Calorimetric and chiroptical evidence of aggregate-driven helix formation in carrageenan systems

Thermally induced, order-disorder transitions of iota- and kappa-carrageenan have been monitored by optical rotation and differential-scanning calorimetry in various ionic environments. Conformational ordering in kappa-carrageenan is observed only in the presence of cations that have been shown previously to promote helix-helix aggregation, and shows marked hysteresis between heating and cooling. Iota-carrageenan, by contrast, shows an order-disorder transition in the non-aggregating, tetramethylammonium salt form, at substantially lower temperature than for kappa-carrageenan, and without hysteresis. In the presence of potassium ions, which are known to promote aggregation, iota-carrageenan shows two distinct thermal-transitions, one without hysteresis at the same temperature as observed under non-aggregating conditions, and one with significant hysteresis close to the temperature of the kappa-carrageenan transition. We interpret these transitions as helix-to-coil and aggregated helix-to-coil, respectively. This interpretation is supported by measurements of the enthalpy changes of the transitions; ΔH values show a systematic increase with increasing aggregation and hysteresis. We conclude that the double helix of iota-carrageenan can exist as a stable entity in isolation, but may be further stabilised by aggregation, whereas the kappa-carrageenan helix is stable only when aggregated.     

Cross-species induction of antibacterial activity produced by epibiotic bacteria isolated from Indian marine sponge Pseudoceratina purpurea

Four antibiotic producing bacteria were isolated from the surface of the marine sponge Pseudoceratina purpurea and exposed to living cells of two human pathogenic bacteria as well as some marine fouling bacteria to induce the production of antimicrobial activity. Experimental results showed that these four marine epibiotic bacteria enhanced their antibacterial production, when exposed to these test strains. The highest induction was exhibited by the sponge isolate PS79 against fouling bacterium FB-9 (from 3 mm to 7 mm inhibition zone). All the four strains were induced and showed increased activity specifically against the challenged pathogenic or fouling bacteria tested. Specific induction by these species suggests that the induction might be attributed to the response to the chemical signals received from potential challenger strains.     

Comparison between a moving bed membrane bioreactor and a conventional membrane bioreactor on membrane fouling

A membrane bioreactor filled with carriers instead of activated sludge named a moving bed membrane bioreactor (MBMBR) was investigated to minimize the effect of suspended solids on membrane fouling. The MBMBR and a conventional membrane bioreactor (CMBR) were operated in parallel for about two months. Unexpectedly, the rate of membrane fouling in MBMBR was about three times of that in CMBR. MBMBR showed a higher cake layer resistance than CMBR due to plenty of filamentous bacteria inhabited in suspended solids in MBMBR. Protein and polysaccharide contents of soluble EPS in MBMBR were obviously larger than those in CMBR. It could be speculated that the overgrowth of filamentous bacteria in MBMBR resulted in severe cake layer and induced a large quantity of EPS, which deteriorated the membrane fouling.     

Synergistic Interaction between Kappa-Carrageenan and Locust-bean Gum in Aqueous Media

Although neither kappa-carrageenan nor locust-bean gum gelled alone, a mixed aqueous solution of the above gums gave a gel at the concentration of 0.6% total gums in a range of low temperatures. The solution also gelled even at the concentration of 0.4% total gums in the presence of 0.1% KC1. The maximum dynamic modulus was obtained with a series of the samples composed of kappa-carrageenan and locust-bean gum in the mixing ratios of 1:1 and 3:1 at the concentration of 0.6 and 0.8% total gums at 0°C. The dynamic modulus of a mixed solution of kappa-carrageenan and locust-bean gum was not influenced by pH between pH 7.0 to 11.5, but decreased in the acidic range.

We concluded that intermolecular interactions, at low temperature, between kappa-carrageenan and locust-bean gum may take place on the K⁺-bridge of the former and the backbone of the latter molecule at low concentrations, but at high concentration of the gums, self-association of kappa-carrageenan molecules might also occurred.     

