D and 2D NMR spectroscopy and by rheology.

Polybenzofulvene derivatives bearing dynamic binding sites as potential anticancer drug delivery systems.

In order to obtain new advanced functional materials capable of recognizing drug molecules, the polybenzofulvene backbone of molecular brush poly-6-MOEG-9-TM-BF3k has been functionalized with a "synthetic dynamic receptor" composed of two 1-adamantylurea moieties linked together by means of a dipropyleneamino bridge as in Meijer's bis(adamantylurea) pincer (BAUP). This functional material, bearing synthetic receptors potentially capable of recognizing/loading and then delivering drug molecules, was used to prepare colloidal drug delivery systems (by means of soft interaction with BAUP) for delivering the model anti-cancer drug doxorubicin (DOXO). The resulting nanostructured drug delivery systems containing the physically loaded drug were characterized in terms of drug loading and release, dimensions and zeta potential, and in vitro cell activity and uptake on two different cell lines (i.e. the human bronchial epithelial 16HBE and the human colon cancer HCT116). On normal cells, free DOXO was found to be more cytotoxic than DOXO-loaded nanogels at the higher tested concentration and, only on cancer cells, DOXO-loaded nanogels show similar or slightly higher cytotoxicity values than free DOXO, suggesting potential advantages in the treatment of cancer. These results were supported by fluorescence microscopy studies, which suggested that DOXO-loaded nanogels provide an extracellular reservoir of the drug, which is gradually released and internalized within the cells.

Wound healing properties of hyaluronan derivatives bearing ferulate residues.

HAFA macromolecules were designed as graft copolymers combining ferulic acid (FA) structure and the hyaluronic acid (HA) backbone linked through an ester bond. These materials were prepared by feruloylation of HA with bisimidazolide 3 [i.e. (E)-4-(3-(1H-imidazol-1-yl)-3-oxoprop-1-enyl)-2-methoxyphenyl 1H-imidazole-1-carboxylate] and obtained with different grafting degree (GD) values, which could be tuned by applying suitable reaction conditions. Among the numerous applications envisioned for HAFA graft copolymers on the basis of the physico-chemical, biological, and pharmacological properties of the starting natural products and the grafting-derived features such as physical cross-linking, potential wound healing properties have been evaluated in vitro and in vivo in preclinical models. In human keratinocyte (HaCaT) cells, our data showed the ability of HAFA-17 (GD = 7%) to ameliorate the in vitro scratch wound significantly with respect to the control HA and FA alone, and this effect was associated with the ability of HAFA-17 to also induce keratinocyte proliferation as determined by BrdU assay. In addition, experiments on wound healing in SKH1 mice confirmed the ability of HAFA-17 to improve the wound closure rate also in vivo. Overall, the data presented herein suggest HAFA-17 as a possible future drug for the therapeutic treatment of acute and chronic wounds.