| 1. |
- Allahgholi, Leila, et al.
(författare)
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Composition analysis and minimal treatments to solubilize polysaccharides from the brown seaweed Laminaria digitata for microbial growth of thermophiles
- 2020
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Ingår i: Journal of Applied Phycology. - : Springer Science and Business Media LLC. - 0921-8971 .- 1573-5176. ; 32:3, s. 1933-1947
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Tidskriftsartikel (refereegranskat)abstract
- Brown macroalgae (Phaeophyta) hold high potential as feedstock for biorefineries due to high biomass productivity and carbohydrate content. They are, however, a challenging, unconventional feedstock for microbial refining and several processing problems need to be solved to make them a viable option. Pre-treatment is necessary to enhance accessibility and solubility of the biomass components but should be minimal and mild to assure sustainable and cost-effective processing. Here, two routes to pre-treatLaminaria digitata to release polysaccharides were investigated: hot water pre-treatment by autoclaving (121 °C, 20 min or 60 min) and a two-step extraction with mild acid (0.1 M HCl) followed by alkaline treatment. Hot water pre-treatment resulted in partial extraction of a mixture of polysaccharides consisting of alginate, fucoidan and laminarin. After mild acid pre-treatment, alginate was found in the remaining insoluble residues and was extracted in a second step via alkaline treatment using Na2CO3 (0.15 M) at 80 °C and CaCl2 (10%) for the precipitation. In addition to carbohydrates, a fraction of other components such as proteins, phenolic compounds, minerals and trace elements was detected in the extracts. Cultivation of the thermophilic bacterial strains Rhodothermus marinus DSM 16675 and Bacillus methanolicus MGA3 (ATCC 53907) in media supplemented with the respective extracts resulted in growth of both strains, indicating that they were able to utilize the available carbon source for growth. R. marinus displayed the highest cell density in the medium containing the extract from acid pre-treatment, whereas B. methanolicus growth was highest with the extract from hot water pre-treatment.
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| 2. |
- Allahgholi, Leila, et al.
(författare)
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Exploring a novel β-1,3-glucanosyltransglycosylase, MlGH17B, from a marine Muricauda lutaonensis strain for modification of laminari-oligosaccharides
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Ingår i: Glycobiology. - 1460-2423.
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Tidskriftsartikel (refereegranskat)abstract
- The marine environment, contains plentiful renewable resources, e.g. macroalgae with unique polysaccharides, motivating search for enzymes from marine microorganisms to explore conversion possibilities of the polysaccharides. In this study, the first GH17 glucanosyltransglycosylase, MlGH17B, from a marine bacterium (Muricauda lutaonensis), was characterized. The enzyme was moderately thermostable with Tm at 64.4 °C and 73.2 °C, but an activity optimum at 20 °C, indicating temperature sensitive active site interactions. MlGH17B uses β-1,3 laminari-oligosaccharides with a degree of polymerization (DP) of 4 or higher as donors. Two glucose moieties (bound in the aglycone +1 and + 2 subsites) are cleaved off from the reducing end of the donor while the remaining part (bound in the glycone subsites) is transferred to an incoming β-1,3 glucan acceptor, making a β-1,6-linkage, thereby synthesizing branched or kinked oligosaccharides. Synthesized oligosaccharides up to DP26 were detected by mass spectrometry analysis, showing that repeated transfer reactions occurred, resulting in several β-1,6-linked branches. The modelled structure revealed an active site comprising five subsites: three glycone (-3, -2 and - 1) and two aglycone (+1 and + 2) subsites, with significant conservation of substrate interactions compared to the only crystallized 1,3-β-glucanosyltransferase from GH17 (RmBgt17A from the compost thriving fungus Rhizomucor miehei), suggesting a common catalytic mechanism, despite different phylogenetic origin, growth environment, and natural substrate. Both enzymes lacked the subdomain extending the aglycone subsites, found in GH17 endo-β-glucanases from plants, but this extension was also missing in bacterial endoglucanases (modelled here), showing that this feature does not distinguish transglycosylation from hydrolysis, but may rather relate to phylogeny.
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| 3. |
- Allahgholi, Leila, et al.
