Surface-grafted, environmentally sensitive polymers for biofilm release.
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Controlling bacterial biofouling is desirable for almost every human enterprise in which solid surfaces are introduced into nonsterile aqueous environments. One approach that is used to decrease contamination of manufactured devices by microorganisms is using materials that easily slough off accumulated material (i.e., fouling release surfaces). The compounds currently used for this purpose rely on low surface energy to inhibit strong attachment of organisms. In this study, we examined the possible use of environmentally responsive (or "smart") polymers as a new class of fouling release agents; a surface-grafted thermally responsive polymer, poly(N-isopropylacrylamide) (PNIPAAM), was used as a model compound. PNIPAAM is known to have a lower critical solubility temperature of approximately 32 degrees C (i.e., it is insoluble in water at temperatures above 32 degrees C and is soluble at temperatures below 32 degrees C). Under experimental conditions, >90% of cultured microorganisms (Staphylococcus epidermidis, Halomonas marina) and naturally occurring marine microorganisms that attached to grafted PNIPAAM surfaces during 2-, 18-, 36-, and 72-h incubations were removed when the hydration state of the polymer was changed from a wettability that was favorable for attachment to a wettability that was less favorable. Of particular significance is the observation that an organism known to attach in the greatest numbers to hydrophobic substrata (i.e., H. marina) was removed when transition of PNIPAAM to a more hydrated state occurred, whereas an organism that attaches in the greatest numbers to hydrophilic substrata (i.e., S. epidermidis) was removed when the opposite transition occurred. Neither solvated nor desolvated PNIPAAM exhibited intrinsic fouling release properties, indicating that the phase transition was the important factor in removal of organisms. Based on our observations of the behavior of this model system, we suggest that environmentally responsive polymers represent a new approach for controlling biofouling release. (+info)
Effects of palytoxin on isolated intestinal and vascular smooth muscles.
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Palytoxin (PTX), the most potent marine toxin isolated from the Zoanthid, Palythoa tuberculosa, was studied to determine the effect on isolated smooth muscles. In guinea pig taenia coli PTX at above 3 X 10(-10) g/ml caused a contraction which slowly subsided under isotonic recording. Under isometric recording PTX at above 1 X 10(-10) g/ml caused a contraction which depended on the spontaneous activity. The PTX-induced contraction was not affected by atropine, tripelenmamine or tetrodotoxin but was inhibited by 5 mM Mg, norephinrphrine, isoprenaline or papaverine. PTX at above 1 X 10(-9) g/ml induced an increase in spike frequency and a slight depolarization accompanied with a contraction when measured using a sucrose gap method. In some cases the spike generation was almost abolished after a long exposure to higher dose of PTX and the developed tension gradually decreased. Under isometric recording PTX caused a sustained contraction in rabbit aorta, dog mesenteric and coronary arteries at above 1 X 10(-10) and 1 X 10(-11) g/ml, respectively, in a dose-dependent manner. The coronary artery was most sensitive among the preparation used. PTX-induced contraction in aorta was irreversible, was not influenced by phentolamine but diminished with 5 mM Mg and disappeared in a D-600 or Ca-free medium. PTX is thus an extremely potent and direct stimulant which acts on smooth muscles. (+info)
New biodegradable hydrogels based on a photocrosslinkable modified polyaspartamide: synthesis and characterization.
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alpha,beta-Poly(N-2-hydroxyethyl)-DL-aspartamide (PHEA), a synthetic water-soluble biocompatible polymer, was derivatized with glycidyl methacrylate (GMA), in order to introduce in its structure chemical residues having double bonds and ester groups. The obtained copolymer (PHG) contained 29 mol% of GMA residues. PHG aqueous solutions at various concentrations ranging from 30 to 70 mg/ml were exposed to a source of UV rays at lambda 254 nm in the presence or in the absence of N,N'-methylenebisacrylamide (BIS); the formation of compact gel phases was observed beginning from 50 mg/ml. The obtained networks were characterized by FT-IR spectrophotometry and swelling measurements which evidenced the high affinity of PHG hydrogels towards aqueous media at different pH values. In vitro chemical or enzymatic hydrolysis studies suggested that the prepared samples undergo a partial degradation both at pH 1 and pH 10 and after incubation with enzymes such as esterase, pepsin and alpha-chymotrypsin. Finally, the effect of irradiation time on the yield and the properties of these hydrogels was investigated and the sol fractions coming from irradiated samples, properly purified, were characterized by FT-IR and 1H-NMR analyses. (+info)
Modification of liposomes with N-substituted polyacrylamides: identification of proteins adsorbed from plasma.
