Digestive enzymes characterization of krill (Euphausia superba) residues deoiled by supercritical carbon dioxide and organic solvents

In this study, Enzyme activities of krill were characterized before and after lipid extraction by supercritical carbon dioxide (SC-CO₂) and organic solvent, n-hexane and acetone. Krill SC-CO₂ extraction was performed under the conditions of temperature range from 35 to 45 °C and pressure, 150–250 bar for 2.5 h with a constant flow rate of 22 g/min. Extraction yields of lipids increased with pressure and temperature. The digestive enzyme activities of protease, lipase and amylase of SC-CO₂ treated krill residues were slightly decreased comparing to organic solvent, n-hexane and acetone treated residues. In SC-CO₂ treated samples, all of the digestive enzymes showed slightly higher temperature stability. In the other hand the crude extracts of SC-CO₂ and n-hexane treated krill samples showed almost same optimum pH and pH stability for each of the digestive enzymes. It was also found in SDS-PAGE that there are no significant differences in protein patterns of the crude extracts of untreated, SC-CO₂, n-hexane and acetone treated krill indicating no denaturation of proteins.     

Characterization of oil including astaxanthin extracted from krill (Euphausia superba) using supercritical carbon dioxide and organic solvent as comparative method

Krill oil including astaxanthin was extracted using supercritical CO₂ and hexane. The effects of different parameters such as pressure (15 to 25MPa), temperature (35 to 45 °C), and extraction time, were investigated. The flow rate of CO₂ (22 gmin⁻¹) was constant for the entire extraction period of 2.5 h. The maximum oil yield was found at higher extraction temperature and pressure. The oil obtained by SC-CO₂ extraction contained a high percentage of polyunsaturated fatty acids, especially EPA and DHA. The acidity and peroxide value of krill oil obtained by SC-CO₂ extraction were lower than that of the oil obtained by hexane. The SC-CO₂ extracted oil showed more stability than the oil obtained by hexane extraction. The amount of astaxanthin in krill oil was determined by HPLC and compared at different extraction conditions. The maximum yield of astaxanthin was found in krill oil extracted at 25 MPa and 45 °C.     

Effect of reaction parameters on conversion of krill (Euphausia superba) oil by immobilized lipase ethanolysis

Monoglyceride and diglyceride were produced by performing ethanolysis of krill oil with immobilized lipase and the influence of various parameters on the enzymatic ethanolysis was assessed. As an immobilized lipase, lipozyme TL-IM (thermonuces lanuginose) was used. Ethanolysis was done in non-pressurized and pressurized system to compare the reaction rate and yield. The optimal condition was found at 2.0 of ethanol mole ratio, temperature of 60 °C, lipases amount of 5 wt% in non-pressurized system. At pressurized system the optimal temperature and pressure was found at 50 °C and 10 MPa. However, at 50 °C monoglyceride was higher in pressurized system than in non-pressurized system.     

Lipid,sterol and fatty acid composition of antarctic krill (Euphausia superba Dana)

The lipid classes, fatty acids of total and individual lipids and sterols of Antarctic krill (Euphausia superba Dana) from two areas of the Antarctic Ocean were analyzed by thin layer chromatography (TLC), gas liquid chromatography (GLC) and gas liquid chromatography/mass spectrometry (GLC/MS). Basic differences in the lipid composition of krill from the Scotia Sea (caught in Dec. 1977) and krill from the Gerlache Strait (caught in Mar. 1981) were not observed. The main lipid classes found were: phosphatidylcholine (PC) (33–36%), phosphatidylethanolamine (PE) (5–6%), triacylglycerol (TG) (33–40%), free fatty acids (FFA) (8–16%) and sterols (1.4–1.7%). Wax esters and sterol esters were present only in traces. More than 50 fatty acids could be identified using GLC/MS, the major ones being 14∶0, 16∶0, 16∶1(n−7), 18∶1(n−9), 18∶1(n−7), 20∶5(n−3) and 22∶6(n−3). Phytanic acid was found in a concentration of 3% of total fatty acids. Short, medium-chain and hydroxy fatty acids (C≤10) were not detectable. The sterol fraction consisted of cholesterol, desmosterol and 22-dehydrocholesterol.     

