| 1. |
- Ekman, Anna, et al.
(författare)
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Bioresource utilisation by sustainable technologies in new value-added biorefinery concepts - two case studies from food and forest industry
- 2013
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Ingår i: Journal of Cleaner Production. - : Elsevier BV. - 0959-6526 .- 1879-1786. ; 57, s. 46-58
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Tidskriftsartikel (refereegranskat)abstract
- This paper presents a trans-disciplinary assessment of new and innovative biorefinery concepts producing high-value chemical compounds from residues from agriculture, food and forest industries. There is a significant potential of biomass residues in Sweden suitable for the extraction of various compounds, including upgrading by biocatalytic processes, in addition to current energy generation. Two examples presented are quercetin extracted from onion waste by pressurised hot water in conjunction with enzymatic hydrolysis, and betulin from birch bark extracted by liquid CO2 containing ethanol. Inherent in these two extraction processes and production routes is the ability to show good environmental performance from a life cycle perspective. Extraction of high-value compounds also provides possibilities for innovation in the current agricultural, food and forest industry potentially leading to socio-economical benefits. (C) 2013 The Authors. Published by Elsevier Ltd. All rights reserved.
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| 2. |
- Gondo, Thamani Freedom, et al.
(författare)
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Extractability, selectivity, and comprehensiveness in supercritical fluid extraction of seaweed using ternary mixtures of carbon dioxide, ethanol, and water
- 2023
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Ingår i: Journal of chromatography. A. - 0021-9673. ; 1706, s. 464267-464267
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Tidskriftsartikel (refereegranskat)abstract
- It is well-known that an ideal extraction method enabling quantitative analysis should give complete extraction of the target analytes as well as minimal co-extraction of unwanted matrix substances. If the extraction method is part of a nontarget screening protocol, the desired analytes can differ widely in terms of chemical properties. In chromatography, terminologies such as recovery, selectivity, and comprehensiveness are well-established and can easily be determined. However, in extraction, these concepts are much less developed. Hence, the aim of our research is to develop and scrutinize theory in extraction with respect to numerical descriptors for extractability, selectivity, and comprehensiveness. Our approach is based on experiments determining the extractability of target analytes and selected interferences. As a case study, we use a pooled sample of three species of seaweed (Alaria esculenta, Laminaria digitata and Ascophyllum nodosum). Target analytes are β-carotene, fucoxanthin, δ-tocopherol, and phloroglucinol; and selected interferences are carbohydrates, proteins, ash, arsenic, and chlorophyll a. As a "green and clean" extraction technique, supercritical fluid extraction (SFE) using mixtures of CO 2, ethanol and water were explored using a design of experiment. The temperature was varied between 40-80°C, and the pressure was held constant at 300 bar. Obtained results clearly demonstrate that highest relative selectivity was achieved with CO 2 containing only 5 vol% of ethanol and no water, which primarily enabled high extractability of β-carotene, and yielding an extract free of carbohydrates, proteins, and toxic metals such as arsenic. Best methods for highest extractability of the other target analytes varied quite widely. Analytes requiring the highest water content (fucoxanthin and phloroglucinol), also resulted in the lowest relative selectivity. Maximum relative comprehensiveness was achieved using CO 2/ethanol/water (40/55/5, v/v/v) at 70°C and 300 bar. Our study demonstrates the feasibility of using relative quantitative descriptors for extractability, selectivity, and comprehensiveness, in optimization strategies for analytical extractions.
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| 3. |
- Karlsson, Eva Nordberg, et al.
(författare)
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Sustainable resource technology (SURETECH)
- 2012
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Ingår i: ; , s. 323-324
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Konferensbidrag (refereegranskat)abstract
- We all need to contribute to a more sustainable development – a fact that has become quite clear observing extreme weathers and diminishing fossil fuel sources. In line with the 12 Principles of Green Chemistry [1], (recommending the use of safer solvents, catalysed reactions, energy efficient processes and renewable feedstocks), research within the SuReTech program aims at developing technologies for the recovery of high-value compounds from forestry and agricultural byproducts. The idea is to create value addition, by adding products to already utilized resources in line with biorefinery concepts. Extractions of antioxidizing compounds from byproducts are thus made by use of carbon dioxide and water as sustainable solvents and combined with biocatalytic conversions for the creation of compounds with desired properties. The byproducts, used as starting materials, are selected based on annual volumes from the Swedish agricultural, food and forestry industry. Substituents on the compounds are modified by biocatalysis, e.g. using thermostable carbohydrate converting enzymes selected to fit the conditions in the extraction step. The importance of combining different scientific fields to enable creativity in sustainable development is highlighted.
