Contents
Cover
Half Title page
Title page
Copyright page
Preface
Chapter 1: Marine Polymers
1.1 Marine Microbes
1.2 Marine Microgels
1.3 Polymer Production from Marine Algae
1.4 Marine Bioadhesive Analogs
1.5 Medical Applications
1.6 Polymer Production from Marine Sponge
1.7 Chitin and Chitosan from Marine Origin
1.8 Carbohydrates
1.9 Poly(3-hydroxy butyrate) from Marine Bacteria
1.10 Metal Ion Absorption
1.11 Fish Elastin Polypeptide
1.12 Cosmetic Uses
1.13 Protein Hydrolyzate
References
Chapter 2: Marine Applications
2.1 Marine Polymer Coatings
2.2 Foams
2.3 Antifouling
2.4 Electrochemical Impedance and Noise Data for Polymer Coated Steel
2.5 Seawater Immersion Ageing of Glass-Fiber Reinforced Polymer Laminates
2.6 Post-Fire Mechanical Properties of Marine Polymer Composites
2.7 Corrosion
2.8 Marine Ropes
2.9 Marine Diesel Engine Lubricants
2.10 Lubricant for Smoothing Caulking Joints
2.11 Marine Well Applications
References
Chapter 3: Waterborne Polymers
3.1 Analytical and Characterization Techniques
3.2 Synthesis Methods
3.3 Aqueous Dispersions of Pigments
3.4 Waterborne Coatings
3.5 Special Applications
References
Chapter 4: Water-Resistant Polymers
4.1 Coatings
4.2 Biodegradable Resins
4.3 Water-Based Printing Inks
4.4 Reinforcing Fibers
4.5 Paper Industry Applications
4.6 Masonry Products
4.7 Medical Uses
4.8 Membranes
4.9 Personal Care Compositions
4.10 Package Uses
4.11 Grouting Compositions
4.12 Xerogels
References
Index
General Index
Marine, Waterborne and Water-Resistant Polymers
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Co-published by John Wiley & Sons, Inc. Hoboken, New Jersey, and Scrivener Publishing LLC, Salem, Massachusetts.
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Library of Congress Cataloging-in-Publication Data:
ISBN 978-1-119-18486-7
Preface
This book focuses on the chemistry of marine polymers, waterborne polymers, and water resistant polymers, as well as special applications of these materials.
After an introductory section on the general aspects of the field, the types and uses of these polymers are summarized, followed by an overview of some testing methods.
In passing, as it so happens in the literature for these types of polymers, a lot of special organic compounds are used which for the ordinary organic and polymer chemist are not too familiar. Therefore, the structures of these organic compounds are reproduced in many of the figures.
The text focuses on the literature of the past decade. Beyond education, this book may serve the needs of industry engineers and specialists who have only a passing knowledge of these issues, but need to know more.
Utmost care has been taken to present reliable data. Because of the vast variety of material presented here, however, the text cannot be complete in all aspects, and it is recommended that the reader study the original literature for more complete information.
The reader should be aware that mostly US patents have been cited where available, but not the corresponding equivalent patents of other countries. In particular, in this field of science, most of the original patents are of Japanese origin.
For this reason, the author cannot assume responsibility for the completeness, validity or consequences of the use of the material presented here. Every attempt has been made to identify trademarks; however, there were some that the author was unable to locate.
There are three indices: an index of acronyms, an index of chemicals, and a general index. In the index of chemicals, compounds that occur extensively, e.g., acetone, are not included at every occurrence, but rather when they appear in an important context. When a compound is found in a figure, the entry is marked in boldface letters in the chemical index.
I am indebted to our university librarians, Dr. Christian Hasenhüttl, Dr. Johann Delanoy, Franz Jurek, Margit Keshmiri, Dolores Knabl, Friedrich Scheer, Christian Slamenik, Renate Tschabuschnig, and Elisabeth Groß for their support in literature acquisition. In addition, many thanks to the head of my department, ProfessorWolfgang Kern, for his interest and permission to prepare this text.
