| 1. |
- Cavonius, Lillie, 1981, et al.
(författare)
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Effects of pH-shift processing on the bioaccessibility of lipids in the microalgae Nannochloropsis – an in vitro model approach
- 2015
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Ingår i: Lipidforum Symposium, Reykjavik, June 2015.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Introduction: Microalgae are a potential source of human nutrition and animal feed due to the high content of long-chained n-3 fatty acids (LC n-3 PUFA) and favorable amino acid profile of some species. Furthermore, microalgae can be cultivated with minimal nutrient requirements on otherwise non-arable land. We have previously shown that application of the pH-shift process on Nannochloropsis yields a product enriched in essential amino acids and with the original amount of LC n-3 PUFA retained (120 mg/g dry weight). Therefore, the product has potential as a functional food and/or feed ingredient. However, it is not known to which extent the nutrients are bio-accessible; earlier work has suggests that algal cell walls limit digestibility in the gastrointestinal tract. Based on the physical changes that take place during pH-shift processing, e.g. cell lysis and protein solubilization/precipitation, we hypothesize that the pH-shift process renders the algal nutrients more available for hydrolysis. The aim of the present study is to test this hypothesis using a static in vitro digestion model.Method: Nannochloropsis is subjected to the pH-shift process and nutrient content (fatty acids, protein, amino acids) is determined before/after processing. To determine the bio-accessibility of Nannochloropsis processed in various ways, the microalgae is digested using a static in vitro model of human digestion, based on the recent international consensus method emerging from the COST FA 1005 Infogest. Intestinal digesta will then be analyzed in respect to degree of lipolysis using solid phase extraction (SPE) combined with gas chromatography-mass spectrometry (GC-MS).Results and conclusion: The study is ongoing, and results will be presented in June.
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| 2. |
- Cavonius, Lillie, 1981, et al.
(författare)
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Fatty acid contaminations originating from commercially available solid phase-extraction columns
- 2015
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Ingår i: Chemical Science Review and Letters. - 2278-6783. ; 4:16, s. 1206-1209
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Tidskriftsartikel (refereegranskat)abstract
- It has been shown previously that commercially available solid phase extraction (SPE) columns release contaminants such as palmitic acid and stearic acid during separation of lipid classes. The presence of contaminating fatty acids in the fatty acid fraction is particularly troublesome. We here confirm that the overwhelming majority of the contaminants originate from the barrels, and have identified two contaminants as palmitic acid and stearic acid or their equivalents. We urge readers to take fatty acid contaminants into careful consideration when planning their experiments, and if necessary use sorbent packed in glass barrels instead of plastic such as polypropylene.
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| 3. |
- Cavonius, Lillie, 1981
(författare)
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Fractionation of lipids and protein from the microalga Nannochloropsis oculata
- 2016
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- To feed Earth’s growing population, microalgae have been proposed as a source of protein and long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFAs). Currently, microalgae are mainly sold either as whole biomass, or the economically valuable LC n-3 PUFA fraction is extracted with organic solvents. As an alternative to solvent extraction, pH-driven protein solubilization in water followed by isoelectric precipitation, known as the pH-shift process, has been proposed to recover both lipids (i.e. oils) and functional proteins, while removing insoluble material.In this project, the pH-shift process was applied to Nannochloropsis oculata, a marine microalga containing LC n-3 PUFA and essential amino acids in more than adequate amounts for human nutrition. It was hypothesized that the pH-shift process would render separate lipid and protein fractions. A pH-shift process for Nannochloropsis was developed based on documentation of its pH-dependent protein solubility: Nannochloropsis proteins were solubilized at pH 7, insoluble material was removed by centrifugation at 4 000×g and the proteins were then recovered by precipitation at pH 3. By using seawater, a process in which algal culture medium was used directly in the pH-shift process was simulated, thus reducing freshwater consumption. Contrary to the hypothesis, the developed process did not result in two fractions, but in a combined product of 2.3% LC n-3 PUFA and 23% protein per dry weight, compared to 1.9% and 19% respectively in the initial material, suggesting that the product had potential as a functional food ingredient. However, nutrients need to also be accessible for uptake by the gastrointestinal tract. To assess the link between processing and accessibility of Nannochloropsis fatty acids and proteins, a static in vitro digestion model was applied to whole Nannochloropsis, and various products of the pH-shift process. The results indicated that whole Nannochloropsis cannot be digested by mammalian enzymes at all, hence the lipids and proteins cannot be absorbed. However, in cell-disrupted Nannochloropsis ca. 35% fatty acids and protein were hydrolyzed, with hydrolysis somewhat increased with further pH-shift processing. This project indicates that the nutritional profile of Nannochloropsis oculata is favorable for human consumption, but that cell disruption is paramount to make the lipids and proteins accessible to the digestive enzymes. The pH-shift process applied here provided such cell disruption, slightly increased the concentration of proteins and lipid digestibility, and demonstrated a scalable process.
