Free Fatty Acid Fraction Research Articles

Efficient Degumming of Rice Bran Oil by Immobilized PLA1 from Thermomyces lanuginosus.

Phospholipase A1 (PLA1) immobilized in calcium alginate can effectively overcome the mass transfer resistance at the lipid-water interface making more room for the enzyme to separate itself from the products of reaction and to bind with the next available molecule at the interface. The reaction of an immobilized PLA1 hydrolase from Thermomyces lanuginosus was comparatively faster than of its free form. The rate of phospholipid hydrolysis by PLA1 was studied in calcium-rich and calcium-depleted environments; and the extent of phosphorus removed from the crude rice bran oil as well as the amount of free fatty acids produced during the reaction were used as indices for analysing the rate of enzymatic hydrolysis under standard conditions of pH, temperature, time of incubation and agitation. The immobilized PLA1 was found to be superior in removing phosphorus in the presence of 10 mM bivalent calcium ions in a solution. As compared to a maximum of 72.52% phosphorus removed by 0.01 kg of free enzyme per kg of oil, the same amount of immobilized PLA1 removed phosphorus from oil by 94.12% under the same experimental conditions (pH=6, 60 °C, 1-hour incubation). Both the free PLA1 and its immobilized form had shown extended rates of hydrolysis in a calcium-rich environment. The mass fractions of free fatty acids produced by the free enzyme and by its immobilized form were 14.9 and 14.16%, respectively, under the above experimental conditions. The removal of phosphorus from oil was accompanied by a significant reduction in colour and restoration of iodine value to the desired level.

Free fatty acid availability is closely related to myocardial lipid storage and cardiac function in hypoglycemia counterregulation.

Hypoglycemia, a major side effect of intensive glucose-lowering therapy, was recently linked to increased cardiovascular risk in patients with diabetes. Whether increased circulating free fatty acids (FFA) owing to catecholamine-induced lipolysis affect myocardial energy metabolism and thus link hypoglycemia to cardiac vulnerability is unclear. Therefore, this study investigated the impact of hypoglycemia counterregulation (± inhibition of lipolysis) on myocardial lipid content (MYCL) and left ventricular function in healthy subjects. Nine healthy men were studied in randomized order: 1) insulin/hypoglycemia test (IHT; ins+/aci-), 2) IHT during inhibition of adipose tissue lipolysis by acipimox (ins+/aci+), 3) normoglycemia with acipimox (ins-/aci+), and 4) normoglycemia with placebo (ins-/aci-). MYCL and cardiac function were assessed by employing magnetic resonance spectroscopy/imaging at baseline and at 2 and 6 h. In response to acute hypoglycemia, plasma FFA (P<0.0001) and ejection fraction (EF; from 63.2±5.5 to 69.6±6.3%, P=0.0001) increased significantly and were tightly correlated with each other (r=0.68, P=0.0002); this response was completely blunted by inhibition of adipose tissue lipolysis. In the presence of normoglycemia, inhibition of lipolysis was associated with a drop in EF (from 59.2±5.5 to 53.9±6.9%,P=0.005) and a significant decrease in plasma FFA, triglycerides, and MYCL (by 48.5%, P=0.0001). The present data indicate that an intact interorgan cross-talk between adipose tissue and the heart is a prerequisite for catecholamine-mediated myocardial contractility and preservation of myocardial lipid stores in response to acute hypoglycemia.

Synthesis of trans-10,cis-12 conjugated linoleic acid-enriched triacylglycerols via two-step lipase-catalyzed esterification