I also want to express my gratitude to all the scientists who have carefully published their results concerning the topics dealt with herein. This book could not have been otherwise compiled.
Last, but not least, I want to thank the publisher, Martin Scrivener, for his abiding interest and help in the preparation of the text. In addition, my thanks go to Jean Markovic, who made the final copyedit with utmost care.
Johannes Fink
Leoben, July 15, 2015
An overview of the methods, applications, and products of marine biotechnology has been presented (1). A large portion of the surface of the earth is covered by the ocean. Therefore, more than 80% of living organisms can be found in aquatic ecosystems. Thus, these organisms constitute a rich reservoir for various chemical materials and biochemical processes.
The literature for marine natural products has been extensively reviewed (2–4). Compounds isolated from marine microorganisms and phytoplankton, green, brown and red algae, sponges, cnidarians, bryozoans, mollusks, tunicates, echinoderms, mangroves and other intertidal plants and microorganisms have been collected. Also, biosynthetic studies, and syntheses that lead to the revision of structures or stereochemistries, have been dealt with (4).
Biochemical materials and processes from marine sources make these materials available to applications in pharmaceuticals, cosmeceuticals or nutraceuticals, as well as for the production of biopolymers, bioenergy and biofuels (1). Also, biomaterials from marine-origin biopolymers have been reviewed (5).
The marine microbial biosphere is a large resource of biotechnological interest. The potential of marine microbes in biotechnology has been reviewed (6).
The biotechnological potential ranges from the synthesis of bioactive molecules to the production of biofuels, cosmeceuticals, nutraceuticals, and biopolymers.
Marine microbes can be used for biomedical purposes and also for the degradation of pollutants. Marine viruses have a great biotechnological potential. Marine archaea have been exploited for the isolation of enzymes. Also, bacteria and microbial eukaryotes are of importance for biotechnological uses (6).
The ocean plays a critical role in the global carbon cycle (7). It handles around 50% of the global primary production, yielding the world’s largest stock of reduced organic carbon (ROC) that supports one of the world’s largest biomasses. However, the mechanisms whereby ROC becomes mineralized remain unresolved. A review has been presented that focuses on laboratory and field observations that of dissolved organic carbon (DOC) self-assembles, and the formation of self-assembled microgels (SAGs).
Self-assembly has approximately 10% yield, generating an estimated global seawater SAG budget of some 1016 g carbon. The transects at depths of 10 to 4,000 m reveal concentrations of 106 to 3 × 1012 SAG l–1, thus respectively forming an estimated ROC stock larger than the global marine biomass (7).
Because hydrogels have 1% solids, i.e., 10 gl–1, whereas seawater DOC reaches only 10–3 gl–1, SAGs contain 104 more bacterial substrate than seawater.
For this reason, microgels represent an unsuspected and huge micron-level ocean patchiness that could profoundly influence the passage of DOC through the microbial loop, with ramifications that may scale to global cycles of bioactive elements (7).
The broad class of polymeric materials includes polymers with excellent processability, chemical resistance, and mechanical properties. These properties allow polymers to be used to produce extrusion molded articles, injection molded articles, hollow molded articles, films, sheets, among many others.
Numerous polymers are derived from petroleum and natural gas. Actually, the market prices for these fossil fuels are increasing due to a number of factors, including a depletion of easily accessible deposits, growth of emerging economies, political instabilities, and environmental concerns. Therefore, polymer production methods that do not rely on fossil fuels are desirable (8).
The production of biopolymers from algae has already been described in 1975 (9). Long-chain polymers with flocculating properties have been produced.
The special issues of algae have been summarized (10). Algae are a very large and diverse group of eukaryotic organisms, ranging from unicellular genera and the diatoms to multicellular forms. An eukaryote is an organism whose cells contain a nucleus and other organelles enclosed within membranes (11).
Algae can produce 10–100 times more mass then terrestrial plants in the course of a year. Algae also produce oils and starches that may be converted into biofuels. Algae useful for biofuel production are also microalgae, consisting of small, often unicellular, types.
