| 1. |
- Rupar, Katarina, et al.
(författare)
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Solid Phase Micro Extraction Fibers, Calibration for Use in Biofilter Applications
- 2006
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Ingår i: Biochemical Engineering Journal. - : Elsevier BV. - 1369-703X. ; 31:2, s. 107-112
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Tidskriftsartikel (refereegranskat)abstract
- The main purpose of this study is to develop a SPME calibration method suitable for use in evaluation of concentrations of hydrophobic substances in environmental samples. The analyte used in the experiments was alpha-pinene, a hydrophobic organic compound commonly found in wood, and therefore found in wood storage facilities, wood processing industries and wood based biofilters. The SPME fibres were calibrated for different concentrations of alpha-pinene at different temperatures and relative humidities. The method was used to evaluate the removal efficiency of a lab-scale biofilter.
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| 2. |
- de Maré, Lena, et al.
(författare)
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A cultivation technique for E. coli fed-batch cultivations operating close to the maximum oxygen transfer capacity of the reactor
- 2005
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Ingår i: Biotechnology Letters. - : Springer Science and Business Media LLC. - 0141-5492 .- 1573-6776. ; 27:14, s. 983-990
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Tidskriftsartikel (refereegranskat)abstract
- A cultivation strategy combining the advantages of temperature-limited fed-batch and probing feeding control is presented. The technique was evaluated in fed-batch cultivations with E. coli BL21(DE3) producing xylanase in a 3 liter bioreactor. A 20% increase in cell mass was achieved and the usual decrease in specific enzyme activity normally observed during the late production phase was diminished with the new technique. The method was further tested by growing E. coli W3110 in a larger bioreactor (50 l). It is a suitable cultivation technique when the O2 transfer capacity of the reactor is reached and it is desired to continue to produce the recombinant protein.
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| 3. |
- Ahlqvist, Josefin, et al.
(författare)
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Monitoring the production of inclusion bodies during fermentation and enzyme-linked immunosorbent assay analysis of intact inclusion bodies using cryogel minicolumn plates
- 2006
-
Ingår i: Analytical Biochemistry. - : Elsevier BV. - 1096-0309 .- 0003-2697. ; 354:2, s. 229-237
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Tidskriftsartikel (refereegranskat)abstract
- A novel minicolumn chromatgraphic method to monitor the production of inclusion bodies during fermentation and anenzyme-linked immunosorbent assay (ELISA) system allowing direct analysis of the particles with surface-displayed antigens are described. A 33-kDa protein containing 306 amino acids with three sulfur bridges produced its inclusion bodies wits labeled with polyclonal antibodies against 15 amino acid (anti-A15) and 17 amino acid (anti-B17) residues at the N- and C-terminal ends of the protein, respectively. Labeled particles were bound to macroporous Monolithic protein A-cryogel adsorbents inserted into the open-ended wells of a 96-well plate (referred to as protein A-cryogel minicolumn plate). The concept behind this application is that the binding degree of inclusion bodies from lysed fermentation broth to the cryogel minicolumns increases with an increase in their concentration during fermentation. The technique allowed LIS to monitor the increase in the production levels of the inclusion bodies as the fermentation process progressed. The system also has a built-in quality parameter to ensure that the target protein has been fully expressed. Alternatively, inclusion bodies immobilized on phenyl-cryogel minicolumn plate were used in indirect ELISA based on anti-A15 and anti-B17 antibodies against terminal amino acid residues displayed oil the surface of inclusion bodies. Drainage-protected properties of the cryogel minicolumns allow performance of successive reactions with tested immunoglobulin G (IgG) samples and enzyme-conjugated secondary I-G and of enzymatic reaction within the adsorbent. (c) 2006 Elsevier Inc. All rights reserved.
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| 4. |
- Lundgren, Joakim, et al.
(författare)
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Integration of a hemicellulose extraction process into a biomass based heat and power plant
- 2009
-
Ingår i: Proceedings of ECOS 2009. - Foz do Iguaçú : ABCM, Brazilian Society of Mechanical Sciences and Engineering.
-
Konferensbidrag (refereegranskat)abstract
- The development of processes where lignocellulosic biomass can be refined to several different end-products in the same plant, i.e. a biorefinery, will be important in the development towards a more sustainable society where fossil fuels are replaced. This paper presents the idea to integrate the production of green chemicals via hot water hemicellulose extraction of birch wood (hardwood) into a small-scale combined heat and power plant (CHP), in this case an externally fired gas turbine. A techno-economically successful concept could provide the option to turn a small- to medium scale CHP plant into a small- to medium scale biorefinery. The results show that the extracted wood-chips would serve very well as a fuel for combustion and gasification processes due to the relatively high heating value, low ash content and significantly lower concentrations of alkali metals. Under the assumed economic conditions, electricity can be produced to a cost in the range of €85.6 to €196.2 per MWhel and a fermentable feedstock stream with a xylose concentration of 65 g/L to a cost in between €0.44 to €4.15 per kg xylose depending on plant size and number of annual operational hours.
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| 5. |
- Enman, Josefine, et al.