(författare)
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Fermentation of the Brown Seaweed Alaria esculenta by a Lactic Acid Bacteria Consortium Able to Utilize Mannitol and Laminari-Oligosaccharides
- 2023
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Ingår i: Fermentation. - 2311-5637. ; 9:6
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Tidskriftsartikel (refereegranskat)abstract
- The brown seaweed Alaria esculenta is the second most cultivated species in Europe, and it is therefore of interest to expand its application by developing food products. In this study, a lactic acid bacteria consortium (LAB consortium) consisting of three Lactiplantibacillus plantarum strains (relative abundance ~94%) and a minor amount of a Levilactobacillus brevis strain (relative abundance ~6%) was investigated for its ability to ferment carbohydrates available in brown seaweed. The consortium demonstrated the ability to ferment glucose, mannitol, galactose, mannose, and xylose, of which glucose and mannitol were the most favored substrates. No growth was observed on fucose, mannuronic and guluronic acid. The consortium used different pathways for carbohydrate utilization and produced lactic acid as the main metabolite. In glucose fermentation, only lactic acid was produced, but using mannitol as a carbohydrate source resulted in the co-production of lactic acid, ethanol, and succinate. Xylose fermentation resulted in acetate production. The consortium was also able to utilize laminari-oligosaccharides (DP2-4), obtained after enzymatic hydrolysis of laminarin, and produced lactic acid as a metabolite. The consortium could grow directly on A. esculenta, resulting in a pH decrease to 3.8 after 7 days of fermentation. Incubation of the same seaweed in corresponding conditions without inoculation resulted in spoilage of the seaweed by endogenous bacteria.
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| 4. |
- Allahgholi, Leila
(författare)
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Innovative processing of brown seaweed for value-added products
- 2023
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The increasing depletion of fossil fuels, rising greenhouse gas emissions, and a growing global population have increased the need for sustainable food and energy sources. Traditional industrial biotechnology, which relies on plant-based feedstocks, competes with food production for arable land. In contrast, marine macroalgae do not require cultivatable land, irrigation by fresh water or fertilizers, and are a promising alternative due to fast growth, productivity, and high carbohydrate content. This study focuses on the potential of brown macroalgae as a sustainable source for production of value-added chemicals, food products, and ingredients. In this research, various pre-treatment methods, including acid, heat, and enzymatic treatment, were performed to solubilize carbohydrates from brown seaweed for subsequent biorefinery utilization. The extracts from acid and heat treatment of Laminaria digitata showed its potential to be used as feed for cultivating thermophilic bacteria, such as Rhodothermus marinus and Bacillus methanolicus, respectively, which are able to produce bioactive compounds and can be used as microbial cell factories, demonstrating the suitability of the extracts for replacing terrestrial carbohydrates for bacterial cultivation. Biofuel production through fermentation using various microbes, revealed that Thermoanaerobacterium AK17 could utilize not only glucose but also mannitol and glucans released by enzymatic treatment of the brown seaweed L. digitata in ethanol production. Additionally, Clostridial strains studied in this thesis showed different nutrient requirements, but, however, they were able to utilize glucan and mannitol available in the L. digitata hydrolysates for the production of acetone/isopropanol, butanol, and ethanol (A/IBE). A lactic acid bacteria consortium was shown to be able to ferment mannitol from A. esculenta. This could be a way to increase the shelf life of the seaweed, an alternative to current storage techniques.Laminarin is a glucan composed of β-1,3(6) linked glucose unit and is a storage carbohydrate in brown seaweed. Enzymes capable of degrading and modifying laminarin and laminari-oligosaccharides were explored for potential valorization of this glucan. In this study, two novel GH17 enzymes were identified in the marine bacterium Muricauda lutaonensis; one is an endo-acting laminarinase, capable of degrading laminarin into short-chain oligosaccharides with a degree of polymerization (DP) 2-4, with potential prebiotic properties. The other is β-1,3-glucanosyltransglycosylase which utilizes β-1,3 linked oligosaccharides with DP above 5 as a donor to introduce β-1,6 linked branches or kinks into β-1,3-glucans, which have potential bioactive properties. These findings collectively offer invaluable insights into the potential of brown macroalgae as a renewable, sustainable source for biofuels and value-added biomolecules.
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| 5. |
- Daugbjerg Christensen, Monica, et al.