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| 4. |
- Cavonius, Lillie, 1981, et al.
(författare)
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Glazing herring (Clupea harengus) fillets with herring muscle press juice: effect on lipid oxidation development during frozen storage
- 2017
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Ingår i: International Journal of Food Science and Technology. - : Wiley. - 1365-2621 .- 0950-5423. ; 52:5, s. 1229-1237
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Tidskriftsartikel (refereegranskat)abstract
- In this study, fish muscle press juice, PJ, was evaluated as an antioxidative liquid for glazing of fish filletsprior to frozen storage. PJ has in previous studies efficiently protected fish mince lipids against haemoglobin-mediated oxidation. Frozen herring fillet were glazed with water, PJ or 50 mM phosphate buffer of thesame pH as herring PJ (6.5). Over 52 weeks, peroxide value (PV), TBA-reactive substances (TBARS), rancidodour and colour were monitored. From 26 to 52 weeks, water-glazed fillets were significantly(P
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| 5. |
- Cavonius, Lillie, 1981, et al.
(författare)
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Imaging lipids in live microalgae
- 2013
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Ingår i: Algae Biomass Summit 2013.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Microalgae are capable of producing lipids from CO2 and sunlight and as such the primary producers of n-3 fatty acids. Intense research is underway to understand the conditions under which optimal lipid accumulation occurs, not only for neutraceutical applications, but also for biodiesel production. To aid this research we propose the application of a powerful microscopic technique that allows monitoring of lipids with chemical specificity at intra-cellular level in living cells: Coherent anti-Stokes Raman spectroscopy (CARS). CARS is a non-linear microscopy technique that can be used to probe C-H bonds especially abundant in lipids by a process involving four-wave mixing: two or three coherent beams of different near-infra red (NIR) wavelengths are tuned to induce a resonant vibration in the C-H bonds, and generate a blue-shifted CARS signal. The NIR light used to probe the sample allows good penetration which in turn makes optical sectioning possible. By taking many optical sections of the sample, a 3D image can be constructed, from which the volume of lipids in the cell can be calculated, allowing quantitative studies of lipid accumulation in single microalgae. In our study, we used Phaeodactylum tricornutum grown under normal, light-starved and nitrogen-starved conditions. CARS microscopy detected statistically significant differences in lipid droplet number and their volumes when comparing growth conditions at single cell level. The whole population was then subjected to traditional lipid extraction and chromatographic separation of fatty acids. Average lipid volumes, as calculated from CARS microscopy, correlated well with traditional chemical analysis. As a conclusion, CARS could be applied to the in vivo study of culture condition effect on lipid accumulation in microalgae.
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| 6. |
- Cavonius, Lillie, 1981, et al.
(författare)
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Imaging lipids in live microalgae
- 2012
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Ingår i: Novel Sources of Omega-3 in Food and Feed, Copenhagen 2012.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Microalgae are the primary producers of n-3 fatty acids in the aquatic food web, capable of producing lipids from CO2 and sunlight. Intense research is underway to understand the conditions under which optimal lipid accumulation occurs. To aid this research we propose the application of a powerful microscopic technique that allows monitoring of lipids with chemical specificity at intra-cellular level in living cells. While fluorescence microscopy with an appropriate dye can be highly specific, cells must be fixed and permeabilized, excluding live-cell studies; other techniques, such as phase contrast microscopy can be applied to living cells, but they lack chemical specificity. Coherent anti-Stokes Raman spectroscopy (CARS) is a non-linear microscopy technique that probes C-H bonds especially abundant in lipids by a process involving four-wave mixing: two or three coherent beams of different near-infra red (NIR) wavelengths are tuned to induce a resonant vibration in the C-H bonds, and generate a blue-shifted CARS signal in the visible region. Multi-photon autofluorescence from e.g. pigments in the sample can be detected simultaneously. The NIR light used to illuminate the sample allows good penetration which in turn makes optical sectioning possible. By taking many optical sections of the sample a 3D image can be constructed, from which the volume of lipids in the cell can be calculated, allowing quantitative studies of lipid accumulation in single microalgae. We herein present a proof-of-concept in the comparison of Phaeodactylum tricornutum grown under normal light-starved and nitrogen-starved conditions.
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| 7. |
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| 8. |
- Cavonius, Lillie, 1981, et al.