Extraction of phospholipids from canola with supercritical carbon dioxide and ethanol

The potential use of supercritical (SC)-CO₂/ethanol mixture for the extraction and fractionation of phospholipids (PL) from flaked canola seeds, canola meal, and acetone insolubles (AI) was investigated. PL extraction was possible when ethanol was used as a cosolvent in SC-CO₂. PL recovery of 20.8% was achieved when canola flakes were extracted at 70°C and 55.2 MPa with SC-CO₂/10%EtOH after iol removal with neat SC-CO₂. Soaking of canola meal with ethanol prior to SC-CO₂/EtOH extraction increased PL recovery to 30.4%. PL content of the extracts increased with decreasing triglyceride concentration in the feed material and increasing amounts of ethanol added to SC-CO₂ or used for soaking. Fractionation of Al gums resulted in extracts containing 50% PL, of which 90% was phosphatidylcholine (PC); but yields were low, even after soaking treatment, due to caking. SC-CO₂/EtOH mixture may be used to extract PC-enriched PL from flaked canola seeds, canola meal, and AI. However, further research is needed to improve extraction efficiency.     

Oxidative stability and non‐enzymatic browning reactions in Antarctic krill oil (Euphausia superba)

Antarctic krill oil has gained much consideration recently due to its rich content of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in the form of phospholipids and its powerful antioxidant known as astaxanthin. To secure these valuable bioactive nutrients in krill oil, a gentle and immediate on‐board processing of freshly captured krill is recommended. Compared to fish oil, krill oil has a more complex matrix, which leads to the formation of additional compounds from non‐enzymatic browning reactions. Lipid oxidation occurs through different pathways in krill oil and cannot be detected through classical analytical techniques such as determination of peroxide and anisidine value. Therefore selection of appropriate methods to evaluate the oxidative stability of krill oil is of high importance.

Figure 1 Open in figure viewer PowerPoint Development of lipid derived volatiles: (a & b) 1‐penten‐3‐ol and (c & d) 2‐pentylfuran in krill oil upon storage at two different incubation temperatures (20 and 40°C). The same pattern as (a & b) was obtained for (Z)‐2‐penten‐1‐ol and benzaldehyde, whereas the same pattern as (c & d) was obtained for 2‐heptanone and 2‐octanone. Values are mean ± SD (n = 3). Means sharing the same letter are not significantly different at 5% significant level. Reproduced from Lu et al. 2 with permission from Elsevier.     

Cloning and Characterization of Fructose-1,6-Bisphosphate Aldolase from Euphausia superba

Fructose-1,6-bisphosphate aldolase (EC 4.1.2.13) is a highly conserved enzyme that is involved in glycolysis and gluconeogenesis. In this study, we cloned the fructose-1,6-bisphosphate aldolase gene from Euphausia superba (EsFBA). The full-length cDNA sequence of EsFBA is 1098 bp long and encodes a 365-amino-acid protein. The fructose-1,6-bisphosphate aldolase gene was expressed in Escherichia coli (E. coli). A highly purified protein was obtained using HisTrap HP affinity chromatography and size-exclusion chromatography. The predicted three-dimensional structure of EsFBA showed a 65.66% homology with human aldolase, whereas it had the highest homology (84.38%) with the FBA of Penaeus vannamei. Recombinant EsFBA had the highest activity at 45 °C and pH 7.0 in phosphate buffer. By examining the activity of metal ions and EDTA, we found that the effect of metal ions and EDTA on EsFBA’s enzyme activity was not significant, while the presence of borohydride severely reduced the enzymatic activity; thus, EsFBA was confirmed to be a class I aldolase. Furthermore, targeted mutations at positions 34, 147, 188, and 230 confirmed that they are key amino acid residues for EsFBA.     