Rheological properties of mixtures of protein-polysaccharide-dynamic viscoelasticity of blend gels of acylated gelatin, kappa-carrageenan, and agarose

Complex Young's modulus of blend gels of gelatin and kappa-carrageenan or agarose has been measured in order to clarify the protein-polysaccharide interaction in biological systems. The mixture of gelatin and kappa-carrageenan showed phase separation in the intermediate volume fraction of gelatin, and it formed a homogeneous gel when the volume fraction of gelatin is very large or very small. Since the dynamic Young's modulus for blend gels of kappa-carrageenan and gelatin was larger than the calculated one from a theory for dispersed systems, some structural reinforcing must occur. The mixture of agarose and gelatin showed the inverse tendency. It was concluded that the role of electrolytic groups was dominant in dilute gels, while molecular entanglement became more important in concentrated gels.     

A comparative study of the non-acidic chemically mediated antifoulant properties of three sympatric species of ascidians associated with seagrass habitats

The present study investigated aspects of the antifoulant properties of three sympatric species of ascidians found in seagrass habitats of the Gulf of Mexico, Southern Atlantic Ocean, and Caribbean. Field observations in Saint Joseph Bay, Florida indicate that all three species are common and that the tunic of the solitary ascidian Molgula occidentalis is often heavily fouled, while the outer surfaces of both the colonial ascidians Amaroucium stellatum and Botryllus planus are free of fouling organisms. Antifoulant activities of a suite of increasing hydrophilic organic extracts prepared from the tunic of M. occidentalis and whole colonies of A. stellatum and B. planus were measured using both sympatric microbial (bacteria) and macroinvertebrate (cyprid larvae of Balanus amphitrite) fouling organisms in laboratory bioassays. In addition, field antifoulant assays were conducted by combining organic extracts with controlled-release resin and subsequently coating this material on to acrylic rods deployed in the field for a 72 h period. Extracts of the tunic of M. occidentalis generally did not inhibit bacterial growth. The exception was the methanol extract, which inhibited growth in one of the six marine bacteria tested. Moreover, only the highest concentrations of hexane and methanol tunic extracts tested prevented attachment of cyprid larvae. Field assays revealed no antifoulant activity on rods coated with resin containing extracts of M. occidentalis. Inhibition of both microbial growth and cyprid settlement were much more pronounced in whole-organism extracts of the two colonial ascidians. Most potent were the aqueous methanol extracts of colonies of B. planus and A. stellatum which inhibited growth in five of the six marine bacteria tested. In addition, hydrophilic and lipophilic extracts of the colonial ascidians significantly inhibited attachment of cyprid larvae, in many instances across a wide range of extract concentrations. Field antifoulant assays indicated that extracts of both colonial ascidians inhibited settlement of bryozoans and barnacles. The findings indicate that the colonial ascidians B. planus and A. stellatum possess chemical antifoulant properties. In contrast, the solitary ascidian M. occidentalis appears to either tolerate fouling or possess other non-chemical mechanisms to cope with the risks associated with epibiont overgrowth.     

Biomimetic characterisation of key surface parameters for the development of fouling resistant materials

Material science provides a direct route to developing a new generation of non-toxic, surface effect-based antifouling technologies with applications ranging from biomedical science to marine transport. The surface topography of materials directly affects fouling resistance and fouling removal, the two key mechanisms for antifouling technologies. However, the field is hindered by the lack of quantified surface characteristics to guide the development of new antifouling materials. Using a biomimetic approach, key surface parameters are defined and quantified and correlated with fouling resistance and fouling removal from the shells of marine molluscs. Laser scanning confocal microscopy was used to acquire images for quantitative surface characterisation using three-dimensional surface parameters, and field assays correlated these with fouling resistance and fouling release. Principle component analysis produced a major component (explaining 54% of total variation between shell surfaces) that correlated with fouling resistance. The five surface parameters positively correlated to increased fouling resistance were, in order of importance, low fractal dimension, high skewness of both the roughness and waviness profiles, higher values of isotropy and lower values of mean surface roughness. The second component (accounting for 20% of variation between shells) positively correlated to fouling release, for which higher values of mean waviness almost exclusively dictated this relationship. This study provides quantified surface parameters to guide the development of new materials with surface properties that confer fouling resistance and release.     

