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The rapid ionic crosslinking of alginate has been actively studied for biomedical applications including hydrogel scaffolds for tissue engineering, injectable gels, and 3D bioprinting. However, the poor structural stability of ionic crosslinks under physiological conditions limits the widespread applications of these hydrogels. Moreover, the lack of cell adhesion to the material combined with the inability of proteases to degrade alginate further restrict utility as hydrogel scaffolds. Blends of alginate with silk fibroin have been proposed for improved structural and mechanical properties, but potential phase separation between the hydrophobic protein and the hydrophilic polysaccharide remains an issue. In this study, we demonstrated the synthesis of a hybrid biopolymer composed of a silk backbone with side chains of poly(guluronate) isolated from alginate to introduce rapid ionic crosslinking on enzymatically crosslinked silk-based hydrogels for on-demand and reversible stiffening and softening properties. Dual crosslinked macro- and microgels of silk fibroin-poly(guluronate) (SF-PG) hybrid polymers displayed dynamic morphology with reversible shrinking and swelling behavior. SF-PG hydrogel discs demonstrated dynamic mechanics with compressive moduli ranging from less than 5 kPa to over 80 kPa and underwent proteolytic degradation unlike covalently crosslinked alginate controls. SF-PG gels supplemented with gelatin substituted with tyramine or both tyramine and PG also supported the attachment and survival of murine fibroblasts, suggesting potential uses of these new hydrogels in mammalian cell culture to investigate cellular responses to dynamic mechanics or modeling of diseases defined by matrix mechanics, such as fibrosis and cancer.

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Introduction: Developing a culture system that can effectively maintain chondrocyte phenotype and functionalization is a promising strategy for cartilage repair. Methods: An alginate/collagen (ALG/COL) hybrid hydrogel using different guluronate/mannuronate acid ratio (G/M ratio) of alginates (a G/M ratio of 64/36 and a G/M ratio of 34/66) with collagen was developed. The effects of G/M ratios on the properties of hydrogels and their effects on the chondrocytes behaviors were evaluated. Results: The results showed that the mechanical stiffness of the hydrogel was significantly affected by the G/M ratios of alginate. Chondrocytes cultured on Mid-G/M hydrogels exhibited better viability and phenotype preservation. Moreover, RT-qPCR analysis showed that the expression of cartilage-specific genes, including SOX9, COL2, and aggrecan was increased while the expression of RAC and ROCK1 was decreased in chondrocytes cultured on Mid-G/M hydrogels. Conclusion: These findings demonstrated that Mid-G/M hydrogels provided suitable matrix conditions for cultivating chondrocytes and may be useful in cartilage tissue engineering. More importantly, the results indicated the importance of taking alginate G/M ratios into account when designing alginate-based composite materials for cartilage tissue engineering.

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Alginate oligosaccharides are degradation products of alginate and have attracted increasing attention due to their versatile biological functions. In the present study, C57BL/6 mice were used to assess the ameliorative effects and mechanisms of guluronate oligosaccharides (GAOS), mannuronic oligosaccharides (MAOS), and heterozygous alginate oligosaccharides (HAOS), which are the three alginate oligosaccharides of dextran sulfate sodium (DSS)-induced ulcerative colitis. The study showed that alginate oligosaccharides alleviated pathological histological damage by slowing down weight loss, inhibiting colonic length shortening, and reducing disease activity index (DAI) and histopathological scores. Alginate oligosaccharides modulated the colonic inflammatory response by reducing colonic MPO levels and downregulating the expression of IL-6 and IL-1ß. Alginate oligosaccharides reduced intestinal permeability and reversed intestinal barrier damage by increasing the number of goblet cells, decreasing LPS levels, downregulating Bax protein levels, upregulating Bcl-2 protein levels, and enhancing the expression of the E-cadherin. Furthermore, alginate oligosaccharides modulated the composition of the gut microbiota and restored the production of short-chain fatty acids (SCFAs), especially acetate and butyrate. In conclusion, our study provides a scientific basis for the role of alginate oligosaccharides in relieving ulcerative colitis.

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Unsaturated guluronate oligosaccharide (GOS) is generated via alginate-derived polyguluronate (PG) degradation by alginate lyase, followed by formation of a double bond between C-4 and C-5 at the nonreducing end. In this study, GOS was first used as a stabilizer to fabricate O/W nanoemulsions loaded with resveratrol (GOS-RES). Our results revealed that both the GOS-RES and normal O/W resveratrol nanoemulsions (water-RES) showed small droplet sizes and narrow size distributions under certain experimental conditions. However, the particle size and stability of the GOS-RES were slightly greater than those of the water-RES in acidic and neutral environments and at high temperatures. Furthermore, the GOS-RES exhibited a better sustained release effect for resveratrol than the water-RES. Moreover, the GOS-RES showed a significant superoxide radical scavenging effect. All these results demonstrated that GOS has good prospects for preparing nanoemulsions to encapsulate hydrophobic nutrients, which could be applied as food-grade components in beverages and other foods.