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Liposomes prepared from DMPC (80%) and cholesterol (20%) were modified with a series of hydrophobically modified N-substituted polyacrylamides, namely, poly[N-isopropylacrylamide] (PNIPAM), poly[N,N-bis(2-methoxyethyl) acrylamide] (PMEAM), and poly[(3-methoxypropyl)acrylamide] (PMPAM). The hydrophobic group, N-[4-(1-pyrenylbutyl)-N-n-octadecylamine was attached to one end of the polymer chains to serve as an anchor for incorporation into the liposome bilayer. Liposome-polymer interactions were confirmed using fluorescence spectroscopy and chemical analysis. Microscopy revealed differences in aggregation tendency between unmodified and polymer-modified liposomes. Proteins adsorbed to liposome surfaces during exposure to human plasma were identified by immunoblot analysis. It was found that both unmodified and polymer-modified liposomes adsorb a wide variety of plasma proteins. Contact phase coagulation proteins, complement proteins, cell-adhesive proteins, serine protease inhibitors, plasminogen, antithrombin III, prothrombin, transferrin, alpha(2)-microglobulin, hemoglobin, haptoglobin and beta-lipoprotein as well as the major plasma proteins were all detected. Some differences were found between the unmodified and polymer-modified liposomes. The unmodified liposomes adsorbed plasminogen mainly as the intact protein, whereas on the modified liposomes plasminogen was present in degraded form. Also, the liposomes modified with PNIPAM in its extended conformation (below the lower critical solution temperature) appeared to adsorb less protein than those containing the 'collapsed' form of PNIPAM (above the LCST). (+info)
Characterization of a palytoxin-induced non-selective cation channel in mouse megakaryocytes.
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We used the whole-cell clamp and fura-2 techniques to study the membrane current and intracellular Ca2+ concentration ([Ca2+]i) changes of mouse megakaryocytes in response to palytoxin (PTX), a highly potent marine toxin. At a holding potential of -60 mV, PTX induced a sustained inward current in a dose-dependent manner. The reversal potentials measured in the presence of various extracellular major cations indicated that the PTX-induced channel had a non-selective permeability to alkali metal ions. Although elimination of intracellular Ca2+ had no effect on the PTX-induced current, removal of external Ca2+ inhibited the current activation. During the sustained phase of the PTX-induced current, treatment with ADP activated an additional current. Pretreatment with ouabain, an inhibitor of Na+-K+-ATPase, suppressed the PTX-induced current. During the stable phase of the PTX-induced current, challenge with NiCl2 (5 mM) or 2,4-dichlorobenzamil (DCB, 25 microM), a non-selective cation channel blocker, partially reversed the current. Simultaneous measurement of the membrane current and [Ca2+]i showed that PTX induced the current response without increasing the [Ca2+]i. Taken together, these results indicate that PTX induces a non-selective cation channel in mouse megakaryocytes. This channel is distinct from the ADP-operated channel and is sensitive to ouabain, NiCl2 and DCB. (+info)
Determination of acrylamide monomer in polyacrylamide degradation studies by high-performance liquid chromatography.
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A high-performance liquid chromatography method using C18 and ion-exchange columns in series is developed for the determination of acrylamide and acrylic acid monomers in polymeric samples. The C18 column acts as a guard column, trapping surfactants and impurities and retaining the nonionic species. The ion-exchange column then separates the monomers according to their respective ionic strengths. This method has been proven in the laboratory to work successfully for all types of acrylamide/acrylic acid polymers and matrices. Detection limits for both monomers can be achieved in the parts-per-billion range. The method is used to study the possible degradation of polyacrylamide to acrylamide monomer in the presence of glyphosate (a herbicide) and sunlight. Polyacrylamide is used as a spray drift reduction aid in combination with glyphosate. In normal applications, the polymer and herbicide are in contact with each other in the presence of sunlight. The results show that the polymer does not degrade to acrylamide in the presence of glyphosate or sunlight or any combination of the two. It is also observed that glyphosate influences the solubility of polyacrylamide, and care must be used when combining the two. (+info)
In vitro characterization of a novel polymeric-based pH-sensitive liposome system.