This study aimed to synthesize trans-10,cis-12 conjugated linoleic acid (t10,c12-CLA)-enriched triacylglycerols (TAGs) with potential anti-obesity effects via a two-step lipase-catalyzed reaction. Commercial CLA isomer mixtures, containing 33.3 g/100 g t10,c12-CLA, were esterified with dodecan-1-ol to selectively enrich the t10,c12-CLA in a free fatty acid (FFA) fraction. The reaction was performed in a recirculating packed bed reactor using Candida rugosa lipase (immobilized on Immobead 150) as the biocatalyst. An FFA fraction containing 54.7 g/100 g t10,c12-CLA was produced in a yield of 21.8 g/100 g initial t10,c12-CLA under the optimal conditions, i.e., temperature, 20 °C; CLA mixtures-to-dodecan-1-ol molar ratio, 1:1; water content, zero (no added water); reaction time, 36 h. A t10,c12-CLA-enriched FFA fraction was esterified with glycerol to prepare t10,c12-CLA-enriched TAGs. The reaction was performed in a stirred batch reactor using Candida antarctica lipase B (immobilized on macroporous acrylic resin) as the biocatalyst. The optimal combination of temperature, glycerol-to-FFA fraction molar ratio, enzyme loading, reaction time, and vacuum level was 60 °C, 1:3, 10 g/100 g (based on total substrates), 12 h, and 0.4 kPa, respectively. Under these conditions, the TAG content reached 93.7 g/100 g and t10,c12-CLA was evenly distributed throughout the glycerol backbone of the TAG.

About the origin of asclepic acid derived from crude homo- and heterolipids during successive solvent extraction of rapeseeds

To improve oil yield at the final stage of process, relation among solvent extraction mechanism and the removal of homo- and heterolipid classes with different acyl chain domains was studied. Selected technological parameter was decreasing quality of crude oil in the last successively extracted fractions. The composition of fatty acids, originally linked as lipid building-blocks, indicated no differences up to 94 wt% of hexane extractable oil from the rapeseed material. During the same time interval, it was released 95.8 cumulative wt% of oleic acid and only 76.9 cumulative wt% of (Z)-11-octadecenoic (asclepic, cis-vaccenic) acid. Steep decrease of oleic acid content was correlated with the down-trend of total monounsaturated fatty acid concentrations in the simple glycerolipid fractions (from 57.0 –59.0 to 26.7 wt%) and free fatty acid fractions (from 59.0–60.0 to 40.1 wt%). On the other hand, we have observed 7.14-fold increase of asclepic acid concentration derived from the simple glycerolipids (mono-, di- and triacylglycerols), wax and sterol esters. The increment of asclepic acid was 4.78-fold in the fraction of the complex glycerolipids and acylate steryl glycosides. Lipidomic and proteomic analyses of isolated oil bodies, rapeseed hull and dehulled seeds proved an origin of (Z)-11-octadecenoic acid. Furthermore, we determined similar fatty acid composition of lipid fractions derived from pure oil bodies and from dehulled seeds. In both cases, the low content of total asclepic acid: 3.13–3.15 wt% in aleurone cells was observed. Low amount of (Z)-11-octadecenoic acid is novel finding, which indicated that (Z)-11-octadecenoic acid originated from seed coat species that are less accessible to the hexane. It was confirmed that the main portion of asclepic acid moieties were derived from wax and sterol esters, triacylglycerols and heterolipids originally present in rapeseed coat (epidermal layer, testa and aleurone layer). Therefore, increasing amounts of asclepic acid and its precursor, palmitoleic acid, released from seed coats was significant indicator of solvent extraction process efficiency.

Fate of palmitic, palmitoleic and eicosapentaenoic acids during anaerobic digestion of Phaeodactylum tricornutum at varying lipid concentration

The literature is showing an increasing trend in the use of microalgal biomass for the production of biofuel, with a recent keen interest for the conversion of microalgae into methane. The methane potential from Phaeodactylum tricornutum was similar for the whole algae, 362±5L/kg total volatile solid (TVS), compared with a sample from which the lipids were extracted, at 358±6L/kg TVS. The anaerobic digestion of long chain fatty acids (LCFAs) representative of the dominant acids found in P. tricornutum during 22weeks of incubation demonstrated that the usual incubation time of typical anaerobic digesters (3–6weeks) is insufficient to hydrolyze the lipids and maximize the energy yield contained in the microalgae. The results from methane potential assays targeting the LCFAs that were found in significant concentrations in P. tricornutum, namely palmitic, palmitoleic and eicosapentaenoic (EPA) acids), at concentration of 2.5 and 1.25g/L, showed a slow but high yield with palmitic acid (802±16L/kg), inhibition from palmitoleic acid and fair degradation of EPA (442±67L/kg). There was a 27% increase in the methane yield from a reconstituted P. tricornutum containing 25% of LCFA compared to 10%, at 456±1 vs 363±5mLSTP/g TVS. Increasing the free fatty acid fraction to 50% in the microalgal biomass resulted in the inhibition of the digestion after production of methane during the first 11days of incubation.