(författare)
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Raman analysis of synthetic eritadenine
- 2008
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Ingår i: Journal of Raman Spectroscopy. - : Wiley. - 0377-0486 .- 1097-4555. ; 39:10, s. 1464-1468
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Tidskriftsartikel (refereegranskat)abstract
- Eritadenine, 2(R),3(R)-dihydroxy-4-(9-adenyl)-butyric acid, is a cholesterol-reducing compound naturally occurring in the shitake mushroom (Lentinus edodes). To identify the unknown Raman spectrum of this compound, pure synthetic eritadenine was examined and the vibrational modes were assigned by following the synthesis pathway. This was accomplished by comparing the known spectra of the starting compounds adenine and D-ribose with the spectra of a synthesis intermediate, methyl 5-(6-Aminopurin-9H-9-yl)-2,3-O-isopropylidene-5-deoxy-β-D-ribofuranoside (MAIR) and eritadenine. In the Raman spectrum of eritadenine, a distinctive vibrational mode at 773 cm-1 was detected and ascribed to vibrations in the carbon chain, ν(C--C). A Raman line that arose at 1212 cm-1, both in the Raman spectrum of MAIR and eritadenine, was also assigned to ν(C--C). Additional Raman lines detected at 1526 and at 1583 cm-1 in the Raman spectrum of MAIR and eritadenine were assigned to ν(N--C) and a deformation of the purine ring structure. In these cases the vibrational modes are due to the linkage between adenine and the ribofuranoside moiety for MAIR, and between adenine and the carbon chain for eritadenine. This link is also the cause for the disappearance of adenine specific Raman lines in the spectrum of both MAIR and eritadenine. Several vibrations observed in the spectrum of D-ribose were not observed in the Raman spectrum of eritadenine due to the absence of the ribose ring structure. In the Raman spectrum of MAIR some of the D-ribose specific Raman lines disappeared due to the introduction of methyl and isopropylidene moieties to the ribose unit. With the approach presented in this study the so far unknown Raman spectrum of eritadenine could be successfully identified and is presented here for the first time.
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| 6. |
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| 7. |
- Malinovskiy, Dmitry, et al.
(författare)
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Determination of the isotopic composition of molybdenum in the bottom sediments of freshwater basins
- 2007
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Ingår i: Geochemistry International. - 0016-7029 .- 1556-1968. ; 45:4, s. 381-389
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Tidskriftsartikel (refereegranskat)abstract
- This paper presents the results of measurements of the Mo isotopic composition in the bottom sediments (BS) of freshwater basins. Mo isotopic ratios were measured using a multicollector inductively coupled plasma mass spectrometer (MC ICP MS). Efficient methods were used in this study for Mo separation from the elements of the sample matrix and correction for instrumental mass discrimination. This allowed us to achieve a high accuracy of 0.06, 0.08, and 0.14‰ (2 σ) for the measurement of 97Mo/95Mo, 98Mo/95Mo, and 100Mo/95Mo, respectively. The range of variations in Mo isotope ratios observed in the collected BS columns was ∼2.2‰ in terms of δ97Mo/95Mo. The results obtained here suggest that geochemical processes occurring during Mo migration with land water can change the isotopic composition of Mo. It is pointed out that the potential use of Mo isotopic systematics for reconstructions of redox conditions in seawater over the geologic past requires the quantification of isotopic effects of Mo accompanying its migration on land and the extent of possible variations in the isotopic composition of Mo entering the ocean.
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| 8. |
- Andersson, Christian
(författare)
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Biobased production of succinic acid by Escherichia coli fermentation
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The prospects of peak oil, climate change and the dependency of fossil carbon have urged research and development of production methods for the manufacture of fuels and chemicals from renewable resources (biomass). The present thesis illustrates different aspects of biobased succinic acid production by a metabolically engineered E. coli strain. The main areas of the thesis are sugar utilisation and feedstock flexibility, and fermentation inhibition, both due to toxic compound derived from the raw material and the fermentation products themselves.The first part of this thesis aimed to investigate the fermentation characteristics of AFP184 in a medium consisting of corn steep liquor, inorganic salts and different sugar sources without supplementation with high-cost nutrients such as yeast extract and peptone. The effects of different sugars, sucrose, glucose, fructose, xylose, equal mixtures of glucose-fructose and glucose-xylose, on succinic acid production kinetics and yields in an industrially relevant medium were investigated. AFP184 was able to utilise all sugars and sugar combinations except sucrose for biomass generation and succinate production. Using glucose resulted in the highest yield, 0.83 (g succinic acid per g sugar consumed anaerobically). Using a high initial sugar concentration resulted in volumetric productivities of almost 3 g L-1 h-1, which is above estimated values for economically feasible production. However, succinic acid production ceased at final concentrations greater than 40 g L-1. To further increase succinic acid concentrations, fermentations using NH4OH, NaOH, KOH, K2CO3, and Na2CO3 as neutralising agents were performed and compared. It was shown that substantial improvements could be made by using alkali bases to neutralise the fermentations. The highest concentrations and productivities were achieved when Na2CO3 was used, 77 g L-1 and 3 g L-1 h-1 respectively. A gradual decrease in succinate productivity was observed during the fermentations, which was shown to be due to succinate accumulation in the broth and not as a result of the addition of neutralising agent or the subsequent increase in osmolarity.To maintain high succinate productivity by keeping a low extracellular succinic acid concentration fermentations were interrupted and cells recovered and resuspended in fresh media. By removing the succinate it was possible to maintain high succinic acid productivity for a prolonged time. Cells subjected to high concentrations of succinate were also able to regain high productivity once transferred into a succinate-free medium.In the last part of the thesis succinic acid production from softwood dilute acid hydrolysates was demonstrated. This study involved establishing the degree of detoxification necessary for growth and fermentation using industrial hydrolysates. Detoxification by treatment with lime and/or activated carbon was investigated and the results show that it was possible to produce succinate from softwood hydrolysates in yields comparable to those for synthetic sugars.The work done in this thesis increases the understanding of succinic acid production with AFP184, illustrate its limitations, and suggests improvements in the current technology with the long term aim of increasing the economical feasibility of biochemical succinic acid production.