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This study demonstrates rapid and pH-sensitive release of a highly water-soluble fluorescent aqueous content marker, pyranine, from egg phosphatidylcholine liposomes following incorporation of N-isopropylacrylamide (NIPA) copolymers in liposomal membranes. The pH-sensitivity of this system correlates with the precipitation of the copolymers at acidic pH. In vitro release can be significantly improved by increasing the percentage of anchor in the copolymer and thus favoring its binding to the liposomal bilayer. In the case of liposomes containing a poly(ethylene glycol)-phospholipid conjugate, the insertion of the pH-sensitive copolymer in the liposomal membrane appears to be sterically inhibited. Dye release from these formulations at acidic pH can still be achieved by varying the anchor molar ratio and/or molecular mass of the polymers or by including the latter during the liposome preparation procedure. Removal of unbound polymer results in decreased leakage only when the copolymer is inserted by incubation with preformed liposomes, but can be overcome by preparing liposomes in the presence of polymer. Aqueous content and lipid mixing assays suggest contents release can occur without membrane fusion. The results of this study indicate that the addition of pH-sensitive copolymers of NIPA represents promising strategy for improving liposomal drug delivery. (+info)
Antitumor activity of N-(2-hydroxypropyl) methacrylamide copolymer-Mesochlorine e6 and adriamycin conjugates in combination treatments.
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This study demonstrates the selective tumor targeting and the antitumor efficacy of the N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer-bound mesochlorin e6 monoethylenediamine (Mce6) and HPMA copolymer-bound Adriamycin (ADR) in combination photodynamic therapy (PDT) and chemotherapy against human ovarian OVCAR-3 carcinoma xenografted in female athynmic mice. The concentrations of Mce6 and ADR in blood and tissues, in free or HPMA copolymer-bound form, were determined by fluorescence and high-performance liquid chromatography fluorescence assays, respectively. Xenograft responses to single and combination therapies were recorded. The peak concentration of HPMA copolymer-Mce6 conjugate in tumor was achieved 18 h after administration. For HPMA copolymer-bound drugs, the concentration ratios of liver and spleen versus muscle were significantly higher than those of free drugs. The HPMA copolymer-bound drugs demonstrated selective targeting and accumulation in the tumor, probably attributed to the enhanced permeability and retention effect. In vivo studies revealed that all tumors in the treatment groups showed significant responses after receiving any of the various types of therapy as compared with controls (P < 0.001). PDT with HPMA copolymer-Mce6 conjugate (PDTMC) at a dose of 13.4 mg/kg (1.5 mg/kg of Mce6 equivalent) and light doses of 110 J/cm2 at 12 and 18 h, respectively, resulted in significant suppression of the growth of OVCAR-3 tumors. Three courses of chemotherapy using 35 mg/kg (2.2 mg/kg of ADR equivalent) of HPMA copolymer-ADR conjugate (CHEMO) were effective in suppressing the growth of tumors. Single PDTMC plus multiple CHEMO exhibited significantly greater therapeutic efficacy than multiple CHEMO. In the group of mice receiving multiple PDTMC, tumor recurrence became obvious after day 20. However, 10 of 12 tumors exhibited complete responses in the group of mice receiving multiple PDTMC plus multiple CHEMO. The least to most effective treatments were ranked as follows: multiple CHEMO < single PDTMC plus multiple CHEMO < multiple PDTMC < multiple PDTMC plus multiple CHEMO. The results clearly demonstrate that: (a) HPMA copolymer-bound drugs exhibited selective tumor accumulation contrary to free drugs; (b) PDT using HPMA copolymer-Mce6 conjugate with multiple light irradiations was a better therapy than that with single light irradiation; and (c) combination chemotherapy and photodynamic therapy with HPMA copolymer-ADR and HPMA copolymer-Mce6 conjugates was the most effective regimen. (+info)