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| 4. |
- Khan, Samiullah, et al.
(författare)
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Aglycone specificity of Thermotoga neapolitana beta-glucosidase 1A modified by mutagenesis, leading to increased catalytic efficiency in quercetin-3-glucoside hydrolysis
- 2011
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Ingår i: BMC Biochemistry. - : Springer Science and Business Media LLC. - 1471-2091. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- Background: The thermostable beta-glucosidase (TnBgl1A) from Thermotoga neapolitana is a promising biocatalyst for hydrolysis of glucosylated flavonoids and can be coupled to extraction methods using pressurized hot water. Hydrolysis has however been shown to be dependent on the position of the glucosylation on the flavonoid, and e. g. quercetin-3-glucoside (Q3) was hydrolysed slowly. A set of mutants of TnBgl1A were thus created to analyse the influence on the kinetic parameters using the model substrate para-nitrophenyl-beta-D-glucopyranoside (pNPGlc), and screened for hydrolysis of Q3. Results: Structural analysis pinpointed an area in the active site pocket with non-conserved residues between specificity groups in glycoside hydrolase family 1 (GH1). Three residues in this area located on beta-strand 5 (F219, N221, and G222) close to sugar binding sub-site +2 were selected for mutagenesis and amplified in a protocol that introduced a few spontaneous mutations. Eight mutants (four triple: F219L/P165L/M278I, N221S/P165L/M278I, G222Q/P165L/M278I, G222Q/V203M/K214R, two double: F219L/K214R, N221S/P342L and two single: G222M and N221S) were produced in E. coli, and purified to apparent homogeneity. Thermostability, measured as T-m by differential scanning calorimetry (101.9 degrees C for wt), was kept in the mutated variants and significant decrease (Delta T of 5 -10 degrees C) was only observed for the triple mutants. The exchanged residue(s) in the respective mutant resulted in variations in K-M and turnover. The K-M-value was only changed in variants mutated at position 221 (N221S) and was in all cases monitored as a 2-3 x increase for pNPGlc, while the K-M decreased a corresponding extent for Q3. Turnover was only significantly changed using pNPGlc, and was decreased 2-3 x in variants mutated at position 222, while the single, double and triple mutated variants carrying a mutation at position 221 (N221S) increased turnover up to 3.5 x compared to the wild type. Modelling showed that the mutation at position 221, may alter the position of N291 resulting in increased hydrogen bonding of Q3 (at a position corresponding to the +1 subsite) which may explain the decrease in K-M for this substrate. Conclusion: These results show that residues at the +2 subsite are interesting targets for mutagenesis and mutations at these positions can directly or indirectly affect both K-M and turnover. An affinity change, leading to a decreased K-M, can be explained by an altered position of N291, while the changes in turnover are more difficult to explain and may be the result of smaller conformational changes in the active site.
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| 5. |
- Khan, Sami, et al.
(författare)
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Immobilization of thermostable beta-glucosidase variants on acrylic supports for biocatalytic processes in hot water
- 2012
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Ingår i: Journal of Molecular Catalysis B: Enzymatic. - : Elsevier BV. - 1873-3158 .- 1381-1177. ; 80, s. 28-38
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Tidskriftsartikel (refereegranskat)abstract
- Two variants of the thermostable beta-glucosidase TnBgl1A (wt and N221S/P342L) from Thermotoga neapolitana were immobilized on acrylic supports (Eupergit (R) C, Eupergit (R) C250L, and cryogel) and evaluated at conditions close to the boiling point of water. Thermo-gravimetric analysis showed the supports to be stable <250 degrees C. Both wt and N221S/P342L were covalently bound to oxirane-groups respectively via glutaraldehyde spacers, and for coupling reactions 26 Lys and 20 Ser/Thr were surface-located. Immobilized enzymes were active on all supports in the temperature range 80-95 degrees C, but the observed specific activity was low (<= 19 U mg(-1)) using the cryogel. More than 91% of the initial activity was maintained after ten times recycling, and the same was recovered after 3 months storage at 4 degrees C for Eupergit (R) supports by simply incubating the preparation with bovine serum albumin. No storage loss was detectable on cryogels. The glutaraldehyde spacer improved activity on cryogels, but not on Eupergie supports. Immobilization on Eupergit (R) C250L yielded the highest observed specific activity (254 U mg(-1) for N221S/P342L) in a procedure including blocking of free oxirane-groups by BSA. This biocatalyst was used for on-line hydrolysis of quercetin-glucosides in a yellow onion extract at 80 degrees C, proving the immobilized biocatalyst to be promising in on-line systems for extraction and hydrolysis using hot pressurized water. (C) 2012 Elsevier B.V. All rights reserved.