These algae can grow almost everywhere, but are most commonly found at latitudes between 40 degrees north and 40 degrees south. The algae can grow rapidly in nearly any environment, with almost any kind of water, including marginal areas with a limited or poor quality water. Micrographs from the algae are shown in Figure 1.1.
It has been found that certain algae species of the Isochrysis family produce polyunsaturated long-chain alkenones. In the studies, methyl and ethyl alkenones with 35–40 carbons having 2–4 double bonds have been detected.
Lipid-producing algae can include a wide variety of algae. Algae are classified as follows: diatoms are bacillariophytes, green algae are chlorophytes, blue-green algae are cyanophytes, golden-brown algae are chrysophytes, and phylum of algae are haptophytes. Most common algae are listed in Table 1.1.
Bacillariophytes | Chlorophytes |
Amphipleura | Ankistrodesmus |
Amphora | Botryococcus |
Chaetoceros | Chlorella |
Cyclotella | Chlorococcum |
Cymbella | Dunaliella |
Fragilaria | Monoraphidium |
Hantzschia | Oocystis |
Navicula | Scenedesmus |
Nitzschia | Tetraselmis |
Phaeodactylum | |
Thalassiosira | |
Cyanophytes | Haptophytes |
Oscillatoria | Isochrysis |
Synechococcus | Pleurochrysis |
Methods have been developed for producing polymers from algae. Such methods comprise (8):
The alkenone-producing alga can be a species of the Isochrysis family, such as Isochrysis galbana, Isochrysis sp. T-Iso, and Isochrysis sp. C-Iso. The alkenones of the alga should be alkenones with a number of carbons of 35–40. The alkenones may be converted into hydrocarbons by catalytic hydroprocessing.
Eventually, the alkenones can be converted into a liquid fuel such as diesel and gasoline. Likewise, the alkenones may be processed into a gaseous fuel such as synthesis gas or methane, propane, and butane. The alga may also deliver fatty acid methyl esters.
The growth condition for culturing the alga may include a stationary growth phase, a high temperature, sufficient light, and nutrient limitation. The algae can also be directly converted into methane via a hydrothermal gasification process (8).
Algae store lipids inside the cell body, sometimes but not always in vesicles (8). The lipids can be recovered in various ways, including solvents, heat, pressure, or depolymerization, such as by biologically breaking the walls of the algal cell or oil vesicles.
At least one of three types of biological agents may be used to release algae energy stores, for example, enzymes such as cellulase or glycoproteinase, structured enzyme arrays or system such as a cellulosome, a viral agent, or a combination of these agents.
A cellulase is an enzyme that breaks down cellulose, especially in the wall structures. A cellulosome is an array or sequence of enzymes or cellulases which is more effective and faster than a single enzyme or cellulase. In both cases, the enzymes break down the cell wall or lipid vesicles and release lipids from the cell.
Cellulases used for this purpose may be derived from fungi, bacteria, or yeast. Examples include cellulase produced by fungus Trichoderma reesei and many genetic variations of this fungus, cellulase produced by the bacteria genus Cellulomonas, and cellulase produced by the yeast genus Trichosporon.
A glycoproteinase provides the same function as a cellulase, but is more effective on the cell walls of microalgae, many of which have a structure more dependent on glycoproteins than cellulose (8).
A process for converting the algal alkenones into hydrocarbons is catalytic hydroprocessing, or cracking (8). The catalytic hydroprocessing technology is well known in the art of petroleum refining and generally refers to converting at least large hydrocarbon molecules into smaller hydrocarbon molecules by breaking the carboncarbon bonds (12).
The long chains of carbon in the alkenones produced by algae can be used to produce a wider range of biofuels or lubricating oils than those derived from glycerides (8).
The algal lipids can be used as feedstock in the industrial chemical field, particularly in the manufacture of polymers. The algal lipids can be polymerized, either directly or after some chemical modification (8).
Also, the algal lipids can be pyrolyzed or cracked into smaller molecules to permit the generation of standard monomers such as acrylic acids and esters, alkenes, vinyl chloride, vinyl acetate, diacids, diamines, diols, lactic acid, and others (8).