Characterization and supercritical carbon dioxide extraction of walnut oil

Walnut (Juglans regia L.) oil was extracted with compressed carbon dioxide (CO₂) in the temperature range of 308 to 321 K and in the pressure range of 18 to 23.4 MPa. The influence of particle size was also studied at a superficial velocity of 0.068 cm/s, within a tubular extractor of 0.2 L capacity (cross-sectional area of 16.4 cm²). FFA, sterol, TAG, and tocopherol compositions were not different from those of oil obtained with n-hexane. The main FA was linoleic acid (56.5%), followed by oleic acid (21.2%) and linolenic acid (13.2%). The main TAG was LLL (linoleic, linoleic, linoleic) (24.4%), followed by OLL (oleic, linoleic, linoleic) (19.6%) and LLLn (linoleic, linoleic, linolenic) (18.4%). The main component of sterols was β-sitosterol (85.16%), followed by campesterol (5.06%). The amount of cholesterol was low (0.31 and 0.16% for oils extracted by n-hexane and supercritical fluid extraction, respectively. The CO₂-extracted oil presented a larger amount of tocopherols (405.7 μg/g oil) when compared with 303.2 μg/g oil obtained with n-hexane. Oxidative stability determined by PV and the Rancimat method revealed that walnut oil was readily oxidized. Oil extracted by supercritical CO₂ was clearer than that extracted by n-hexane, showing some refining. A central composite, nonfactorial design was used to optimize the extraction conditions using the software Statistica, Version 5. The best results were found at 22 MPa, 308 K, and particle diameter (Dp) −0.1 mm.     

Concentration of tocopherols from soybean sludge by supercritical carbon dioxide

A supercritical fluid extraction method has been applied to test the feasibility of tocopherol concentration from soybean sludge with carbon dioxide at temperatures and pressures ranging from 35 to 70°C and 200 to 400 bar, respectively. The supercritical solubility of the esterified soybean sludge was over 4–6 times greater than that of the original soybean sludge. By a simple batch-type one-stage method the tocopherols in the esterified soybean sludge could be concentrated up to 40 wt%. The overall results of the present study show that soybean sludge initially containing about 13–14 wt% tocopherols may require a countercurrent multistage column to be highly and effectively concentrated.     

Characterization of wheat bran oil obtained by supercritical carbon dioxide and hexane extraction

Supercritical carbon dioxide (SC-CO₂) and soxhlet extraction using was carried out to extract oil from wheat bran oil. For SC-CO₂, the pressure and temperature were ranging from 10 to 30 MPa and 313.15–333.15 K. The extraction was performed in a semi batch process with a CO₂ flow rate of 26.81 g/min for 2 h. Wheat bran oil was characterized to investigate the quality. Acid value (AV) and peroxide value (POV) were higher in hexane extracted oil compared to SC-CO₂ extracted oil. Induction period was measured by rancimat test. The oil obtained by SC-CO₂ extraction had higher capability to delay the oxidation by surrounding environment. The DPPH radical scavenging activity was also measured. The SC-CO₂ extracted oil showed higher radical scavenging activity compared to hexane extracted oil.     

Characterization and processing of cottonseed oil obtained by extraction with supercritical carbon dioxide

Extraction of flaked cottonseed with supercritical carbon dioxide at temperatures of 50–80 C and pressures of 8,000–15,000 psi yields an improved crude cottonseed oil compared to those obtained by conventional solvent or expeller processes. Improvements include lighter initial color, less refining loss and lighter refined bleached colors. Crude cottonseed oils obtained by supercritical fluid extraction require less refining lye and show less tendency to undergo color fixation while in storage. Presented at the AOCS annual meeting, Chicago, May 1983.     

Characterization of expanded polystyrene and its composites by supercritical carbon dioxide foaming approach

Journal of Porous Materials - Supercritical carbon dioxide (scCO2) has been used as a physical blowing agent to produce polymer expanded materials. Firstly, a statistic study was performed to...     