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| 6. |
- Kristjansdottir, Thordis, et al.
(författare)
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Engineering the carotenoid biosynthetic pathway in Rhodothermus marinus for lycopene production
- 2020
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Ingår i: Metabolic Engineering Communications. - : Elsevier BV. - 2214-0301. ; 11
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Tidskriftsartikel (refereegranskat)abstract
- Rhodothermus marinus has the potential to be well suited for biorefineries, as an aerobic thermophile that produces thermostable enzymes and is able to utilize polysaccharides from different 2nd and 3rd generation biomass. The bacterium produces valuable chemicals such as carotenoids. However, the native carotenoids are not established for industrial production and R. marinus needs to be genetically modified to produce higher value carotenoids. Here we genetically modified the carotenoid biosynthetic gene cluster resulting in three different mutants, most importantly the lycopene producing mutant TK-3 (ΔtrpBΔpurAΔcruFcrtB::trpBcrtBT.thermophilus). The genetic modifications and subsequent structural analysis of carotenoids helped clarify the carotenoid biosynthetic pathway in R. marinus. The nucleotide sequences encoding the enzymes phytoene synthase (CrtB) and the previously unidentified 1′,2′-hydratase (CruF) were found fused together and encoded by a single gene in R. marinus. Deleting only the cruF part of the gene did not result in an active CrtB enzyme. However, by deleting the entire gene and inserting the crtB gene from Thermus thermophilus, a mutant strain was obtained, producing lycopene as the sole carotenoid. The lycopene produced by TK-3 was quantified as 0.49 g/kg CDW (cell dry weight).
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| 7. |
- Lindahl, Sofia, et al.
(författare)
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An on-line method for pressurized hot water extraction and enzymatic hydrolysis of quercetin glucosides from onions.
- 2013
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Ingår i: Analytica Chimica Acta. - : Elsevier BV. - 1873-4324 .- 0003-2670. ; 785, s. 50-59
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Tidskriftsartikel (refereegranskat)abstract
- A novel environmentally sound continuous-flow hot water extraction and enzymatic hydrolysis method for determination of quercetin in onion raw materials was successfully constructed using a stepwise optimization approach. In the first step, enzymatic hydrolysis of quercetin-3,4'-diglucoside to quercetin was optimized using a three level central composite design considering temperature (75-95°C), pH (3-6) and volume concentration of ethanol (5-15%). The enzyme used was a thermostable β-glucosidase variant (termed TnBgl1A_N221S/P342L) covalently immobilized on either of two acrylic support-materials (Eupergit(®) C 250L or monolithic cryogel). Optimal reaction conditions were irrespective of support 84°C, 5% ethanol and pH 5.5, and at these conditions, no significant loss of enzyme activity was observed during 72h of use. In a second step, hot water extractions from chopped yellow onions, run at the optimal temperature for hydrolysis, were optimized in a two level design with respect to pH (2.6 and 5.5), ethanol concentration (0 and 5%) and flow rate (1 and 3mLmin(-1)) Obtained results showed that the total quercetin extraction yield was 1.7 times higher using a flow rate of 3mLmin(-1) (extraction time 90min), compared to a flow rate of 1mLmin(-1) (extraction time 240min). Presence of 5% ethanol was favorable for the extraction yield, while a further decrease in pH was not, not even for the extraction step alone. Finally, the complete continuous flow method (84°C, 5% ethanol, pH 5.5, 3mLmin(-1)) was used to extract quercetin from yellow, red and shallot onions and resulted in higher or similar yield (e.g. 8.4±0.7μmolg(-1) fresh weight yellow onion) compared to a conventional batch extraction method using methanol as extraction solvent.
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| 8. |
- Lindahl, Sofia, et al.