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| 9. |
- Andersson, Christian, et al.
(författare)
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Effect of different carbon sources on the production of succinic acid using metabolically engineered Escherichia coli
- 2007
-
Ingår i: Biotechnology progress (Print). - : Wiley. - 8756-7938 .- 1520-6033. ; 23:2, s. 381-388
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Tidskriftsartikel (refereegranskat)abstract
- Succinic acid (SA) is an important platform molecule in the synthesis of a number of commodity and specialty chemicals. In the present work, dual-phase batch fermentations with the E. coli strain AFP184 were performed using a medium suited for large-scale industrial production of SA. The ability of the strain to ferment different sugars was investigated. The sugars studied were sucrose, glucose, fructose, xylose, and equal mixtures of glucose and fructose and glucose and xylose at a total initial sugar concentration of 100 g L-1. AFP184 was able to utilize all sugars and sugar combinations except sucrose for biomass generation and succinate production. For sucrose as a substrate no succinic acid was produced and none of the sucrose was metabolized. The succinic acid yield from glucose (0.83 g succinic acid per gram glucose consumed anaerobically) was higher than the yield from fructose (0.66 g g-1). When using xylose as a carbon source, a yield of 0.50 g g-1 was obtained. In the mixed-sugar fermentations no catabolite repression was detected. Mixtures of glucose and xylose resulted in higher yields (0.60 g g-1) than use of xylose alone. Fermenting glucose mixed with fructose gave a lower yield (0.58 g g-1) than fructose used as the sole carbon source. The reason is an increased pyruvate production. The pyruvate concentration decreased later in the fermentation. Final succinic acid concentrations were in the range of 25-40 g L-1. Acetic and pyruvic acid were the only other products detected and accumulated to concentrations of 2.7-6.7 and 0-2.7 g L-1. Production of succinic acid decreased when organic acid concentrations reached approximately 30 g L-1. This study demonstrates that E. coli strain AFP184 is able to produce succinic acid in a low cost medium from a variety of sugars with only small amounts of byproducts formed.
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| 10. |
- Andersson, Christian, et al.
(författare)
-
Effects of neutralising agent, organic acids, and osmolarity on succinic acid production by Escherichia coli AFP184
- 2008
-
Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Using a low-cost medium Escherichia coli AFP184 has previously been reported to produce succinic acid with volumetric productivities close to 3 g L-1 h-1. At a total organic acid concentration of 30 g L-1 the productivity decreased drastically resulting in final succinate concentrations of 40 g L-1. The economical viability of biochemical succinic acid production would benefit from higher final succinic acid concentrations and volumetric productivities maintained at >2.5 g L-1 h-1 for an extended period of time. In the present work the effects of osmolarity and neutralising agent (NH4OH, KOH, NaOH, K2CO3, and Na2CO3) on succinic acid production by AFP184 were investigated. Highest concentration of succinic acid was obtained with Na2CO3, 75 g L-1. It was also found that the osmolarity resulting from succinate production and subsequent base addition, only marginally affected the productivity per viable cell. Organic acid inhibition due to the produced succinic acid on the other hand significantly reduced succinic acid productivity per viable cell. When using NH4OH productivity completely ceased at approximately 40 g L-1. Volumetric productivities remained at 2.5 g L-1 h-1 for 5 to 10 hours longer when using K- or Na-bases than when using NH4OH. However, loss of cell viability occurred, and together with the acid inhibition decreased the volumetric productivities. In this study it was demonstrated that by altering the neutralising agent it was possible to increase the period of high volumetric productivity in the anaerobic phase and improve the final succinic acid concentration by almost 100 %
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| 11. |
- Andersson, Christian, et al.
(författare)
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Inhibition of succinic acid production in metabolically engineered Escherichia Coli by neutralizing agent, organic acids, and osmolarity
- 2009
-
Ingår i: Biotechnology progress (Print). - : Wiley. - 8756-7938 .- 1520-6033. ; 25:1, s. 116-123
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Tidskriftsartikel (refereegranskat)abstract
- The economical viability of biochemical succinic acid production is a result of many processing parameters including final succinic acid concentration, recovery of succinate, and the volumetric productivity. Maintaining volumetric productivities >2.5 g L-1 h(-1) is important if production of succinic acid from. renewable resources should be competitive. In this work, the effects of organic acids, osmolarity, and neutralizing agent (NH4OH, KOH, NaOH, K2CO3, and Na2CO3) on the fermentative succinic acid production by Escherichia coli AFP184 were investigated. The highest concentration of succinic acid, 77 g L-1. was obtained with Na2O3. In general, irrespective of the base used, succinic acid productivity per viable cell was significantly reduced as the concentration of the produced acid increased. Increased osmolarity resulting from base addition during succinate production only marginally affected the productivity per viable cell. Addition of the osmoprotectant glycine betaine to cultures resulted in an increased aerobic growth rate and anaerobic glucose consumption rate, but decreased succinic acid yield. When using NH4OH productivity completely ceased at a succinic acid concentration of similar to 40 g L-1. Volumetric productivities remained at 2.5 g L-1 h(-1) for tip to 10 h longer when K- or Na-bases where used instead of NH4OH. The decrease in cellular succinic acid productivity observed during the anaerobic phase was found to be due to increased organic acid concentrations rather than medium osmolarity.