Overexpression of a phosphatidic acid phosphatase type 2 leads to an increase in triacylglycerol production in oleaginous Rhodococcus strains.

Oleaginous Rhodococcus strains are able to accumulate large amounts of triacylglycerol (TAG). Phosphatidic acid phosphatase (PAP) enzyme catalyzes the dephosphorylation of phosphatidic acid (PA) to yield diacylglycerol (DAG), a key precursor for TAG biosynthesis. Studies to establish its role in lipid metabolism have been mainly focused in eukaryotes but not in bacteria. In this work, we identified and characterized a putative PAP type 2 (PAP2) encoded by the ro00075 gene in Rhodococcus jostii RHA1. Heterologous expression of ro00075 in Escherichia coli resulted in a fourfold increase in PAP activity and twofold in DAG content. The conditional deletion of ro00075 in RHA1 led to a decrease in the content of DAG and TAG, whereas its overexpression in both RHA1 and Rhodococcus opacus PD630 promoted an increase up to 10 to 15% by cellular dry weight in TAG content. On the other hand, expression of ro00075 in the non-oleaginous strain Rhodococcus fascians F7 promoted an increase in total fatty acid content up to 7% at the expense of free fatty acid (FFA), DAG, and TAG fractions. Moreover, co-expression of ro00075/atf2 genes resulted in a fourfold increase in total fatty acid content by a further increase of the FFA and TAG fractions. The results of this study suggest that ro00075 encodes for a PAP2 enzyme actively involved in TAG biosynthesis. Overexpression of this gene, as single one or with an atf gene, provides an alternative approach to increase the biosynthesis and accumulation of bacterial oils as a potential source of raw material for biofuel production.

Nonpolar Compounds and Free Fatty Acids from Several Marine Isolates of Fungus Aspergillus ustus and Actinobacterium Nocardiopsis umidischolae

In continuation of research on secondary metabolites of marine fungi and actinobacteria [1, 2], hexane fractions and fractions of free fatty acids and the sterol fraction of actinobacterium were obtained and analyzed from cultures of marine isolates of the fungus Aspergillus ustus KMM 4642 and KMM 4664 isolated from sediment (Okhotsk Shelf, Sakhalin Island, 26.5 m depth) and the actinobacterium Nocardiopsis umidischolae KMM 7036 isolated from Mycale sp. of sponge (Deryugin Basin, Okhotsk Sea). Strain KMM 4642 was cultivated in malt-agar medium prepared with sea water for 14 d [3] and on rice medium prepared with seawater for 21 d [4]. Strain KMM 4664 was cultivated only on rice medium prepared with seawater. Actinobacterium was cultivated in a special medium containing peptone (5.0 g/L), meat extract (3.0), starch (20.0), agar (16.0) and seawater at pH 7.5 and 23°C for 23 d. Cultures were extracted with EtOAc. The extracts were evaporated to dryness. The resulting residues were dissolved in EtOH (10%) and extracted successively with hexane, EtOAc, and BuOH. The hexane fractions were evaporated at reduced pressure and analyzed by GC-MS. The data were compared with the mass spectrometric fragmentation of standards using the NIST98 database. The hexane fraction of A. ustus KMM 4642 consisted of 96.88% diethylhexylphthalate. The remainder consisted of fatty acid ethyl esters and pentadecane. The hexane fraction of A. ustus KMM 4664 contained fatty acid ethyl esters, squalene, octadecane, and sterols such as 24-cholest-4-en-3-one, 24-ethylcholest-7-en-3 -ol, 24-ethylcholesta-3,5-diene, and 24-ethylcholest-5-en-3 -ol at concentrations of 1–2% of the studied mixture. These results differed from those obtained earlier for marine isolates of A. ustus [1]. The hexane fraction of actinobacterium N. umidischolae contained hydrocarbons at concentrations of 1.5–3% that included linear C15, C16, C17, C18, C20, and C22; C18, C19, C20, C22, and C24 with an iso-carbon chain; C18, C19, and C22 with a terminal double bond; and dibutylphthalate (5%) and diethylhexylphthalate (45%). The EtOAc fraction of each culture was chromatographed over a column of silica gel using a hexane–EtOAc gradient (100:0 90:10) to afford fractions of free fatty acids. The obtained total acids were analyzed as the methyl esters (methylated by diazomethane in Et2O) and pyrrolides [5] using GC-MS. Derivatives were identified by comparing their mass spectra with those of standards using the NIST98 database. The composition and content of A. ustus acid esters are given below (mass%):