It has been shown that a phenolic polymer extracted from Fucus serratus can be crosslinked using a vanadium-dependent bromoperoxidase (13). The methanol extracted phenolic polymer was adsorbed onto a quartz crystal sensor and the crosslinking was initiated by the addition of bromoperoxidase, KBr, and H2O2.
The decreased dissipation upon addition of the crosslinking agents, as measured with the quartz crystal microbalance with the quartz crystal microbalance dissipation (QCM-D), was interpreted as intramolecular crosslinks being formed between different phloroglucinol units in the phenolic polymer.
With surface plasmon resonance, it was shown that no desorption occurred from the sensor surface during the crosslinking reaction. UV/VIS spectroscopy verified the results achieved with QCM-D that all components, i.e., bromoperoxidase, KBr, and H2O2, were necessary in order to achieve intramolecular oxidative crosslinking of the polymer (13).
Nature has evolved materials that possess mechanical properties surpassing many manmade composites. A nanostructured composite film that takes advantage of two different natural materials has been prepared (14). These materials are layered nacre and the marine adhesive of mussels.
L-3,4-Dihydroxyphenylalanine molecules impart an unusual adhesive strength to a clay composite and the hardening mechanism found in the natural cement plays an equally important role in the strengthening of the nanostructured nacre (14).
The in-situ molecular physicochemical characterization of bioadhesives at solid or liquid interfaces has been reviewed (15). The adhesion strategies that lie at the root of marine biofouling have been elucidated. Three major fouling organisms have been assessed: mussels, algae and barnacles.
The dispersal of these organisms, their colonization on the surfaces, and ultimately their survival are critically dependent on the ability of the larvae or spores of the organisms to locate a favorable settlement site and undergo metamorphosis. In this way their sessile existence is initiated.
Differences in the composition of the adhesive secretions and the strategies employed for their temporary or permanent implementation between the larval and adult life stages are obvious.
Until now, only a few adhesive secretions from marine fouling organisms have been adequately described in terms of their chemical composition. The presence of certain recurrent functional groups, specifically catechol, carboxylate, monoester-sulphate and -phosphate are used for this purpose. The binding modes of such functionalities to wet mineral and metal oxides surfaces have been described in detail.
A plausible explanation for the propensity of these adhesive functionalities to bind to hydrous metal oxide surfaces has been based on the basis of the hard and soft acids and bases principle, Hofmeister effects and entropic considerations.
From the in-situ analysis of marine organism bioadhesives and adsorption studies of functionalities relevant to the bioadhesion process, insights suitable for antifouling strategies and the synthesis of durable adhesive materials can be obtained (15).
Marine organisms are constituted by materials with a vast range of properties and characteristics that may justify their potential application within the biomedical field (16). Moreover, assuring the sustainable exploitation of natural marine resources, the valorization of residues from marine origin, like those obtained from food processing, constitutes a highly interesting platform for the development of novel biomaterials, with both economic and environmental benefits.
In the last decade, many different biomaterials, like various types of polymers and bioactive ingredients, have been identified, isolated, and characterized. These biomaterials can be used in controlled drug delivery, tissue engineering, and diagnostic devices (17).
In this perspective, an increasing number of different types of compounds can be isolated from aquatic organisms and transformed into profitable products for health applications, including controlled drug delivery and tissue engineering devices.
The issues of marine structural proteins in biomedicine and tissue engineering have been reviewed (18). Actually, the development of biocompatible composites and vehicles of marine biopolymer origin for growth, retention, delivery, and differentiation of stem cells is of crucial importance for regenerative medicine.
Also, the current techniques for the isolation and characterization of polysaccharides, proteins, glycosaminoglycans and ceramics from marine raw materials have been reviewed (16). Specific compound classes for medical applications are listed in Table 1.2.