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| 12. |
- Andersson, Christian
(författare)
-
Succinic acid production using metabolically engineered Escherichia coli
- 2007
-
Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The prospects of peak oil, climate change and the dependency of fossil carbon have urged research and development of production methods for the manufacture of fuels and chemicals from renewable resources (biomass). To date, the primary emphasis has been placed on the replacement of oil for transportation fuels. A highly significant subset of petroleum usage is the production of chemicals, which represents 10-15% of the petroleum usage. White biotechnology, also called industrial biotechnology, is a fast evolving technology with a large potential to have a substantial impact on the industrial production of fuels and chemicals from biomass. This work addresses the issue of chemical production by investigating the production of bio-based succinic acid, which can be used in a wide range of applications to replace petroleum based chemicals. Succinic acid can be produced by fermentation of sugar by a number of organisms; one is Escherichia coli (E. coli). It is known that E. coli under anaerobic conditions produces a mixture of organic acids. In order to obtain a cost-effective production it is necessary to metabolically engineer the organism to produce succinic acid in greater yield than the other acids. In the current work, E. coli mutant AFP184 was used. AFP184 originates from a near wild type strain, the C600 (ATCC 23724), which can ferment both five and six carbon sugars and has mutations in the glucose specific phosphotransferase system (ptsG), the pyruvate formate lyase system (pfl) and in the fermentative lactate dehydrogenase system (ldh). The previous studies using different organisms have all used cultivation mediums supplemented to some degree with different nutrients like biotin, thiamine and yeast extract. In order to apply the technology to large scale, production must be cost-effective and it is important to minimise the use of additional supplements. The first part of this work aimed to investigate the fermentation characteristics of AFP184 in a medium consisting of corn steep liquor, inorganic salts and different sugar sources without supplementation of other additional nutrients. It addresses questions regarding the effect of different sugars on succinic acid kinetics and yields in an industrially relevant medium. In order to gain a sustainable production of succinic acid from biomass feedstocks (sugar from biomass) it is important to investigate how well the organism can utilise different sugars in the biomass. The sugars studied were sucrose, glucose, fructose, xylose and equal mixtures of glucose-fructose and glucose-xylose at a total initial sugar concentration of 100 g L-1. AFP184 was able to utilise all sugars and sugar combinations except sucrose for biomass generation and succinate production. Using glucose resulted in the highest yield, 0.83 (g succinic acid per g sugar consumed anaerobically). Fructose resulted in a yield of 0.66 and xylose of 0.5. Using a high initial sugar concentration made it possible to obtain volumetric productivities of almost 3 g L-1h-1, which is above estimated values for feasible economic production. Succinic acid production ceased at final concentrations greater than 40 g L-1. In order to further increase succinic acid concentrations, this inhibitory effect was studied in the second part of the present work. The inhibitory effects can be two-fold including pH-based inhibition and an anion specific effect on metabolism. It has been reported that high concentrations of ammonia inhibit E. coli growth and damage cell membranes. In order to limit toxic and inhibitory effects different neutralising agents were tested. First the use of NH4OH was optimised with respect to fermentation pH and it was found that the best results were obtained at pH 6.5-6.7. Optimal pH was then used with NaOH, KOH, and Na2CO3 as neutralising agents and it was shown that NaOH, KOH, and Na2CO3 neutralised fermentations could reach succinic acid concentrations of 69 and 61 and 78 g L-1 respectively without any significant decrease in succinic acid productivity. It was observed that cells lost viability during the cause anaerobic phase. It resulted in decreasing succinic acid productivities. It is believed that the viability decrease is a combined effect of organic acids concentration and the osmolarity of the medium. The work done in this thesis is aimed towards increasing the economical feasibility of a biochemical succinic acid production.
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| 13. |
- Berglund, Kris, et al.
(författare)
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Fermentative Upgrading of Xylose
- 2009
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Ingår i: NWBC-2009. - Helsinki : KCL Re-inventing paper.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Chemical pulp mills such as Kraft, soda, or sulfite mills are current examples of biorefineries that can convert lignocellulosic biomass into energy, pulp or cellulose derivatives, and tall oil. While existing viscose pulps use a hemicellulose extraction to generate soluble sugars for ethanol production, in general there still exists a large potential for other more profitable applications of the biomass (Fig. 1), i.e. the mill needs to present a widespread product portfolio. The biofuels under development from fermentation that will be discussed are the diesel fuel oxygenates dibutyl succinate and diethyl succinate to be used for reduced particulate emissions and fossil fuel replacement for diesel engines and butanol for Otto engines. It's important to stress that succinic acid, butanol and ethanol, needed for the production of the diesel additives and gasoline replacement will be produced from renewable resources and hence replacing products currently produced from non-renewable fossil sources. Since wood will be used, there will be no issue of competing with raw material used for food production. Besides biofuel production, succinic acid and butanol, can be used directly or further refined into numerous different products classified as green chemicals.