Acinar inflammatory response to lipid derivatives generated in necrotic fat during acute pancreatitis

Lipids play a role in acute pancreatitis (AP) progression. We investigate the ability of pancreatic acinar cells to trigger inflammatory response in the presence of lipid compounds generated in necrotic areas of peripancreatic adipose tissue (AT) during AP induced in rats by 5% sodium taurocholate. Lipid composition of AT was analyzed by HPLC–mass spectrometry. Acinar inflammatory response to total lipids as well as to either the free fatty acid (FFA) fraction or their chlorinated products (Cl-FFAs) was evaluated. For this, mRNA expression of chemokine (C-C motif) ligand 2 (CCL2) and P-selectin as well as the activation of MAPKs, NF-κB and STAT-3 were analyzed in pancreatic acini. Myeloperoxidase (MPO) activity, as an inducer of Cl-FFA generation, was also analyzed in AT. MPO activity significantly increased in necrotic (AT-N) induced changes in lipid composition of necrotic fat, such as increase in FFA and phospholipid (PL) content, generation of Cl-FFAs and increases in saturated FFAs and in the poly-:mono-unsaturated FFA ratio. Total lipids from AT-N induced overexpression of CCL2 and P-selectin in pancreatic acini as well as MAPKs phosphorylation and activation of NF-κB and STAT3. FFAs, but not Cl-FFAs, up-regulated CCL2 and P-selectin in acinar cells. We conclude that FFAs are capable of up-regulating inflammatory mediators in pancreatic acini and given that they are highly produced during AP, mainly may contribute to the inflammatory response triggered in acinar cells by fat necrosis. No role is played by Cl-FFAs generated as a result of neutrophil infiltration.

Obtaining a hydrolyzed milk fat fraction enriched in conjugated linoleic acid and trans-vaccenic acid

Anhydrous milk fat (AMF) rich in conjugated linoleic acid (CLA) and trans-vaccenic acid (TVA) was enzymatically hydrolyzed and dry fractionated. Oxidation kinetic parameters of hydrolyzed AMF was determined by differential scanning calorimetry. Hydrolysis yielded 90.4% free fatty acids (FFAs) after 24 h at 50 °C, using a ratio of 70 for water to fat (w/w) and a ratio of 40 for enzyme to fat (w/w). The oxidation of hydrolyzed AMF started at a lower temperature (108.3 ± 4.1 °C) compared with non-hydrolyzed AMF (155.7 ± 3.4 °C). Upon fractionation of hydrolyzed AMF, three fractions were obtained (high-, middle- and low-melting point FFA fractions). The middle fraction contained the highest concentration of CLA and TVA (64.8 and 249.3 mg g−1 fat, respectively). The middle fraction shows great potential for fortification of dairy products but further optimization is needed to maximize the CLA and TVA contents, while minimizing the presence of the other fatty acids.

Gamma Irradiation Prolongs the Sea Bass (Dicentrarchus Labrax L.) Storage and Delays the Lipids Membrane Degradation

Mediterranean sea bass (Dicentrarchus labrax L.) fish were gamma irradiated at doses 0, 0.5, 1, 2 or 3 kGy and stored at 1°C for 21 days in the dark to assess whether the delay of fish alteration by irradiation involved the protection of membrane lipids. Total basic volatile nitrogen (TVB-N) and the trimethylamine (TMA) contents, the pH, and the lipid composition of membrane were determined in the muscle tissue during storage. The TVB-N and TMA contents and the pH of irradiated and non-irradiated samples increased during storage whereas the external quality decreased in correlation with a reduction in the amount of phospholipids (PL). The degree of unsaturation of PL and of free fatty acids (FFA) fractions decreased, whereas, the ratio of sterol to PL increased. The catabolism of PL was delayed by irradiation during storage as compared to the untreated sample and it was always positively correlated with the total viable counts (TVC) of fish muscle which was also reduced by the treatments. We conclude that the delay of alteration of sea bass tissue during storage by irradiation involved probably a protection of membrane lipids from degradation which seemed to be partially reliable to the micro-organisms load of muscle tissue.