Extraction of essential oil from geranium (Pelargonium graveolens) with supercritical carbon dioxide

This study investigated the supercritical fluid extraction (SFE) of geranium essential oil from geranium (Pelargonium graveolens) using supercritical carbon dioxide solvent. The extraction yield was measured as a function of pressure, temperature and carbon dioxide flow rate. At low pressure (10 MPa) and high temperature (343 K), waxes were co‐extracted with the essential oil, resulting in artificially elevated essential oil extraction yields as no method was available with the SFE apparatus used to separate co‐extracted waxes and oil. At high pressure (30 MPa) and low temperature (313 K), the amount of wax co‐extracted decreased. Under these ‘optimum’ conditions, the extraction yield increased with decrease in flow rate giving a maximum extraction yield of 2.53%. All samples were analyzed by gas chromatography–mass spectrometry and the effect of pressure and extraction time on oil composition was studied. The percentage compositions of terpene hydrocarbons, terpenols, geraniol and geranyl esters were significantly affected by pressure and extraction time. The oil samples obtained by SFE were also compared with commercially obtained steam distilled samples. All major components of the commercially obtained oils were present in the SFE‐obtained oils; however, the percentage composition of the major components differed greatly between steam distilled and SFE oils. Copyright © 2005 Society of Chemical Industry     

Elastomeric foam prepared by supercritical carbon dioxide

Biodegradable elastomer poly[(1,4‐butylene terephthalate)‐co‐(1,4‐butylene adipate)] foam was successfully prepared using supercritical CO₂. The elastomer foam has closed and uniform cell structure with density about 90 g/L. Narrow size distribution with average cell diameter 32 μm were obtained. The foamed balls show rubbery properties and can recover well (>90%) to their original shape quickly after releasing press stress. Cell growth needs to take place at melting state where the migration and orientation of polymer chains are permanently kept without remained force of springback. Some unique phenomena were observed during foaming the elastomer: post‐expansion (about 40%) was obtained after removing the foamed samples from a cylinder mold, which results in forming a ball rather than a cylinder like the mold. This phenomenon was explained by the lower glass transition temperature of the elastomer, in which residual CO₂ foam continuously in the room temperature; and “memory” of internal strength. © 2016 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2017 , 134, 44354.     

Controlling cellular morphology by supercritical carbon dioxide

A series of high‐performance porous materials, including open‐cell and closed‐cell foams, were prepared by a two‐stage batch foaming process from fluorinated poly(arylene ether)s, and the porous morphology was characterized with scanning electron microscopy. The effects of saturation pressures, saturation temperatures, transfer times, molecular structures, and solvent traces on the cell sizes, cell densities, and bulk densities of porous materials were examined. According to the conclusions, the porous structures could be controlled with cellular sizes between the nanoscale and microscale. All nanoporous materials have dielectric constants in the ultralow‐dielectric‐constant range (1.4–2.0). The open structures can be prepared by two methods, and there are two kinds of morphologies from the two methods, including foams with small, spotlike openings (diameters between 10 and 100 nm) in the cell walls and bicontinuous nanoporous foams. Another special method for controlling the porous morphology was used here. Films with closed‐cell microcellular foams were placed in supercritical carbon dioxide again. The original morphology of the microcellular foam collapsed. The collapsed morphology was also characterized with scanning electron microscopy, and three kinds of cellular morphology were observed. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2009     

Extraction of boric acid from colemanite mineral by supercritical carbon dioxide

The use of H₂SO₄ in boric acid production from colemanite mineral has several problems, related to product impurities, corrosion and environmental discharge limits. To overcome these problems and to increase extraction efficiency of boric acid, heterogeneous reaction between colemanite and CO₂ dissolved in H₂O was studied at and above supercritical CO₂ conditions. Supercritical conditions enhanced the extraction efficiency of boric acid from colemanite mineral, with 96.9% boric acid extraction efficiency being obtained from CO₂–colemanite reaction at 60 °C, for 2 h of reaction time for particles in the range of +20–40 μm. A powder crystallized from filtrate of reaction was determined as H₃BO₃ and the solid formed at the end of reaction was characterized mostly as CaCO₃ according to FTIR, XRD, TGA and SEM analyses. The use of supercritical CO₂ as a leaching agent in colemanite does not only produce boric acid but also helps to reduce the amount of CO₂ in the atmosphere. Based on these facts, supercritical CO₂ as extractant makes this process green and sustainable for recovering boric acid from boron minerals.     