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| 14. |
- Berglund, Kris
(författare)
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Succinic acid from xylan : fermentations using mixed sugar feedstocks
- 2008
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Ingår i: NWBC 2008. - Stockholm : STFI-Packforsk. - 9789186018139 ; , s. 121-124
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Konferensbidrag (refereegranskat)abstract
- The next generation of cost effective, economically viable biorefinery scenarios requires the development and deployment of higher valued added products whose production can be integrated into a forest biorefinery. We present preliminary results for the production of one such product, succinic acid, from mixed sugars streams including glucose and xylose, as would be present from the hydrolysis of xylan. Examples are presented for the base laboratory case, softwood hydrolysis streams, and hardwood extracts
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| 15. |
- Claus, Michael, et al.
(författare)
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Fruit brandy production by batch column distillation with reflux
- 2005
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Ingår i: Journal of food process engineering. - : Wiley. - 0145-8876 .- 1745-4530. ; 28:1, s. 53-67
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Tidskriftsartikel (refereegranskat)abstract
- The relationship between the operating parameters of batch fruit spirits column stills with reflux and the congener (trace compounds that provide flavors and aromas) concentrations in resulting fruit spirits has not been widely studied. Congener concentrations were determined in three different collection fractions, or "cuts," during batch distillation. Acetaldehyde and ethyl acetate were found in higher concentrations in the head cut, first overhead fraction, of the distillation and have lower boiling points relative to ethanol. 1-Propanol and isoamyl alcohol (isopentanol) were present in higher concentrations in the tail cut, third or final fraction, of the distillation and have boiling points that are higher than ethanol. Methanol has a unique concentration profile as it has higher concentrations in both the head and tail cuts, but a lower concentration in the heart cut, the middle fraction which is the desired product of the distillation. Methanol was of particular interest because the distillate must adhere to governmental regulations that limit its concentration in the product. Operating-condition parameters that were studied include the number of trays used in the distillation as well as the use of a "catalytic converter," a high surface, copper-packing material thought to catalyze formation of cyanide-containing compounds allowing them to be separated from the distillate. The effect of the number of trays used in a distillation on the concentration of ethanol and the congeners, methanol, acetaldehyde, ethyl acetate, 1-propanol and isoamyl alcohol in the final distilled spirits product is presented. An additional result of acetaldehyde production at the copper surface of the catalytic converter was also discovered in the analysis of the data.
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| 16. |
- Claus, Michael J., et al.
(författare)
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Defining still parameters using Chemcad batch distillation model for modeling fruit spirits distillations
- 2009
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Ingår i: Journal of food process engineering. - : Wiley. - 0145-8876 .- 1745-4530. ; 32:6, s. 881-892
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Tidskriftsartikel (refereegranskat)abstract
- Fruit spirits production has always been and continues to be heavily dependent upon the sensory evaluation of the distillate by the distiller. Sensory fatigue can lead to many problems of analysis for the distiller during production. As such, it is desirable to have a method of analysis that is not heavily dependent upon the senses of the distiller. Modeling of the distillation process is difficult because of the number of components present in the fruit mash and distillate, as well as because of the constantly changing thermodynamic interactions present on each tray as a result of the batch distillation process. This paper identifies a method for identifying the still parameters needed to perform predictive modeling. This involves setting a distillate flow rate and reflux ratio, as well as time or volume cutoff frames of analysis. After identifying the still characteristics, it is possible to model the distillation using the CHEMCAD batch distillation program.
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| 17. |
- Enman, Josefine
(författare)
-
Fungal production and solid state chemistry of eritadenine : an integrated approach to development of an active pharmaceutical ingredient
- 2009
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The present thesis demonstrates an integrated approach to the development of a potential active pharmaceutical ingredient, eritadenine, a cholesterol reducing compound originating from the shiitake mushroom (Lentinus edodes). The main areas covered in the thesis are a method for quantification of eritadenine, production of eritadenine by submerged cultivation of fungal mycelia and investigation of the influence of process parameters on mycelial growth and production, and finally solid state characterizations of eritadenine. The usage of the fungus as a source of eritadenine requires an analytical tool for quantification of the compound. An HPLC method was hence developed for identification and quantification of eritadenine, using chemically synthesized eritadenine as a reference. The amount of eritadenine in fruit bodies of selected strains of shiitake was determined and with the method developed in this study, eritadenine concentrations up to ten times higher than previously reported were detected. Since both fruit bodies and mycelia of shiitake have been shown to contain eritadenine submerged cultivation of shiitake mycelia was investigated as an alternative source for this compound. The mycelia were cultivated in various submerged conditions, both in shake flasks and in bioreactors. It was found that both the mycelia and the culture media contained eritadenine, of which the major part was detected in the culture media. While the biomass concentrations were higher in shake flasks, the eritadenine concentrations were considerably higher in the bioreactors, which were assigned to morphological variations. In an attempt to improve the mycelial growth and eritadenine production, a growth promotive substance in the form of a water extract of DDGS, a by-product from drygrind ethanol facilities, was added to the culture media. It was demonstrated that an amendment of the cultivation media with this extract caused a considerable growth promotive effect on shiitake mycelia in bioreactor cultivations, along with enhanced eritadenine production. If eritadenine will be used as a pharmaceutical agent, understanding about the solid state chemistry of the compound is required. Raman spectroscopy is a valuable technique for investigation of structural properties; hence, a Raman reference spectrum with line assignments for the solid state of synthetic eritadenine was established. To further investigate the solid state chemistry of eritadenine, its synthetic analogue was slowly crystallized from water and different ethanol concentrations, at different temperatures. Solids formed from slow cooling of either water or aqueous ethanol showed crystallinity. No polymorphism was detected, irrespective of solvent system or temperature. However, dissimilar thermal behaviours were observed, deducing crystals derived from water as dihydrates and crystals derived from aqueous ethanol as 2.5 hydrates.