(författare)
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Cloning and Characterization of a Novel N-Acetyl-D-galactosamine-4-O-sulfate Sulfatase, SulA1, from a Marine Arthrobacter Strain
- 2024
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Ingår i: Marine Drugs. - 1660-3397. ; 22:3
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Tidskriftsartikel (refereegranskat)abstract
- Sulfation is gaining increased interest due to the role of sulfate in the bioactivity of many polysaccharides of marine origin. Hence, sulfatases, enzymes that control the degree of sulfation, are being more extensively researched. In this work, a novel sulfatase (SulA1) encoded by the gene sulA1 was characterized. The sulA1-gene is located upstream of a chondroitin lyase encoding gene in the genome of the marine Arthrobacter strain (MAT3885). The sulfatase was produced in Escherichia coli. Based on the primary sequence, the enzyme is classified under sulfatase family 1 and the two catalytic residues typical of the sulfatase 1 family—Cys57 (post-translationally modified to formyl glycine for function) and His190—were conserved. The enzyme showed increased activity, but not improved stability, in the presence of Ca2+, and conserved residues for Ca2+ binding were identified (Asp17, Asp18, Asp277, and Asn278) in a structural model of the enzyme. The temperature and pH activity profiles (screened using p-nitrocatechol sulfate) were narrow, with an activity optimum at 40–50 °C and a pH optimum at pH 5.5. The Tm was significantly higher (67 °C) than the activity optimum. Desulfation activity was not detected on polymeric substrates, but was found on GalNAc4S, which is a sulfated monomer in the repeated disaccharide unit (GlcA–GalNAc4S) of, e.g., chondroitin sulfate A. The position of the sulA1 gene upstream of a chondroitin lyase gene and combined with the activity on GalNAc4S suggests that there is an involvement of the enzyme in the chondroitin-degrading cascade reaction, which specifically removes sulfate from monomeric GalNAc4S from chondroitin sulfate degradation products.
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| 6. |
- Jönsson, Madeleine, et al.
(författare)
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Exploration of high-pressure processing (HPP) for preservation of the Swedish grown brown macroalgae Saccharina latissima
- 2023
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Ingår i: Frontiers in Food Science and Technology. - 2374-5533. ; 3
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Tidskriftsartikel (refereegranskat)abstract
- Introducing seaweed to new food markets entails new challenges concerning efficient preservation. Hence, this study explores high-pressure processing (HPP) as an alternative technique to conventional methods by evaluating its effects on the composition, quality, and microbial safety of the Swedish grown macroalgae Saccharina latissima. The results from the physicochemical analysis showed that after high-pressure treatment the color was retained, while the algal texture was altered by up to an 87.7% reduction in hardness and a 60.0% reduction in compression. Biochemical analysis demonstrated some variations in the algal samples, but the nutritional content was overall retained after treatment. The microbial analysis showed a low microbial load of untreated fresh material, which was confirmed by a lack of amplification in polymerase chain reaction attempts and low growth during attempts on spontaneous proliferation using fresh and frozen algae. Additionally, shelf-life studies showed inconsistent growth, but overall, a low increase in unspecific bacteria, an increasing load of Enterobacteriaceae, no growth of Lactobacilli, and low fouling by mold and yeast. The results from this study can be useful in the continued attempts of introducing seaweed to new markets, with different prerequisites for post-harvest treatment.
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| 7. |
- Jönsson, Madeleine, et al.
(författare)
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Extraction and Modification of Macroalgal Polysaccharides for Current and Next-Generation Applications
- 2020
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Ingår i: Molecules. - : MDPI AG. - 1420-3049. ; 25:930
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Tidskriftsartikel (refereegranskat)abstract
- Marine macroalgal (seaweed) polysaccharides are highly promising for next-generation applications in several industries. However, despite the reported comprehensive potential of these polysaccharides, commercial products are scarce on the market. Seaweed cultivations are increasing in number and production quantity, owing to an elevated global trend of utilization interest in seaweed. The extraction of polysaccharides from seaweed generally generates low yields, but novel methods are being developed to facilitate and improve the extraction processes. Current areas of applications for seaweed polysaccharides mainly take advantage of the physicochemical properties of certain polysaccharides, such as gelling, thickening and emulsifying. However, many of the numerous bioactivities reported are still only at research level and lack clinical evidence for commercialization. It has been suggested the construction of smaller units may generate better defined molecules that are more suitable for biomedical applications. Enzymatic modification is a promising tool for the generation of more defined, targeted biomolecules. This review covers; structural differences between the most predominant marine algal polysaccharides, extraction processes, modification alternatives, as well as a summary of current and potential next-generation application areas.
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| 8. |
- Moenaert, Antoine, et al.