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Alginate oligosaccharides (AOS) are degradation products of alginate extracted from brown algae. With low molecular weight, high water solubility, and good biological activity, AOS present anti-inflammatory, antimicrobial, antioxidant, and antitumor properties. They also exert growth-promoting effects in animals and plants. Three types of AOS, mannuronate oligosaccharides (MAOS), guluronate oligosaccharides (GAOS), and heterozygous mannuronate and guluronate oligosaccharides (HAOS), can be produced from alginate by enzymatic hydrolysis. Thus far, most studies on the applications and biological activities of AOS have been based mainly on a hybrid form of HAOS. To improve the directional production of AOS for practical applications, systematic studies on the structures and related biological activities of AOS are needed. This review provides a summary of current understanding of structure-function relationships and advances in the production of AOS. The current challenges and opportunities in the application of AOS is suggested to guide the precise application of AOS in practice.

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Unsaturated guluronate oligosaccharide (GOS) was prepared from alginate-derived homopolymeric blocks of guluronic acid by alginate lyase-mediated depolymerization. In this study, a GOS-based water-in-oil-in-water (W1/O/W2) nanoemulsion was prepared, and different influencing factors were investigated. First, linseed oil was selected as the optimal carrier oil. Then, other optimal conditions of the GOS nanoemulsion were determined based on response surface methodology (RSM). Under the optimal conditions, the obtained GOS nanoemulsion showed a spherical structure with an average particle size of 273.93 ±â€¯8.91 nm, and its centrifugal stability was 91.37 ±â€¯0.45 %. Moreover, the GOS nanoemulsion could achieve the aim of sustained release in vitro and be stably stored at 4°C for at least 5 days. This work prepared a novel GOS-based W1/O/W2 nanoemulsion that may effectively address the storage difficulties of unsaturated GOS and provides a valuable contribution to the application of GOS in the food and medicine fields.

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Alginates are one of the major polysaccharide constituents of marine brown algae in commercial manufacturing. However, the content and composition of alginates differ according to the distinct parts of these macroalgae and have a direct impact on the concentration of guluronate and subsequent commercial value of the final product. The Azotobacter vinelandii mannuronan C-5 epimerases AlgE1 and AlgE4 were used to determine their potential value in tailoring the production of high guluronate low-molecular-weight alginates from two sources of high mannuronic acid alginates, the naturally occurring harvested brown algae (Ascophyllum nodosum, Durvillea potatorum, Laminaria hyperborea and Lessonia nigrescens) and a pure mannuronic acid alginate derived from fermented production of the mutant strain of Pseudomonas fluorescens NCIMB 10,525. The mannuronan C-5 epimerases used in this study increased the content of guluronate from 32% up to 81% in both the harvested seaweed and bacterial fermented alginate sources. The guluronate-rich alginate oligomers subsequently derived from these two different sources showed structural identity as determined by proton nuclear magnetic resonance (1H NMR), high-performance anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) and size-exclusion chromatography with online multi-angle static laser light scattering (SEC-MALS). Functional identity was determined by minimum inhibitory concentration (MIC) assays with selected bacteria and antibiotics using the previously documented low-molecular-weight guluronate enriched alginate OligoG CF-5/20 as a comparator. The alginates produced using either source showed similar antibiotic potentiation effects to the drug candidate OligoG CF-5/20 currently in development as a mucolytic and anti-biofilm agent. These findings clearly illustrate the value of using epimerases to provide an alternative production route for novel low-molecular-weight alginates.

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Sodium salts of the algal uronic-acids, d-mannuronic acid (HManA) and l-guluronic acid (HGulA) have been isolated and characterised in solution by nuclear magnetic resonance (NMR) spectroscopy. A suite of recently-described NMR experiments (including pure shift and compressive sampling techniques) were used to provide confident assignments of the pyranose forms of the two uronic acids at various pD values (from 7.5 to 1.4). The resulting high resolution spectra were used to determine several previously unknown parameters for the two acids, including their pKa values, the position of their isomeric equilibria, and their propensity to form furanurono-6,3-lactones. For each of the three parameters, comparisons are drawn with the behaviour of the related D-glucuronic (HGlcA) and D-galacturonic acids (HGalA), which have been previously studied extensively. This paper demonstrates how these new NMR spectroscopic techniques can be applied to better understand the properties of polyuronides and uronide-rich macroalgal biomass.