Optimization of Lipase‐Catalyzed Fractionation of Two Conjugated Linoleic Acid (CLA) Isomers

AbstractThe separation of two isomers of conjugated linoleic acid is highly significant since each exhibits different biochemical properties. The aim of this study was to investigate and optimize several factors affecting the esterification of l‐menthol with the c9,t11‐CLA isomer in an organic solvent‐free system using lipase from Candida rugosa (Lipase AY‐30). D‐optimal design with 5 factors and 3 levels were employed to evaluate the effects of synthesis parameters; reaction time (8–24 h), temperature (30–50 °C), enzyme content (2–20 U/ml), substrate molar ratio of conjugated linoleic acid oil to l‐menthol (2:1–1:2) and pH (6–8) on esterification of c9,t11‐CLA with l‐menthol. Based on the analysis of the residual amount of c9,t11‐CLA in the free fatty acid fraction after just one‐step esterification, the optimum synthesis conditions were as follows: reaction time 23.12 h, temperature 32.65 °C, enzyme amount 135.40 U, molar ratio of CLA oil to l‐menthol at 1:1.7 and pH at 7.7; the lowest purity of c9,t11‐CLA in free fatty acid fraction based on the total content of c9,t11 and t10,c12‐CLA isomers was 8.6 %.

Determination of free fatty acids in beer wort

The importance of free fatty acids (FFAs) in wort has been known for a long time because of their influence on beer quality and yeast metabolism. Lipids have a beneficial effect on yeast growth during fermentation as well as negative effects on beer quality. Lipids content of beer affects the ability to form a stable head of foam and plays an important role in beer staling. Moreover, the ratio of unsaturated and saturated fatty acids seems to be related to gushing problems. A novel, simple, and reliable procedure for quantitative analysis of FFAs in wort was developed and validated. The determination of FFAs in wort was achieved via liquid–liquid cartridge extraction, purification of FFA fraction by solid phase extraction, boron trifluoride in methanol methylation, and injection into GC-FID system. The proposed method has high accuracy (<0.3%, expressed as the bias), high precision (<1.2%, RSD), and recoveries ranging from 74% to 98%. The method was tested on two different wort samples (9° and 12° Plato).

Fatty Acids and Recovery During First Hours of Acute Myocardial Ischemia

Figure 1. Data for FFA from the study by Oliver et al. Data for creatine kinase-MB (CK-MB) and troponins from “Wikipedia.” Maintenance or restoration of normal myocardial metabolism during acute ischemia has seldom been a priority in its management. The focus for many years has been on overcoming impaired coronary blood supply. Depending on the severity, this may or may not restore abnormal myocardial metabolism. Yet, it is a functioning myocardium that will determine survival and prognosis. Normal myocardial metabolism depends on a balanced supply of fatty acids, glucose, pyruvate, and lactate—the principal substrates. An excess of free fatty acids (FFAs) to local ischemic areas could uncouple oxidative metabolism with irreparable damage and lethal arrhythmias. The fear and pain, which usually occurs during impending infarction, is accompanied by a surge of adrenaline leading to lipolysis of stored tissue glycerides and a large and immediate efflux of FFAs throwing the balance of substrates awry. Studies conducted during the first hours after the onset of symptoms indicate that there is an excess of ventricular arrhythmias and mortality, both of which have been correlated with high concentrations of plasma FFA. In this issue, Kleinfeld et al, using data from the Thrombolysis In Myocardial Infarction II trial of thrombolysis for ST elevation myocardial infarction, have shown that this is also true for unbound FFA (FFAu). FFAu is a very small fraction of total FFA. It increases exponentially with increasing total albumin-bound FFA and is more sensitive to physiological changes. In 1,834 patients, the upper 2 quartiles of FFAu were associated with a significantly higher mortality rate at 1, 7, and 30 days after acute myocardial infarction. In this study, high FFAu was a risk factor independent of age, gender, race, body mass index, diastolic and systolic blood pressures at baseline, history of diabetes, previous myocardial infarction and hypertension, and use of b blockers within 24 hours of enrollment. This confirms earlier reports of the association of ventricular arrhythmias with high plasma concentrations of FFA. Proof that it is the high concentrations of FFA that impair myocardial metabolism has previously been demonstrated in dogs with experimental myocardial ischemia, which were given heparin and protamine sulfate. Heparin induced plasma lipolysis leading to high concentrations of plasma FFA and ventricular arrhythmias. Both could be decreased by protamine sulfate. Also, infusions of