Biomimetic characterisation of key surface parameters for the development of fouling resistant materials

Material science provides a direct route to developing a new generation of non-toxic, surface effect-based antifouling technologies with applications ranging from biomedical science to marine transport. The surface topography of materials directly affects fouling resistance and fouling removal, the two key mechanisms for antifouling technologies. However, the field is hindered by the lack of quantified surface characteristics to guide the development of new antifouling materials. Using a biomimetic approach, key surface parameters are defined and quantified and correlated with fouling resistance and fouling removal from the shells of marine molluscs. Laser scanning confocal microscopy was used to acquire images for quantitative surface characterisation using three-dimensional surface parameters, and field assays correlated these with fouling resistance and fouling release. Principle component analysis produced a major component (explaining 54% of total variation between shell surfaces) that correlated with fouling resistance. The five surface parameters positively correlated to increased fouling resistance were, in order of importance, low fractal dimension, high skewness of both the roughness and waviness profiles, higher values of isotropy and lower values of mean surface roughness. The second component (accounting for 20% of variation between shells) positively correlated to fouling release, for which higher values of mean waviness almost exclusively dictated this relationship. This study provides quantified surface parameters to guide the development of new materials with surface properties that confer fouling resistance and release.     

Adhesion and viability of two enterococcal strains on covalently grafted chitosan and chitosan/kappa-carrageenan multilayers

Chitosans are natural aminopolysaccharides, whose low cytotoxicity suggests their potential use for nonadhesive, antibacterial coatings on biomaterials implant surfaces. Here, the antiadhesive behavior and ability to kill bacteria upon adhesion ("contact killing") of chitosan coatings were evaluated for two strains of Enterococcus faecalis, isolated from clogged biliary stents. Chitosan coatings covalently grafted or applied as chitosan/kappa-carrageenan multilayers were characterized by ellipsometry, scanning force microscopy (SFM), X-ray photoelectron spectroscopy (XPS), and electrokinetic measurements. Decreases in initial bacterial deposition rates and the number of bacteria adhering in a more advanced state of the adhesion process were observed on both types of modified surfaces, with more pronounced effects on highly hydrated multilayers. Adhesion of negatively charged enterococci was slightly enhanced on chitosan-terminated multilayers, but antibacterial effect was absent on kappa-carrageenan-terminated multilayers. Thus, the efficacy of multilayers remains an interesting interplay between the promoting effect of cationically charged groups on adhesion of negatively charged bacteria and, on the other hand, their antibacterial effects.     

Marine microbial natural products in antifouling coatings

Ecological problems associated with current antifouling technologies have increased interest in the natural strategies that marine organisms use to keep their surfaces clean and free from fouling. Bacteria isolated from living surfaces in the marine environment have been shown to produce chemicals that are potential antifoulants. Active compounds from the cells and culture supernatant of two bacterial strains, FS‐55 and NudMB50–11, isolated from surface of the seaweed, Fucus serratus, and the nudibranch, Archidoris pseudoargus, respectively, were extracted using solid phase extraction. The extracts were combined with acrylic base paint resin and assayed for antifouling activity by measuring their ability to inhibit the growth of fouling bacteria. These formulations were found to be active against fouling bacteria isolated from marine surfaces. The formulation of antifouling paints that incorporate marine microbial natural products is reported here for the first time. This is a significant advance towards the production of an environmentally friendly antifouling paint that utilises a sustainable supply of natural biodegradable compounds.     

Bacterial biofilms: from the natural environment to infectious diseases

Biofilms--matrix-enclosed microbial accretions that adhere to biological or non-biological surfaces--represent a significant and incompletely understood mode of growth for bacteria. Biofilm formation appears early in the fossil record (approximately 3.25 billion years ago) and is common throughout a diverse range of organisms in both the Archaea and Bacteria lineages, including the 'living fossils' in the most deeply dividing branches of the phylogenetic tree. It is evident that biofilm formation is an ancient and integral component of the prokaryotic life cycle, and is a key factor for survival in diverse environments. Recent advances show that biofilms are structurally complex, dynamic systems with attributes of both primordial multicellular organisms and multifaceted ecosystems. Biofilm formation represents a protected mode of growth that allows cells to survive in hostile environments and also disperse to colonize new niches. The implications of these survival and propagative mechanisms in the context of both the natural environment and infectious diseases are discussed in this review.     