Dyeing poly(lactic acid) fibres in supercritical carbon dioxide

A study has been conducted into the dyeing of poly(lactic acid) fibres in supercritical carbon dioxide. The fibres were completely dyed using disperse dyes at 50 °C as shown by fibre cross-sections, although high colour depths in dark shades still prove challenging. Dye uptake increased significantly at temperatures ≥80 °C. At 95 °C in supercritical carbon dioxide, shrinkage and hardening of raw poly(lactic acid) were observed which could partly be overcome by the supercritical carbon dioxide extraction step. Afterclearing with cold supercritical carbon dioxide (to remove unfixed dye after dyeing) decreased the colour depth and led to non-uniform dyeing results on poly(lactic acid). Wash and rub fastness was good to very good also when poly(lactic acid) was not aftercleared in supercritical carbon dioxide. Fibre damage and elongation at break in supercritical carbon dioxide were similar to water.     

Deacidification of olive oils by supercritical carbon dioxide

An investigation of the application of supercritical carbon dioxide (SC-CO₂) extraction to the deacidification of olive oils has been made to verify that the nutritional properties of the oil remain unchanged when this technique is applied. Preliminary runs at 20 and 30 MPa in the temperature range of 35–60°C were performed on fatty acids and triglycerides as pure compounds or mixtures, to determine their solubility in SC-CO₂. The solubility data obtained show that CO₂ extracts fatty acids more selectively than triglycerides under specific conditions of temperature and pressure (60°C and 20 MPa). It has been noted that the physical state of the solutes plays an important role in determining the solubility trends as a function of temperature and pressure. Extraction of free fatty acids from olive oil was performed on samples with different free fatty acid (FFA) contents at 20 and 30 MPa and at 40 and 60°C. Experimental data suggest that the selectivity factor for fatty acids is higher than 5 and increases significantly as the fatty acid concentration of the oil decreases. For a FFA content of 2.62%, the selectivity reaches a value of 16. In order to evaluate any variations in the composition, several SC-CO₂ extractions of husk oil with high FFA content (29.3%) were made. The results show that selectivity is still significant (≈5) and the composition in the minor component of the deacidified oil has not changed. On the basis of the experimental results and preliminary process evaluations, the authors conclude that SC-CO₂ extraction could be a suitable technique for the deacidification of olive oils, especially for oils with relatively high FFA (<10%).     

Cosolvent-modified supercritical carbon dioxide extraction of phenolic compounds from bamboo leaves (Sasa palmata)

This study highlights the possibility of supercritical carbon dioxide for extracting phenolic compounds from bamboo leaves that have shown antioxidant and anticancer activities. The CO₂ extraction solvent was modified by adding ethanol–water mixture cosolvent of different concentrations to allow extraction of both polar and non-polar compounds. Conventional Soxhlet extraction was also done to investigate the advantages of supercritical extraction over the conventional extraction method. For addition of 5% (mol) of a 25:75 (mol:mol) ethanol–water mixture solvent to CO₂, the highest amount of polyphenols (7.31 ± 0.06 mg/g bamboo leaves in catechin equivalents) and radical scavenging activity (3.65 ± 0.05 mg/g bamboo leaves in BHA equivalents) at 20 MPa and 95 °C, could be obtained among the mixture cosolvents studied. For Soxhlet extraction with a 25:75 (mol:mol) ethanol–water mixture, 1.48 times the amount of phenolic compounds (10.85 ± 0.52 mg/g bamboo leaves in catechin equivalents), could be isolated compared with the supercritical extraction method, however, the radical scavenging activity (3.30 ± 0.05 mg/g bamboo leaves in BHA equivalents) was 0.90 times lower than the extract obtained from the supercritical extraction method. The seven major antioxidative compounds identified from the SC-CO₂ extraction method were: (1) dl-alanine, (2) gluconic acid, (3) phosphoric acid, (4) ß-siosterol, (5) β-amyrene, (6) α-amyrin acetate and (7) friedelin.     

The lipids of krill (Euphausia species) and red crab (Pleuroncodes planipes)

The composition of the lipids of two samples of krill and one of “red crab” was determined by thin layer, column and gas chromatographic procedures. A large number of unusual fatty acids were present.