Polyunsaturated fatty acid metabolism in enterocyte models: T84 cell line vs. Caco-2 cell line

Human colon carcinoma cell lines such as Caco-2 cells, model of mature enterocytes and T84 cells, model of crypt cells are useful to study interactions between nutrient processing and metabolic functions at intestinal level. Our study aimed at comparing the ability of Caco-2 and T84 cells (1) to incorporate dietary polyunsaturated fatty acids (PUFA), (2) to process them and (3) to sort them into neutral lipids (NL), free fatty acids (FFA) and phospholipids (PL). Caco-2 and T84 cells were exposed to a 7-day long supplementation with PUFA. The amounts of fatty acids accumulated and incorporated into the NL, FFA or PL fractions were higher in Caco-2 than in T84 cells. Caco-2 cells were able to significantly elongate C18 PUFA and C20 PUFA of both n-3 and n-6 families. In contrast, T84 cells were unable to elongate the n-6 fatty acids whereas elongation of n-3 fatty acids was detectable but marginal. Similarly, a Δ6 desaturase activity was observed in Caco-2 but not in T84 cells. In T84 cells, each exogenous fatty acid was predominantly accumulated in the PL fraction. In Caco-2 cells, C20 fatty acids and C18:2n-6 was preferentially accumulated in the PL fraction, while C22 PUFA and C18:3n-3 was preferentially accumulated in the NL fraction. Overall, this study has shown that Caco-2 and T84 cells, as models of intestinal mucosal cells, present large differences in PUFA accumulation capacity, specific elongase and desaturase activities and distribution pattern of exogenous PUFA and of their metabolites in the lipid classes.

Recent Patents on Lipase Catalyzed Vegetable and Fish Oil Modification

Lipase is an enzyme which catalyzes hydrolysis, esterification and transesterification processes. Type (origin) of lipase affects process conditions and final conversion. Use of these biocatalytic properties of lipase in modification of various vegetable oils is depicted in different patents, leading to varieties of beneficial products. Production and concentration of polyunsaturated fatty acids like eicosapentanoic acid, docosapentanoic acid and docosahexanoic acid were carried out through lipase catalyzed hydrolysis. Hydrolysis followed by reesterification was also employed in some patents to concentrate long chain polyunsaturated fatty acids in triacylglycerol fraction or free fatty acid fraction as per specific requirements. Ethanolysis of fish oil hexyl esters was also described in some studies to concentrate similar fatty acids. Combined ethanolysis and esterification, ethanolysis and transesterification were also conducted in some patents to concentrate these desirable fatty acids. Degumming using phospholipase and biodiesel production with lipase as catalyst were also carried out leading to some notable patents. Reactor configuration for maximizing conversion was also patented. Removal of diacylglycerols from their mixture with triacylglycerols led to specifically designed triacylglycerol mixture for catering certain purposes.