The effect of depth on the accrual of marine biofilms on glass substrata deployed in the Clyde Sea, Scotland

Recent concerns about fouling problems caused by biofilms affecting optical oceanographic instruments have highlighted the need for a better understanding of their nature and extent in the marine environment. Glass slides were deployed in April and August for periods of up to 3 weeks at 5, 10, 20, 40, 80 and 160 m in the water column of Loch Fyne, Clyde Sea, W Scotland. Biofilms were enumerated using epifluorescence and bright field microscopy. During April the biofilm community varied significantly with depth, although this effect was attributable solely to changes in the diatom community. Diatom numbers peaked between 10 and 20 m. During August the biofilm community also showed a significant depth effect, although in this case there were significant effects for diatoms, rods, and filamentous bacteria. Cell numbers for diatoms, filamentous bacteria, and rod shaped bacteria peaked at 5 m. There was a significant linear relationship between the number of diatoms and bacteria on the slides deployed in August. No such relationship was found for the April data. The results indicate that optical performance may be significantly degraded after a few weeks and highlights the need for provision of suitable strategies to protect such surfaces from biofilm accumulation.     

Sediment challenge to promising ultra-low fouling hydrophilic surfaces in the marine environment

Hydrophilic coatings exhibit ultra-low fouling properties in numerous laboratory experiments. In stark contrast, the antifouling effect of such coatings in vitro failed when performing field tests in the marine environment. The fouling release performance of nonionic and zwitterionic hydrophilic polymers was substantially reduced compared to the controlled laboratory environment. Microscopy and spectroscopy revealed that a large proportion of the accumulated material in field tests contains inorganic compounds and diatomaceous soil. Diatoms adhered to the accumulated material on the coating, but not to the pristine polymer. Simulating field tests in the laboratory using sediment samples collected from the test sites showed that incorporated sand and diatomaceous earth impairs the fouling release characteristics of the coatings. When exposed to marine sediment from multiple locations, particulate matter accumulated on these coatings and served as attachment points for diatom adhesion and enhanced fouling. Future developments of hydrophilic coatings should consider accumulated sediment and its potential impact on the antifouling performance.     

Stereocontrolled Synthesis of an Octasaccharide Part of I-Active Glycolipids

The effects of glycerin and ethylene glycol on the elastic modulus and DSC thermograms of agarose and kappa-carrageenan gels were examined to clarify the relation between structure and properties. The elastic modulus of these gels as a function of the concentration of polyols increased up to a certain concentration and then decreased with increasing concentration of polyols. These polyols shifted the melting temperature of the gel to higher temperatures in kappa-carrageenan gels but to lower temperatures in agarose gels. The temperature dependence of elastic modulus was changed in opposite directions in agarose and kappa-carrageenan gels by the addition of polyols, and this is discussed on the basis of model consisting of junction zones which are connected by Langevin chains. It was suggested that the mean distance between junction zones became shorter in the presence of a small amount of polyols.     

Desulfation of sulfated galactans with chlorotrimethylsilane. Characterization of beta-carrageenan by 1H NMR spectroscopy

A desulfation method using chlorotrimethylsilane for treatment of pyridinium salts of sulfated galactans was developed. It proved to be appropriate for desulfation of polysaccharides of both agar and carrageenan families. In order to evaluate its efficiency in presence of the maximum content of 3,6-anhydrogalactose, it was applied to commercial kappa-carrageenan, leading to obtention of a product mainly composed by beta-carrageenan. Best experimental conditions for achieving desulfation of kappa-carrageenan--in terms of low sulfate content, high recovery and low degradation of the product--were found. In addition, the complete assignment of the 1H NMR spectrum of beta-carrageenan was achieved by means of 1D and 2D NMR techniques.     

Gel--sol transition in kappa-carrageenan systems: microviscosity of hydrophobic microdomains, dynamic rheology and molecular conformation.