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| 18. |
- Enman, Josefine
(författare)
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Production and quantification of eritadenine, a cholesterol reducing compound in shiitake (Lentinus edodes)
- 2007
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Cardiovascular diseases are among the main causes of death in our society and there is a strong correlation between enhanced blood cholesterol levels and the development of such diseases. The popular edible fungus, shiitake mushroom (Lentinus edodes), has been shown to produce a blood cholesterol lowering compound designated eritadenine, and the hypocholesterolemic action of this compound has been quite extensively examined in rats. Eritadenine is suggested to accelerate the removal of blood cholesterol either by stimulating tissue uptake or by inhibiting tissue release; there are no indications of this compound inhibiting the biosynthesis of cholesterol. If shiitake mushrooms are to be used as a source for a potential cholesterol reducing product, it is of great importance to determine the content of eritadenine in the mushrooms as accurately as possible. Hence, in paper I methanol extraction was used to recover as much as possible of the hypocholesterolemic agent from the fungal cells. In order to analyse the target compound, a reliable and reproducible HPLC method for separation, identification and quantification of eritadenine was developed. The amounts of eritadenine in fruit bodies of four commercially cultivated shiitake mushrooms were determined, and the mushrooms under investigation exhibited up to ten times higher levels of eritadenine (3.17-6.33 mg/g dry mushrooms) than previously reported. Not only the fruit bodies of shiitake, but also its mycelia contain eritadenine. Growing fruit bodies of shiitake is a fairly demanding and time consuming process. Hence, in search for a source of eritadenine, submerged (liquid) cultivation of shiitake mycelia could be an alternative. The reason why shiitake mushrooms synthesize eritadenine is yet not clarified; i.e. the function of this secondary metabolite and the growth conditions that favour its production are not elucidated. In addition, like other filamentous fungi, shiitake exhibits different hyphal morphologies in submerged cultures depending on cultivation conditions such as medium composition, temperature, pH, inoculum concentration, dissolved oxygen and shear. The fungal metabolism and hence production of secondary metabolites is in turn affected by the morphology, as have been shown in several studies on filamentous fungi. Submerged cultivation of shiitake mycelia offers a convenient way to change the cultivating conditions in order to improve eritadenine yield and productivity. The study in paper II focused on cultivation of mycelia at different conditions, both in shake flasks and in bioreactors, to investigate the effect of pH and stirring rate on production of eritadenine. The shiitake mycelia were found to produce eritadenine, and the compound of interest was found in both the fungal cells and the growth media. The major part (90-99%) was found in the culture medium, which offers a facilitated downstream processing if large scale production of the compound is to be conducted. The mycelial morphology in the shake flask cultures were macroscopic aggregates, pellets, and the specific productivity of eritadenine was relatively low; 6.56 mg/g dry cell weight (DCW). In the bioreactor cultivations, the mycelia grew as freely dispersed filaments, showing a higher specific productivity than in the shake flasks, ranging between 26.00- 39.58 mg/g DCW. This indicates the influence of morphology on eritadenine production. The biomass yield in shake flasks and bioreactors was in parity; 0.45 g in the shake flasks and 0.25- 0.62 g in the bioreactors. A stirring rate of 50 rpm in the bioreactors was preferable for eritadenine production, whereas for biomass production it was 250 rpm, indicating the influence of agitation on both growth and productivity. The pH did not have any major impact on growth, whereas the specific productivity in the bioreactors was higher when pH was uncontrolled than controlled at 5.7.
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| 19. |
- Enman, Josefine, et al.
(författare)
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Production of the bioactive compound eritadenine by submerged cultivation of shiitake (Lentinus edodes) mycelia
- 2008
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Ingår i: Journal of Agricultural and Food Chemistry. - : American Chemical Society (ACS). - 0021-8561 .- 1520-5118. ; 56:8, s. 2609-2612
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Tidskriftsartikel (refereegranskat)abstract
- Fruit bodies and mycelia of shiitake mushroom (Lentinus edodes) have been shown to contain the cholesterol-reducing compound eritadenine, 2(R),3(R)-dihydroxy-4-(9-adenyl)butyric acid. In the search for a production method for eritadenine, shiitake mycelia were investigated in the present study. The mycelia were cultivated both in shake flasks and in bioreactors, to investigate the effects of pH, stirring rate, and reactor type on the production and distribution of eritadenine. Both the biomass and the culture broth were examined for their eritadenine content. In the shake flasks, the final concentration of eritadenine was 1.76 mg/L and eritadenine was equally distributed between the mycelia and the growth media. In the bioreactors, the shiitake mycelia were found to contain eritadenine in relatively low levels, whereas the majority, 90.6-98.9%, was detected in the growth media. Applying a stirring rate of 250 rpm during bioreactor cultivation resulted in the highest eritadenine concentrations: 10.23 mg/L when the pH was uncontrolled and 9.59 mg/L when the pH was controlled at 5.7. Reducing the stirring rate to 50 rpm resulted in a decreased eritadenine concentration, both at pH 5.7 (5.25 mg/L) and when pH was not controlled (5.50 mg/L). The mycelia in the shake flask cultures appeared as macroscopic aggregates, whereas mycelia cultivated in bioreactors grew more as freely dispersed filaments. This study demonstrates for the first time the extra- and intracellular distribution of eritadenine produced by shiitake mycelial culture and the influence of reactor conditions on the mycelial morphology and eritadenine concentrations.