Effects of high dose intravenous fish oil on human atrial electrophysiology: Implications for possible anti- and pro-arrhythmic mechanisms in atrial fibrillation

Intravenous omega-3 polyunsaturated fatty acids (ω-3 PUFAs) may prevent atrial fibrillation (AF) inducibility and perpetuation in animal models. We examined the effect of high dose IV ω-3 PUFAs on human atrial electrophysiology. We randomised 88 patients with no structural heart disease to receive saline (control group) or high dose IV ω-3 PUFA infusion prior to detailed atrial electrophysiologic evaluation. Biologically active components, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were measured in total lipids, free fatty acid and phospholipid (membrane incorporated) fraction pre and post infusion. Compared to pre-infusion values, EPA and DHA increased significantly in the total lipids and free fatty acid but were unchanged in the phospholipid fraction. IV ω-3 did not alter atrial refractory periods, however it slowed right, left and global atrial conduction (P<.05). Inducible AF was significantly less likely in ω-3 patients compared to controls (AF ≥ 5 min, 20% vs. 58%, P = .02) and was non-sustained (mean AF duration: 14s vs. 39 s, P<.001), however inducible and sustained atrial flutter was more common (≥ 5 min: 28% vs. 0%, P = .01). Organisation of AF into flutter was observed in a greater proportion of inductions in the ω-3 group (8.5% vs. 0.6%, P<.001). IV ω-3 PUFAs (as free fatty acids) cause acute atrial conduction slowing, suppress AF inducibility, organise AF into atrial flutter and enhance atrial flutter inducibility. These findings provide a novel insight into potential anti and pro-arrhythmic mechanisms of fish oils in human AF.

Strain variability in fatty acid composition of Chattonella marina (Raphidophyceae) and its relation to differing ichthyotoxicity toward rainbow trout gill cells.

Lipid profiles of three strains (Mexico, Australia, Japan) of Chattonella marina (Subrahmanyan) Hara et Chihara were studied under defined growth (phosphate, light, and growth phase) and harvest (intact and ruptured cells) conditions. Triacylglycerol levels were always <2%, sterols <7%, free fatty acids varied between 2 and 33%, and polar lipids were the most abundant lipid class (>51% of total lipids). The major fatty acids in C.marina were palmitic (16:0), eicosapentaenoic (EPA, 20:5ω3), octadecatetraenoic (18:4ω3), myristic (14:0), and palmitoleic (16:1ω7c) acids. Higher levels of EPA were found in ruptured cells (21.4-29.4%) compared to intact cells (8.5-25.3%). In general, Japanese N-118 C.marina was the highest producer of EPA (14.3-29.4%), and Mexican CMCV-1 the lowest producer (7.9-27.1%). Algal cultures, free fatty acids from C.marina, and the two aldehydes 2E,4E-decadienal and 2E,4E-heptadienal (suspected fatty acid-derived products) were tested against the rainbow trout fish gill cell line RTgill-W1. The configuration of fatty acids plays an important role in ichthyotoxicity. Free fatty acid fractions, obtained by base saponification of total lipids from C.marina showed a potent toxicity toward gill cells (median lethal concentration, LC50 (at 1h) of 0.44μg·mL(-1) in light conditions, with a complete loss of viability at >3.2μg·mL(-1) ). Live cultures of Mexican C.marina were less toxic than Japanese and Australian strains. This difference could be related to differing EPA content, superoxide anion production, and cell fragility. The aldehydes 2E,4E-decadienal and 2E,4E-heptadienal also showed high impact on gill cell viability, with LC50 (at 1h) of 0.34 and 0.36μg·mL(-1) , respectively. Superoxide anion production was highest in Australian strain CMPL01, followed by Japanese N-118 and Mexican CMCV-1 strains. Ruptured cells showed higher production of superoxide anion compared to intact cells (e.g., 19 vs. 9.5pmol·cell(-1) ·hr(-1) for CMPL01, respectively). Our results indicate that C.marina is more ichthyotoxic after cell disruption and when switching from dark to light conditions, possibly associated with a higher production of superoxide anion and EPA, which may be quickly oxidized to produce more toxic derivates, such as aldehydes.