Compound class | Compound class |
Agar | Collagen |
Alginates | Chondroitin sulfate |
Carrageenans | Heparin |
Chitin | Hyaluronic acid |
Chitosan | Calcium phosphorous compounds |
Glycosaminoglycans | Biosilica |
Marine-derived bioactive compounds for breast and prostate cancer treatment have been reviewed (19). Marine-derived natural bioactive products, isolated from aquatic fungi, cyanobacteria, sponges, algae, and tunicates, have been found to exhibit various anticancer activities including anti-angiogenic, anti-proliferative, inhibition of topoisomerase activities and induction of apoptosis.
Matrix metalloproteinases are endopeptidases which belong to the group of metalloproteinases that contribute for the extracellular matrix degradation and several tissue remodeling processes (20). An imbalance in the regulation of these endopeptidases may cause severe pathological complications like cancer, cardiac, cartilage, and neurological-related diseases (21).
Synthetic metalloproteinases have some shortcomings. Therefore, many of them cannot be used in clinical applications. Thus, a growing interest metalloproteinases from marine origin has been taking place. Potential matrix metalloproteinase inhibitors from edible marine algae have been reviewed (21).
An imbalance in the regulation of these endopeptidases eventually results in severe pathological complications like cancer, cardiac, cartilage, and neurological-related diseases.
Fucoidans are a class of sulfated, fucose-rich polymers found in brown macroalgae (22, 23). They were identified in the early 20th century by Harald Kylin (24).
Subsequently, the class of these compounds was detailed (25, 26). The systematic isolation of fucoidan from a number of different British seaweeds has been described (27).
Fucoidan was recognized as having a role in the biology of seaweed, and was examined over the next few decades for its activity in a number of biological systems.
The uses of fucoidan as a therapeutic target for cancer have been reviewed (28). Fucoidan was shown to induce a cytotoxicity of various cancer cells; it induces apoptosis, and inhibits invasion, metastasis and angiogenesis of cancer cells. The principle of apoptosis was already described in 1842. It is a process of programmed cell death (29,30). Apoptosis is a critical defense mechanism against the formation and progression of cancer and exhibits distinct morphological and biochemical traits. Targeting the apoptotic pathways has become an important strategy for the development of chemotherapeutic agents. Marine natural products have become important sources in the discovery of antitumor drugs. The effects of selected marine natural products and their synthetic derivatives on apoptosis signaling pathways have been reviewed (31).
Fucoidans from marine algae are also potential matrix metalloproteinase inhibitors. Inhibitory substances of metalloproteinases could be beneficial in the management of pathological events (20).
The current research interest in fucoidan is now global. Research in occurring in Australia, Japan, Korea, Russia and China, in addition to Europe and the American countries. The intensity of biological activities of fucoidan varies with species, molecular weight, composition, structure and the route of administration.
The literature concerning fucoidan drugs has increased considerably in the last decade. These algal-derived marine carbohydrate polymers present numerous valuable bioactivities.
The role of fucoidan in the control of acute and chronic inflammation via selectin blockade, enzyme inhibition and inhibition of the complement cascade has been reviewed (32–34). Most recent data on toxicology and uptake of fucoidan have been detailed together with a discussion on the comparative activities of fractions of fucoidan from different sources. The targets of of fucoidan-derived drugs include (34):
Sulfated fucans and galactans are strongly anionic polysaccharides that are found in marine organisms (33). Their structures vary among the various species, but their major features are conserved among phyla. Sulfated fucans are found in marine brown algae and echinoderms. In contrast, sulfated galactans occur in red and green algae, marine angiosperms, tunicates (ascidians), and sea urchins.
Polysaccharides with 3-linked, ß-galactose units are highly conserved in some taxonomic groups of marine organisms and show a strong tendency toward 4-sulfation in algae and marine angiosperms, and 2-sulfation in invertebrates.
Marine algae mainly express sulfated polysaccharides with complex, heterogeneous structures, whereas marine invertebrates synthesize sulfated fucans and sulfated galactans with regular repetitive structures.
These polysaccharides are structural components of the extracellular matrix. Sulfated fucans and galactans are involved in sea urchin fertilization, acting as species-specific inducers of the sperm acrosome reaction. Galactan is a polysaccharide consisting of polymerized galactose.