(författare)
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Imaging of Lipids in Microalgae with Coherent Anti-Stokes Raman Scattering Microscopy
- 2015
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 1532-2548 .- 0032-0889. ; 167:3, s. 603-616
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Tidskriftsartikel (refereegranskat)abstract
- Microalgae have great prospects as a sustainable resource of lipids for refinement into nutraceuticals and biodiesel, which increases the need for detailed insights into their intracellular lipid synthesis/storage mechanisms. As an alternative strategy to solvent- and label-based lipid quantification techniques, we introduce time-gated coherent anti-Stokes Raman scattering (CARS) microscopy for monitoring lipid contents in living algae, despite strong autofluorescence from the chloroplasts, at approximately picogram and subcellular levels by probing inherent molecular vibrations. Intracellular lipid droplet synthesis was followed in Phaeodactylum tricornutum algae grown under (1) light/nutrient-replete (control [Ctrl]), (2) light-limited (LL), and (3) nitrogen-starved (NS) conditions. Good correlation (r2 = 0.924) was found between lipid volume data yielded by CARS microscopy and total fatty acid content obtained from gas chromatography-mass spectrometry analysis. In Ctrl and LL cells, micron-sized lipid droplets were found to increase in number throughout the growth phases, particularly in the stationary phase. During more excessive lipid accumulation, as observed in NS cells, promising commercial harvest as biofuels and nutritional lipids, several micron-sized droplets were present already initially during cultivation, which then fused into a single giant droplet toward stationary phase alongside with new droplets emerging. CARS microspectroscopy further indicated lower lipid fluidity in NS cells than in Ctrl and LL cells, potentially due to higher fatty acid saturation. This agreed with the fatty acid profiles gathered by gas chromatography-mass spectrometry. CARS microscopy could thus provide quantitative and semiqualitative data at the single-cell level along with important insights into lipid-accumulating mechanisms, here revealing two different modes for normal and excessive lipid accumulation.
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| 9. |
- Cavonius, Lillie, 1981, et al.
(författare)
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In vitro bioaccessibility of proteins and lipids of pH-shift processed Nannochloropsis oculata microalga
- 2016
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Ingår i: Food and Function. - : Royal Society of Chemistry (RSC). - 2042-6496 .- 2042-650X. ; 7:4, s. 2016-2024
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Tidskriftsartikel (refereegranskat)abstract
- The pH-shift process fractionates biomass into soluble proteins and insoluble fractions, followed by precipitation and recovery of the solubilized proteins. Nannochloropsis oculata in seawater was subjected to the pH-shift process, followed by digestion of various intermediates and product fractions of the process, using the Infogest in vitro digestion model (Minekus et al., 2014) with added gastric lipase. As measures for protein and lipid accessibility, degrees of protein hydrolysis and fatty acid liberation were assessed post-digestion and compared to the amounts of peptide bonds and total fatty acids present in the raw materials. Results showed that neither proteins nor lipids of intact Nannochloropsis cells were accessible to the mammalian digestive enzymes used in the digestion model. Cell disruption, and to a lesser extent, further pH-shift processing with protein solubilisation at pH 7 or pH 10, increased the accessibility of lipids. For proteins, differences amongst the pH-shift processed materials were non-significant, though pre-freezing the product prior to digestion increased the accessibility from 32% to 47%. For fatty acids, pH-shift process-products gave rise to 43% to 52% lipolysis, with higher lipolysis for products solubilised at pH 10 as opposed to pH 7. Our results indicate the importance of processing to produce an algal product that has beneficial nutritional properties when applied as food or feed.
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| 10. |
- Cavonius, Lillie, 1981, et al.
(författare)
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pH-shift processing of Nannochloropsis oculata microalgal biomass to obtain a protein-enriched food or feed ingredient
- 2015
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Ingår i: Algae Biomass Summit, Washington D.C., 2015.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- lthough some species of microalgae have potential as human and animal nutrition due to the presence of high-value protein and polyunsaturated fatty acids, there has been relatively little development of algal food products. Fractionation of plant and animal raw materials by pH-shift processing has been widely applied to purify proteins. The principle is to solubilize proteins at high or low pH, removing insoluble material and precipitating the proteins near or at their isoelectric point. We have applied the pH-shift process to the commercially-available, marine microalga Nannochloropsis oculata and studied the partitioning of major nutrients into the various fractions emerging from the process. The process was developed and adapted to the biomass with scalability in mind. Proteins were found to exhibit maximal solubility between pH 7 and 10, with a minimal solubility below pH 4. Two process versions were investigated in this study, with solubilization at either pH 7 (native pH) or 10; in both versions, proteins were efficiently precipitated at pH 3. Up to 85 % of both the protein and total fatty acids were recovered in the final product, compared to the initial algal slurry. The amino acid profile was not altered by the process, apart from a minor increase in phenylalanine, counteracted by a minor decrease in proline. From a processing point of view, solubilization of Nannochloropsis at native pH was found to be preferable, since less chemicals were consumed compared to high-pH solubilization. The final product was on a dry-weight basis 23 % protein (of which 43 % were essential amino acids), 12 % total fatty acids (of which 18 % were eicosapentaenoic acid), 37 % carbohydrates and 34 % ash, and thus has potential as a functional food ingredient.
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