Effects of Echinostoma caproni miracidia dose on the neutral and polar lipids of Biomphalaria glabrata as determined by high-performance thin-layer chromatography

The effects of a 5 versus 25 miracidia exposure of Echinostoma caproni on the lipid composition of Biomphalaria glabrata was studied using high performance thin layer chromatography (HPTLC)-densitometry. A 50 miracidia dose was not used because such a high level of exposure caused severe snail mortality by 3 weeks post-exposure (PE). Lipids were determined in the digestive-gland gonad complex (DGG) of the exposed snails and in the uninfected matched controls at 2 and 4 weeks PE. Extraction of lipids from DGGs was carried out by the Folch method with chloroform-methanol (2:1), and extracts were analyzed on Analtech HPTLC-HLF pre-adsorbent silica gel plates with measurement of separated bands using a CAMAG Scanner 3. For neutral lipids the mobile phase was petroleum ether-diethyl ether-glacial acetic acid (80:20:1) and the detection reagent was 5% ethanolic phosphoric acid, and for polar lipids chloroform-methanol-deionized water (65:25:4) mobile phase and 10% cupric sulfate in 8% phosphoric acid detection reagent were used. No significant differences in the concentrations of free sterols, free fatty acids, triacylglycerols, phosphatidylcholine, and phosphatidylethanolamine were seen at 2 weeks PE in any of the groups. At 4 weeks PE, the free fatty acid concentration increased significantly in the snails exposed to 25 miracidia compared to that of the 5 miracidia/snail group or the controls. Elevation of the free fatty acid fraction in the high dose snail group suggested that some changes occurred in the lipid metabolism of the snails in that group as a function of miracidia dose.

Factors affecting cellular lipid extraction from marine microalgae

Factors affecting intracellular lipid extraction from marine microalgae were investigated using various techniques. The biomass drying method and moisture content, and the solvent extraction system were the factors studied. Lipid was analytically classified into three categories i.e. neutral lipid, free fatty acid (FFA) and polar lipid using solid-phase extraction. Biomass drying methods (freeze-, oven- and solar drying) did not affect lipid yield, but the FFA content of the lipid was three times higher for solar dried biomass. Of the various lipid extraction methods tested, including sonication, homogenization, accelerated solvent extraction (ASE) and Soxhlet extraction, sonication was the least efficient compared to other methods when a partially miscible solvent system i.e. hexane–methanol was used. Chloroform–methanol solvent system had maximum lipid extraction efficiency (33%). A biomass moisture content up to 5% had no impact on lipid extraction efficiency, but higher moisture contents reduced lipid extraction and increased the FFA fraction.

Reduced protein oxidation in Wistar rats supplemented with marine ω3 PUFAs

The potential effects of various dietary eicosapentaenoic acid (EPA; 20:5) and docosahexaenoic acid (DHA; 22:6) ratios (1:1, 2:1, and 1:2, respectively) on protein redox states from plasma, kidney, skeletal muscle, and liver were investigated in Wistar rats. Dietary fish oil groups were compared with animals fed soybean and linseed oils, vegetable oils enriched in ω6 linoleic acid (LA; 18:2) and ω3 α-linolenic acid (ALA; 18:3), respectively. Fish oil treatments were effective at reducing the level of total fatty acids in plasma and enriching the plasmatic free fatty acid fraction and erythrocyte membranes in EPA and DHA. A proteomic approach consisting of fluorescein 5-thiosemicarbazide (FTSC) labeling of protein carbonyls, FTSC intensity visualization on 1-DE or 2-DE gels, and protein identification by MS/MS was used for the protein oxidation assessment. Albumin was found to be the most carbonylated protein in plasma for all dietary groups, and its oxidation level was significantly modulated by dietary interventions. Supplementation with an equal EPA:DHA ratio (1:1) showed the lowest oxidation score for plasma albumin, followed in increasing order of carbonylation by 1:2 <2:1 ≈ linseed < soybean. Oxidation patterns of myofibrillar skeletal muscle proteins and cytosolic proteins from kidney and liver also indicated a protective effect on proteins for the fish oil treatments, the 1:1 ratio exhibiting the lowest protein oxidation scores. The effect of fish oil treatments at reducing carbonylation on specific proteins from plasma (albumin), skeletal muscle (actin), and liver (albumin, argininosuccinate synthetase, 3-α-hydroxysteroid dehydrogenase) was remarkable. This investigation highlights the efficiency of dietary fish oil at reducing in vivo oxidative damage of proteins compared to oils enriched in the 18-carbon polyunsaturated fatty acids ω3 ALA and ω6 LA, and such antioxidant activity may differ among different fish oil sources because of variations in EPA/DHA content.