The algal and invertebrate polysaccharides are also potent anticoagulant agents of mammalian blood and represent a potential source of compounds for antithrombotic therapies (33). α-L-Fucopyranose and galactose are shown in Figure 1.2.
The issue of marine organisms for bone repair and regeneration has been reviewed (35). Various types of polymer scaffolds have been described (36). Chitosan-alginate and chitosan-alginate with fucoidan, were developed by a freeze-drying method. Each of these materials was characterized as a bone graft substitute. The porosity, water uptake and retention ability of the prepared scaffolds showed a similar behavior.
The pore size of the chitosan-alginate and chitosan-alginate with fucoidan scaffolds were measured from scanning electron microscopy and found to be 62–490 μm and 56–437 μm, respectively.
It has been suggested that hydrogen bonding or ion-ion pair interaction between these components usually increases the uniform dispersion (37).
In-vitro studies revealed a more profound cytocompatibility, increased cell proliferation and enhanced alkaline phosphatase secretion in the chitosan-alginate with fucoidan scaffold in comparison to the chitosan-alginate scaffold (36).
Further, protein adsorption and mineralization were about two times greater in the chitosan-alginate with fucoidan scaffold than for the chitosan-alginate scaffold. Therefore, it has been concluded that chitosan-alginate with fucoidan will be a promising biomaterial for bone tissue regeneration (36).
Chitosan composites for bone tissue engineering have been reviewed (38). Bone contains considerable amounts of minerals and proteins. Hydroxyapatite, Ca10(PO4)6(OH)2, is one of the most stable forms of calcium phosphate and it occurs in bones in an amount of 60–65%, together with other materials such as collagen, chondroitin sulfate, keratin sulfate and lipids.
Chitosan has always played a major role in bone tissue engineering. It is a natural polymer that can be obtained from chitin, which forms a major component of the crustacean exoskeleton.
Considerable attention has been given to chitosan composite materials and their applications in the field of bone tissue engineering due to its minimal foreign body reactions, an intrinsic antibacterial nature, biocompatibility, biodegradability, and the ability to be molded into various geometries and forms such as porous structures, suitable for cell ingrowth and osteoconduction.
Composites of chitosan with hydroxyapatite is very popular because of their biodegradability and biocompatibility. Grafted chitosan polymers with carbon nanotubes have been incorporated into the compositions in order to increase the mechanical strength.
Chitosan and hydroxyapatite are among the best bioactive biomaterials in bone tissue engineering and are renowned for their excellent biocompatibility with the human body environment (39).
The recent technological advances with regard to the isolation and manufacture of nanofibrillar chitin and chitosan have been reviewed (40). Chitin and chitosan are obtained either by mechanical chitin disassembly and fibrillation optionally assisted by sonication, or by electro-spinning. Nanosized materials have better performances.
Chemically modified or nanofibrous chitin and chitosan have been developed, and their effects on wound healing have been evaluated. These compounds are beneficial for the wound healing process (41).
The biomedical applications of chitin include hemostasis and wound healing, regeneration of tissues such as joints and bones, cell culture, antimicrobial agents, and dermal protection.
Other biomedical applications of chitosan are epithelial tissue regeneration, bone and dental tissue regeneration, and also protection against bacteria, fungi and viruses (40, 41). In addition, chitins and chitosans can be used as immunoadjuvants and nonallergenic drug carriers (42).
Collagen is the most abundant protein of animal connective tissues (43). It is found in skin, bone or cartilage. Therefore, it is one of the key polymers for biomedical applications, i.e., tissue engineering and drug delivery.
Marine sponges are extremely rich in natural products and are considered a promising biological resource (44). Marine sponge collagen has unique physicochemical properties, but its application is difficult, due to the lack of availability because of inefficient extraction methodologies. The traditional extraction methods are time consuming since they involve several operating steps and large amounts of solvents.
An extraction methodology under mild operating conditions has been proposed, in which water is acidified with carbon dioxide to promote the extraction of collagen or gelatin from different marine sponge species (44).