The effect of gel-sol transition in kappa-carrageenan systems on the microviscosity of hydrophobic microdomains, as well as its relation to macroscopic rheology and molecular conformation, was studied in kappa-carrageenan systems. The microdomains were probed by 1,3-di(-1-pyrenyl)propane (P3P) for which the excimer intensity (Ie) provides relative measures of the microviscosity in the immediate probe surroundings. In particular the applicability of P3P to monitor the gel--sol transition was proved, the results showing a dramatic decrease in microviscosity in the vicinity of the transition point. The corresponding changes in rheological properties and carrageenan conformation were investigated by dynamic viscometry (DV) and optical rotation (OR), respectively. The temperature of onset of the transition as indicated by the microviscosity data (T0) was found to correlate well with the OR and DV-results. The application of microviscosity and OR-measurements allowed an estimation of the helical content at T0 to be determined. P3P-data indicate a microenvironment viscosity for the probe sites in the kappa-carrageenan system comparable to that found in SDS micelles.     

Relation between structural and release properties in a polysaccharide gel system

The potential utility of kappa-carrageenan gels for preparing drug release devices is here shown. Structural properties of kappa-carrageenan gels prepared with different salt composition and containing Ketoprofen sodium salt, as model drug, have been evaluated with static light scattering and rheological measurements. These properties have been correlated with release profiles in vitro at pH 5.5. Release properties from gelled matrices have been compared with those obtained by two commercial products containing the same drug. Results show that: i) in this system it is possible to easily control the gel texture by using different cationic concentration; ii) the kinetics of drug release by kappa-carrageenan gels are dependent on the structural properties of matrices; iii) in the typical interval time used in classical local applications, all gel samples release the loaded drug almost completely, at difference with the commercial products. All these findings can provide useful suggestions for the realization of classical topical release systems.     

Aufwuchs Protozoa and Bacteria on the Gills of the Rock Crab,Cancer irroratus Say: A Survey by Light and Electron Microscopy1

Histological examination of gills of the rock crab (Cancer irroratus Say, 1917) taken from specimens collected in polluted and nonpolluted areas indicated that gill discoloration was not directly related to the degree of microbial fouling. Light microscope studies demonstrated that bacteria, peritrich and suctorian ciliates, and some naviculoid diatoms were the principal members of the fouling community. Examination of the bacteria by electron microscopy showed that several distinct types were present, and that some of these were attached to the gill by holdfast structures while others were enmeshed in a filamentous slime layer. An unusual small marine flagellate and an unidentified amoeba were also examined ultrastructurally.     

Dynamic light scattering studies of irradiated kappa carrageenan

Investigation of the dynamic behavior of irradiated kappa carrageenan (in KCl) as a function of irradiation dose and temperature was done by dynamic light scattering (DLS). The intensity correlation function (ICF) shifted towards shorter relaxation times with increasing radiation dose as a result of radiolysis. The characteristic decay time distribution function, G(gamma), indicates the presence of fast and slow mode peaks respectively at around 0.1-10 ms and 100-1000 ms. A peak broadening of the fast mode peak in G(gamma) appeared with decreasing temperature, indicating that coil-to-helical conformational transition took place. The conformation transition temperature (CTT) decreased with increasing radiation dose. No transition was observed for kappa-carrageenan irradiated at 200 kGy. A new faster relaxation mode appeared at around 0.1-1 ms at temperatures below the CTT. This peak is found in kappa-carrageenan irradiated at doses exclusively between 75 and 175 kGy. The peak height of this mode is largest at 100 kGy which corresponds to the optimum biologic activity of kappa-carrageenan reported previously.     

Physicochemical characterization of carrageenan fromGigartina teedii (Rooth) Lamouroux (Gigartinales,Rhodophyta)

The phycocolloids of female gametophytes ofGigartina teedii (Roth) Lamouroux harvested in Roscoff (Brittany, France) are a hybrid carrageenan resulting from juxtaposition of fragments of kappa-, iota-and nu-carrageenan. They represent 70% of the dry matter of the alga in summer. After alkaline transformation the proportion of iota-carrageenan increased to 76%, demonstrating the presence of nu-carrageenan. Absence of mu-carrageenan, the precursor of kappa-carrageenan, suggests that iota-carrageenan is desulfated enzymically to kappa-carrageenan.