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| 20. |
- Enman, Josefine, et al.
(författare)
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Quantification of the bioactive compound eritadenine in selected strains of shiitake mushroom (Lentinus edodes)
- 2007
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Ingår i: Journal of Agricultural and Food Chemistry. - : American Chemical Society (ACS). - 0021-8561 .- 1520-5118. ; 55:4, s. 1177-1180
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Tidskriftsartikel (refereegranskat)abstract
- Cardiovascular disease is one of the most common causes of death in the Western world, and a high level of blood cholesterol is considered a risk factor. The edible fungus, shiitake mushroom (Lentinus edodes), contains the hypocholesterolemic agent eritadenine, 2(R),3(R)-dihydroxy-4-(9-adenyl)-butyric acid. This study was conducted to quantify the amount of the cholesterol reducing agent eritadenine in shiitake mushrooms, in search of a potential natural medicine against blood cholesterol. The amounts of eritadenine in the fruit bodies of four different shiitake mushrooms, Le-1, Le-2, Le-A, and Le-B, were investigated in this study. To achieve this goal, methanol extraction was used to recover as much as possible of the hypocholesterolemic agent from the fungal cells. In addition, enzymes that degrade the fungal cell walls were also used to elucidate if the extraction could be further enhanced. To analyze the target compound, a reliable and reproducible HPLC method for separation, identification, and quantification of eritadenine was developed. The shiitake strains under investigation exhibit up to 10 times higher levels of eritadenine than previously reported for other shiitake strains. Further, pretreating the mushrooms with hydrolytic enzymes before methanol extraction resulted in an insignificant increase in the amount of eritadenine released. These results indicate the potential for delivery of therapeutic amounts of eritadenine from the ingestion of extracts or dried concentrates of shiitake mushroom strains.
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| 21. |
- Helmerius, Jonas, et al.
(författare)
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Production of value added chemicals from xylan extraction in a Kraft pulp mill and the effect on pulp quality
- 2008
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- In the Kraft process hemicelluloses are lost in the cooking procedure to the black liquor stream, which is subsequently burnt in the recovery boiler to recover cooking chemicals and to produce steam and energy. Hemicelluloses have a low heating value compared to lignin and therefore recovery of hemicelluloses at an earlier stage of the Kraft process followed by biochemical conversionintohighvalue-conversion intohighvalue-into high value-added products might offer a muchbettereconomicopportunity.much better economic opportunityIn collaboration with the research and development department of Smurfit Kappa Kraftliner AB, Piteå, Sweden, alkali and water extractions of birch wood were performed under conditions compatible with the Kraft process, at different times, temperatures and alkali charges. The extraction conditions were set in a range suitable with the current pulp process at Smurfit Kappa Kraftliner. TherequirementsforprocessThe requirements for process configurations, based on either hot water or alkali extraction were also explored. ThexylanyieldsfromdifferentextractiontrialswereThe xylan yields from different extraction trials were measured and the chips from those extraction trials were cooked, refined and turned into sheets of paper. The effects on paper quality were compared with a reference pulp made from the same wooden chips. Recovered xylans from water extracted birch wood chips were subjected to secondary hydrolysis, enzymatic or sulphuricacid.sulphuric acidDetoxification of the hydrolysate with active carbon and regulation of pH were performed before fermentation. FermentationofthexyloseFermentation of the xylose hydrolysate to succinic acid was demonstrated by the use of thethe succinic acid producer Escherichia coli AFP184.
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| 22. |
- Helmerius, Jonas, et al.
(författare)
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Production of value added chemicals from xylan extraction in a Kraft pulp mill and the effect on pulp quality
- 2008
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- In the Kraft process hemicelluloses are lost in the cooking procedure to the black liquor stream, which is subsequently burnt in the recovery boiler to recover cooking chemicals and to produce steam and energy. Hemicelluloses have a low heating value compared to lignin and therefore recovery of hemicelluloses at an earlier stage of the Kraft process followed by biochemical conversion into high value-added products might offer a much better economic opportunity. In collaboration with the research and development department of Smurfit Kappa Kraftliner AB, Piteå, Sweden, alkali and water extractions of birch wood were performed under conditions compatible with the Kraft process, at different times, temperatures and alkali charges. The extraction conditions were set in a range suitable with the current pulp process at Smurfit Kappa Kraftliner. The requirements for process configurations, based on either hot water or alkali extraction were also explored. The xylan yields from different extraction trials were measured and the chips from those extraction trials were cooked, refined and turned into sheets of paper. The effects on paper quality were compared with a reference pulp made from the same wooden chips. Recovered xylans from water extracted birch wood chips were subjected to secondary hydrolysis, enzymatic or sulphuric acid. Detoxification of the hydrolysate with active carbon and regulation of pH were performed before fermentation. Fermentation of the xylose hydrolysate to succinic acid was demonstrated by the use of the succinic acid producer Escherichia coli AFP184.