Actually, there are various reasons to think about marine gelatin as an alternative to the terrestrian gelatin, for example, the risk of transmission of infectious diseases (45). So, an advantage of marine gelatin is that it has no risk associated with bovine spongiform encephalopathy. Practical applications for fish gelatins in the form of gels, films and composite materials have been discussed (45).
Porous structures from marine collagen crosslinked with genipin under high-pressure carbon dioxide have been fabricated (46). Collagen from shark skin has been used to prepare pre-scaffolds by freeze-drying. The poor stability and the low mechanical properties require the crosslinking for scaffold uses.
In a dense carbon dioxide atmosphere, the crosslinking of collagen pre-scaffolds has been done for 16 h. Then, the hydrogels have been foamed and the thus obtained scaffolds showed a highly porous structure.
In-vitro cell culture tests using chondrocyte-like cell line showed a good cell adherence and proliferation. Therefore it has been suggested to use these materials in tissue cartilage (46).
Industrial procedures to extract collagen use bovine and porcine as their main sources. A source of marine origin is one of the alternatives that has been explored through byproducts of fish processing.
In a study, collagen has been extracted from the skin of the shark Scyliorhinus canicula. The thus obtained collagen was evaluated as an alternative for dermal membranes, regarding the sustained release of drugs. The method used for the isolation, as well as the method used for membrane preparation have been described in detail (43).
For the preparation of membranes for drug delivery assessment, dexamethasone was added to the collagen solutions and membranes were prepared. The collagen membranes were characterized by the water contact angle, mechanical properties and their stability in phosphate-buffered saline. Collagen membranes showed more stability in phosphate-buffered saline as long as the degree of crosslinking is higher, which also influences their mechanical properties. The degree of crosslinking also affects the hydrophobicity of the membranes. The properties of the collagen membranes can be tailored by degree of crosslinking (43).
The use of shark collagen as a matrix for cell culture and as a substrate for zymography has been investigated (47). Zymography is an electrophoretic technique for the detection of hydrolytic enzymes, which is based on the substrate repertoire of the enzyme (48).
Fibroblasts were cultured on a gel matrix of shark type I collagen at 30°C. The collagen gel contracted by 4 d of incubation. Individual fibroblasts were visible against the transparent background of the contracted collagen as long, lean star-shaped cells (47). The matrix metalloproteinases from the fibroblasts secreted from the medium digested shark gelatin more easily than pig gelatin.
Exopolysaccharides are high molecular weight polymers that are composed of sugar residues and are secreted by a microorganism into the surrounding environment (49).
Glycosaminoglycans are glycopolymers found in animal tissues and are composed of uronic acid and neutral or hexosamine residues (50). They are covalently bound to a core protein and are the major constituent of proteoglycans. Their carbohydrate backbone is unique for each cell type. Glycosaminoglycans are essential for the life of animals since they are involved in their development and organogenesis (51). Two glycosaminoglycan macromolecules are frequently used in various industrial fields, namely hyaluran and heparin (50).
Several exopolysaccharide-producing marine strains have been studied, which led to the discovery and isolation of new macromolecules (52).
An exopolysaccharide (HE800 EPS) is secreted by a deep-sea hydrothermal bacterium and displays an interesting glycosaminoglycan-like feature resembling hyaluronan (53). Hyaluronan is the new term for hyaluronic acid.
Its effectiveness in enhancing in-vivo bone regeneration and in supporting osteoblastic cell metabolism in culture has been demonstrated. in-vitro reconstructed connective tissues containing the HE800 EPS were achieved. This polysaccharide promotes both collagen structuring and extracellular matrix settle by dermal fibroblasts.
A low-molecular-weight sulfated derivative displays a chemical analogy with heparan sulfate (53). These derivatives can be obtained by means of free radical depolymerization of a native EPS followed by sulfating the resulting depolymerized derivatives (54).
Exopolysaccharides also find applications in environmental biotechnology where they are used for soil and water bioremediation, decontamination and detoxification processes (55, 56).