(författare)
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Evaluation of Laminaria Digitata Hydrolysate for the Production of Bioethanol and Butanol by Fermentation
- 2023
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Ingår i: Fermentation. - : MDPI AG. - 2311-5637. ; 9:1
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Tidskriftsartikel (refereegranskat)abstract
- Seaweeds (macroalgae) are gaining attention as potential sustainable feedstock for the production of fuels and chemicals. This comparative study focuses on the characterization of the microbial production of alcohols from fermentable carbohydrates in the hydrolysate of the macroalgae Laminaria digitata as raw material. The potential of a hydrolysate as a carbon source for the production of selected alcohols was tested, using three physiologically different fermentative microbes, in two main types of processes. For the production of ethanol, Saccharomyces cerevisiae was used as a benchmark microorganism and compared with the strictly anaerobic thermophile Thermoanaerobacterium strain AK17. For mixed production of acetone/isopropanol, butanol, and ethanol (A/IBE), three strictly anaerobic Clostridium strains were compared. All strains grew well on the hydrolysate, and toxicity constraints were not observed, but fermentation performance and product profiles were shown to be both condition- and strain-specific. S. cerevisiae utilized only glucose for ethanol formation, while strain AK17 utilized glucose, mannitol, and parts of the glucan oligosaccharides. The clostridia strains tested showed different nutrient requirements, and were able to utilize glucan, mannitol, and organic acids in the hydrolysate. The novelty of this study embodies the application of different inoculates for fermenting a common brown seaweed found in the northern Atlantic Ocean. It provides important information on the fermentation properties of different microorganisms and pinpoints the value of carbon source utilization when selecting microbes for efficient bioconversion into biofuel and chemical products of interest.
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| 9. |
- Moenaert, Antoine, et al.
(författare)
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Metabolic engineering of Thermoanaerobacterium AK17 for increased ethanol production in seaweed hydrolysate
- 2023
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Ingår i: Biotechnology for Biofuels and Bioproducts. - 2731-3654. ; 16:1
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Tidskriftsartikel (refereegranskat)abstract
- Sustainably produced renewable biomass has the potential to replace fossil-based feedstocks, for generation of biobased fuels and chemicals of industrial interest, in biorefineries. In this context, seaweeds contain a large fraction of carbohydrates that are a promising source for enzymatic and/or microbial biorefinery conversions. The thermoanaerobe Thermoanaerobacterium AK17 is a versatile fermentative bacterium producing ethanol, acetate and lactate from various sugars. In this study, strain AK17 was engineered for more efficient production of ethanol by knocking out the lactate and acetate side-product pathways. This was successfully achieved, but the strain reverted to acetate production by recruiting enzymes from the butyrate pathway. Subsequently this pathway was knocked out and the resultant strain AK17_M6 could produce ethanol close to the maximum theoretical yield (90%), leading to a 1.5-fold increase in production compared to the wild-type strain. Strain AK17 was also shown to successfully ferment brown seaweed hydrolysate from Laminaria digitata to ethanol in a comparatively high yield of 0.45 g/g substrate, with the primary carbon sources for the fermentations being mannitol, laminarin-derived glucose and short laminari-oligosaccharides. As strain AK17 was successfully engineered and has a wide carbohydrate utilization range that includes mannitol from brown seaweed, as well as hexoses and pentoses found in both seaweeds and lignocellulose, the new strain AK17_M6 obtained in this study is an interesting candidate for production of ethanol from both second and third generations biomass.
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| 10. |
- Schiøtt, Morten, et al.
(författare)
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Biorefining of brown seaweeds catalyzed through innovative enzyme processes
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Ingår i: Industrial Biotechnology. - 1550-9087.
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Tidskriftsartikel (refereegranskat)abstract
- The current expansion of seaweed farming to North America and Europe can be a cornerstone in a new “blue bioeconomy” in the Northern Hemisphere. In this domain, the focus of R&D efforts is on creating value-added products through new biorefining processes for valorizing the unique polysaccharides of seaweeds. Apart from direct consumption of seaweeds as food—particularly in the Asian cuisine—commercial seaweed products are primarily natural hydrocolloids used to make viscous suspensions and gels, but new valuable products exerting bioactivity are coming into focus. This recent development rests on targeted, gentle extraction and modification of the seaweed polysaccharides using tailormade bioprocessing enzyme technologies. Since brown seaweed cultivation is rising in the Northern Hemisphere, this article provides an overview of recent advances and prospects in brown seaweed biorefining.
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