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| 23. |
- Hodge, David B., et al.
(författare)
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Soluble and insoluble solids contributions to high-solids enzymatic hydrolysis of lignocellulose
- 2008
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Ingår i: Bioresource Technology. - : Elsevier BV. - 0960-8524 .- 1873-2976. ; 99:18, s. 8940-8948
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Tidskriftsartikel (refereegranskat)abstract
- The rates and extents of enzymatic cellulose hydrolysis of dilute acid pretreated corn stover (PCS) decline with increasing slurry concentration. However, mass transfer limitations are not apparent until insoluble solids concentrations approach 20% w/w, indicating that inhibition of enzyme hydrolysis at lower solids concentrations is primarily due to soluble components. Consequently, the inhibitory effects of pH-adjusted pretreatment liquor on the enzymatic hydrolysis of PCS were investigated. A response surface methodology (RSM) was applied to empirically model how hydrolysis performance varied as a function of enzyme loading (12-40 mg protein/g cellulose) and insoluble solids concentration (5-13%) in full-slurry hydrolyzates. Factorial design and analysis of variance (ANOVA) were also used to assess the contribution of the major classes of soluble components (acetic acid, phenolics, furans, sugars) to total inhibition. High sugar concentrations (130 g/L total initial background sugars) were shown to be the primary cause of performance inhibition, with acetic acid (15 g/L) only slightly inhibiting enzymatic hydrolysis and phenolic compounds (9 g/L total including vanillin, syringaldehyde, and 4-hydroxycinnamic acid) and furans (8 g/L total of furfural and hydroxymethylfurfural, HMF) with only a minor effect on reaction kinetics. It was also demonstrated that this enzyme inhibition in high-solids PCS slurries can be approximated using a synthetic hydrolyzate composed of pure sugars supplemented with a mixture of acetic acid, furans, and phenolic compounds, which indicates that generally all of the reaction rate-determining soluble compounds for this system can be approximated synthetically.
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| 24. |
- Hodge, David, et al.
(författare)
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Detoxification requirements for bioconversion of softwood dilute acid hydrolyzates to succinic acid
- 2009
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Ingår i: Enzyme and microbial technology. - : Elsevier BV. - 0141-0229 .- 1879-0909. ; 44:5, s. 309-316
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Tidskriftsartikel (refereegranskat)abstract
- In this work an Escherichia coli metabolically engineered to ferment lignocellulosic biomass sugars to succinic acid was tested for growth and fermentation of detoxified softwood dilute sulfuric acid hydrolyzates, and the minimum detoxification requirements were investigated with activated carbon and/or overliming treatments. Detoxified hydrolyzates supported fast growth and complete fermentation of all hydrolyzate sugars to succinate at yields comparable to pure sugar, while untreated hydrolyzates were unable to support either growth or fermentation. Activated carbon treatment was able to remove significantly more HMF and phenolics than overliming. However, in some cases, overliming treatment was capable of generating a fermentable hydrolyzate where activated carbon treatment was not. The implications of this are that in addition to the known organic inhibitors, the changes in the inorganic content and/or composition due to overliming are significant to the hydrolyzate toxicity. It was also found that any HMF remaining after detoxification was completely metabolized during aerobic cell growth on the hydrolyzates that were capable of supporting growth.
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| 25. |
- Hodge, David, et al.
(författare)
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Impact of hemicellulose pre-extraction for bioconversion on birch kraft pulp properties
- 2009
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- The carbohydrate portion of lignocellulosic feedstocks are ideally suited to conversion via biochemical transformations because of their crucial role in cellular metabolism. The combination of hemicelluloses extraction with pulping processes could be one way to generate a sugar feedstock amenable to biochemical transformation to fuels and chemical intermediates. White liquor, green liquor, and water HC extractions of birch wood were performed under conditions compatible with the Kraft process, at different times, temperatures and alkali charges. The effective alkali charge was in extractions between 0%-7% and temperature between 130°C-160°C for 20-90 minutes. The xylan yields from different HC extractions were measured and the chips from select HC extractions were cooked, and the refined pulps were made into hand sheets. Several metrics for hand sheet quality were compared with a reference pulp made from the same wood chips. It is possible using white liquor to extract xylan from birch wood chips prior Kraft cooking without decreasing the pulp yield and paper strength properties, and at the same time achieve an impregnation of the wood chips. It is not possible in that extraction to attain extracted and hydrolyzed liquor containing a fermentable concentration of xylose, 2.63 g/L in this study. Increased extracted wood material, increased final acetic acid concentration and decreased final xylan concentration together with increased effective alkali charge at the same extraction temperature and time in white liquor extractions performed support that xylan degradation increases. Using white liquor or green liquor under the conditions investigated degrades xylan resulting in significant losses of xylose that could have been used as substrate in fermentation processes.
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