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Association constants and thermodynamicsparameters on inclusion of four 3-amido coumarins that present trypanocidal activity, into 6-amino-ß-cyclodextrin (1:1 stoichimetry) were determined. In addition, pure homopolymeric-α-l-guluronate fraction prepared by partial hydrolysis of sodium alginate from Sub-Antarctic Kelp Durvillaea antarctica was conjugated with 6-amino-ß-cyclodextrin (64% yield). To glycoconjugates, 3-amido coumarins were incorporated (73% of encapsulation) and supramolecular hydrogels were prepared by gelation with Ca2+ ions. The trypanocidal activity of the inclusion complexes increased by 10%. Likewise, an increase in diffusion in artificial membrane was observed (13%). It was found that the inclusion complexes increased the variation of the mitochondrial potential of T. cruzi (17%). The lowest release of substituted amidocoumarins (ACS) from supramolecular hydrogels occurred at pH 1.2 whereas the maximum release (34%) was observed at pH 8.0. Encapsulation of lipophilic bioactive compounds in supramolecular hydrogels allows the generation of release systems sensitive to pH with potential application in biomedicine.

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Alginate-degrading enzyme, alginate lyase, catalyzes the cleavage of glycosidic 1-4 O-linkages between uronic acid residues of alginate by a ß-elimination reaction leaving a 4-deoxy-l-erythro-hex-4-ene pyranosyluronate as nonreducing terminal end. The enzymes from a wide variety of sources such as marine molluscs, seaweeds, and marine bacteria have been discovered and studied not only from a point of view of enzymological interest of enzyme itself but also for elucidation of fine chemical structure of alginate, structure-activity relationship of alginate, and biological activities and physicochemical features of the enzymatic digestion products. Based on the substrate specificities, alginate lyases are classified into three groups: poly(ß-d-mannuronate) lyase, poly(α-l-guluronate) lyase, and bifunctional alginate lyase, which are specific to mannuronate, guluronate, and both uronic acid residues, respectively. We have studied enzymological aspects of these three types of alginate lyases, and bioactivities of enzymatically digested alginate oligomers. In this chapter, we described the purification and characterization of three types of alginate lyases from different marine origins and overviewed the bioactivities of alginate oligomers.

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Low molecular weight and sulfated low molecular weight guluronate (LMG and SLMG) were prepared and hypolipidemic effects were studied in a human hepatocellular carcinoma HepG2 cell line. Both compounds decreased total cholesterol (TC) and triglycerides (TG) and inhibited 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR) activity in HepG2 cells. In general, SLMG had greater effects than LMG. Activation of sterol regulatory element-binding protein 2 (SREBP-2), low density lipoprotein receptor (LDLR), AMP-activated protein kinase (AMPK), and AMPK's downstream targets were evidenced by increased phosphorylation of AMPK, HMGCR, and acetyl-CoA-carboxylase (ACC), which decreased HMGRC and ACC activity. We further demonstrated that activated AMPK was linked to down-regulated SREBP-1 and up-regulated cholesterol 7α-hydroxylase (CYP7A1).

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To gain insight into the structure-activity relationship of alginate, we examined the effect of alginates with varying molecular weights and M/G ratio on murine macrophage cell line, RAW264.7 cells in terms of induction of tumor necrosis factor-α (TNF-α) secretion. Among the alginates tested, alginate with the highest molecular weight (MW 38,000, M/G 2.24) showed the most potent TNF-α-inducing activity. Alginates having higher M/G ratio tended to show higher activity. These results suggest that molecular size and M/G ratio are important structural parameters influencing the TNF-α-inducing activity. Interestingly, enzymatic depolymerization of alginate with bacterial alginate lyase resulted in dramatic increase in the TNF-α-inducing activity. The higher activity of enzymatically digested alginate oligomers to induce nitric oxide production from RAW264.7 cells than alginate polymer was also observed. On the other hand, alginate polymer and oligomer showed nearly equal hydroxyl radical scavenging activities.

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This study evaluated the immunomodulatory activities, including regulation of nitric oxide (NO), reactive oxygen species (ROS), and tumor necrosis factor (TNF)-α production in RAW264.7 murine macrophages, of alginate oligosaccharides (AOS) and investigated their structure-activity relationships. Our results revealed that unsaturated guluronate oligosaccharide prepared by enzymatic degradation (GOS-ED) induced NO production and inducible nitric oxide synthase (iNOS) expression, dose and time dependently, and stimulated ROS and TNF-α production; however, other AOS prepared by different ways or polymers showed very low and even no such effects. Moreover, GOS-ED induced macrophage activation to release the above-mentioned mediators partly involved in nuclear factor (NF)-κB and mitogen-activated protein (MAP) kinase signaling pathways. We also show that the structural characteristics of AOS, especially the unsaturated terminal structure, molecular size, and M/G ratio, play important roles in determining the macrophage-activating effects. GOS-ED could be applicable for agriculture, drug, and food industry as a potent immune-modulatory agent.