(författare)
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Exploring the possibility of using a thermostable mutant of β-glucosidase for rapid hydrolysis of quercetin glucosides in hot water
- 2010
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Ingår i: Green Chemistry. - : The Royal Society of Chemistry. - 1463-9262 .- 1463-9270. ; 12:1, s. 159-168
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Tidskriftsartikel (refereegranskat)abstract
- The antioxidant quercetin was extracted from yellow onion waste and converted to its aglycone form by a combination of subcritical water extraction and enzymatic hydrolysis. The hydrolytic step was catalysed by a double residue (N221S, P342L) mutant of the thermostable beta-glucosidase (TnBgl1A), isolated from the thermophile Thermotoga neapolitana and cloned and produced in E. coli. The activity of wt TnBgl1A was shown to be dependent on the position of the glucosylation on the quercetin backbone, favouring hydrolysis of quercetin-4'-glucoside over quercetin-3-glucoside. The mutated variant of the enzyme harboured a mutation in the +2 sub-site (N221S) and showed increased catalytic efficiency in quercetin-3-glucoside hydrolysis and also to a certain extent hydrolysis of quercetin-4'-glucoside. The mutated enzyme was used directly in yellow onion extracts, prepared by subcritical water extraction, resulting in complete hydrolysis of the glucosylated flavonoids quercetin-3,4'-diglucoside, quercetin-4'-glucoside, quercetin-3-glucoside, isorhamnetin-4'-glucoside and isorhamnetin-3,4'-diglucoside. To complete hydrolysis within five minutes, 3 mg of TnBgl1A_N221S was used per gramme of onion (dry weight). A life cycle assessment was done to compare the environmental impact of the new method with a conventional solid-liquid extraction-and-hydrolysis method utilising aqueous methanol and hydrochloric acid. Comparison of the methods showed that the new method is preferable regarding primary energy consumption and global warming potential. Another advantage of this method is that handling of toxic chemicals (methanol and HCl) is avoided. This shows that combined subcritical water extraction/enzyme hydrolysis is both a fast and sustainable method to obtain quercetin from onion waste.
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| 9. |
- Plaza, Merichel, et al.
(författare)
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Substituent Effects on in Vitro Antioxidizing Properties, Stability, and Solubility in Flavonoids
- 2014
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Ingår i: Journal of Agricultural and Food Chemistry. - : American Chemical Society (ACS). - 0021-8561 .- 1520-5118. ; 62:15, s. 3321-3333
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Forskningsöversikt (refereegranskat)abstract
- Antioxidants are widely used by humans, both as dietary supplements and as additives to different types of products. The desired properties of an antioxidant often include a balance between the antioxidizing capacity, stability, and solubility. This review focuses on flavonoids, which are naturally occurring antioxidants, and different common substituent groups on flavonoids and how these affect the properties of the molecules in vitro. Hydroxyl groups on flavonoids are both important for the antioxidizing capacity and key points for further modification resulting in O-methylation, -glycosylation, -sulfation, or -acylation. The effects of O-glycosylation and acylation are discussed as these types of substitutions have been most explored in vitro concerning antioxidizing properties as well as stability and solubility. Possibilities to control the properties by enzymatic acylation and glycosylation are also reviewed, showing that depending on the choice of enzyme and substrate, regioselective results can be obtained, introducing possibilities for more targeted production of antioxidants with predesigned properties.
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| 10. |
- Ron, Emanuel, et al.
(författare)
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Characterization of carotenoids in Rhodothermus marinus
- 2018
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Ingår i: MicrobiologyOpen. - : Wiley. - 2045-8827. ; 7:1
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Tidskriftsartikel (refereegranskat)abstract
- Rhodothermus marinus, a marine aerobic thermophile, was first isolated from an intertidal hot spring in Iceland. In recent years, the R. marinus strain PRI 493 has been genetically modified, which opens up possibilities for targeted metabolic engineering of the species, such as of the carotenoid biosynthetic pathway. In this study, the carotenoids of the R. marinus type-strain DSM 4252T, strain DSM 4253, and strain PRI 493 were characterized. Bioreactor cultivations were used for pressurized liquid extraction and analyzed by ultra-high performance supercritical fluid chromatography with diode array and quadropole time-of-flight mass spectrometry detection (UHPSFC-DAD-QTOF/MS). Salinixanthin, a carotenoid originally found in Salinibacter ruber and previously detected in strain DSM 4253, was identified in all three R. marinus strains, both in the hydroxylated and nonhydroxylated form. Furthermore, an additional and structurally distinct carotenoid was detected in the three strains. MS/MS fragmentation implied that the mass difference between salinixanthin and the novel carotenoid structure corresponded to the absence of a 4-keto group on the ß-ionone ring. The study confirmed the lack of carotenoids for the strain SB-71 (ΔtrpBΔpurAcrtBI’::trpB) in which genes encoding two enzymes of the proposed pathway are partially deleted. Moreover, antioxidant capacity was detected in extracts of all the examined R. marinus strains and found to be 2–4 times lower for the knock-out strain SB-71. A gene cluster with 11 genes in two operons in the R. marinusDSM 4252T genome was identified and analyzed, in which several genes were matched with carotenoid biosynthetic pathway genes in other organisms.
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