Neutral Lipid Fraction Research Articles

Separation of polar and neutral lipids from mammalian cell lines by high-performance thin-layer chromatography.

The introduction of high-performance TLC (HPTLC) instrumentation that allows precise control of critical parameters has transformed the technique into an efficient and rapid tool for analyzing various metabolites, namely lipids. Although mass spectrometry (MS) has largely replaced lipid analysis techniques over recent decades due to its comprehensive lipidome profiling capabilities, it typically lacks the rapidity and simplicity of TLC. HPTLC remains advantageous due to its ease of use, simpler data interpretation, and compatibility with complementary techniques. In this study, we established a HPTLC protocol to fractionate both polar and non-polar lipids on a single normal phase plate. Twenty lipid standards were fractionated and the method was successfully applied to whole extracts from six mammalian cell lines. Standards and extracted lipids were applied with an automated sampler, and polar lipids were first fractionated in a 5-step automated gradient elution, followed by the fractionation of neutral lipids in a twin-trough chamber with three different elutions. Plates were automatically sprayed with a modified copper sulfate solution and charred to reveal lipids and obtain the respective chromatograms. LC-MS was used to identify ambiguous bands, thus ensuring the accuracy of lipid identification.

Metal accumulation in female green sea turtles (Chelonia mydas) from Eastern Atlantic affects their egg quality with potential implications for embryonic development

Sea turtles, with their global distribution and complex life cycle, often accumulate pollutants such as metals and metalloids due to their extended lifespan and feeding habits. However, there are limited studies exploring the impact of metal pollution on the reproductive health of female sea turtles, specifically focusing on the quality of their eggs, which has significant implications for the future generations of these charismatic animals. São Tomé Island, a crucial nesting and feeding habitat for green sea turtles, underscores the urgent need for comprehensive research in this ecologically significant area. This study aimed to investigate whether metals and metalloids in the blood of nesting female green sea turtles induce genotoxic effects in their erythrocytes and affect their egg morphometric characteristics and the composition of related compartments. Additionally, this study aimed to evaluate whether the quality of energetic reserves for embryo development (fatty acids in yolk's polar and neutral lipids) is influenced by the contamination status of their predecessors. Results revealed correlations between Cu and Hg levels and increased “lobed” erythrocytes, while As and Cu negatively influenced shell thickness. In terms of energy reserves, both polar and neutral lipid fractions contained primarily saturated and monounsaturated fatty acids, with prevalent 18:1n-9, 18:0, 16:0, 14:0, and 12:0 fatty acids in yolk samples. The yolk polar fraction was more susceptible to contaminant levels in female sea turtles, showing consistent negative correlations between pollution load index and essential n3 fatty acids, including linolenic, eicosatrienoic, eicosapentaenoic, and docosapentaenoic acids, crucial for embryonic development. These metals accumulation, coupled with the reduced availability of these key fatty acids, may disrupt the eicosanoid and other important pathways, affecting reproductive development. This study reveals a negative correlation between metal contamination in female sea turtles' blood and egg lipid reserves, raising concerns about embryonic development and the species' future generations.

Bioactive lipids derived from red wine, beers, and their dealcoholized variants inhibit platelet-activating factor (PAF) induced platelet activation in vitro

This study assesses the in vitro antiplatelet properties of polar lipid extracts of two popular beers, a red wine and their 0% equivalents against platelet aggregation. Total lipids (TL) were isolated using the Bligh and Dyer method from each beverage and the total polar lipid (TPL) and total neutral lipid (TNL) fractions were further isolated using counter-current distribution. The lipid profile of each TPL fraction was analysed using LC-MS. Platelet aggregometry was performed to evaluate the TL, TNL, and TPL extracts of each beverage against platelet-activating factor (PAF) induced platelet aggregation. TPL and TL lipid fractions significantly inhibit platelet aggregation, with TPL fractions showing the highest inhibition as evidenced by generally low IC50 values, ranging from 9 μg to 553 μg. These results indicate that antiplatelet lipid microconstituents are present in beer, wine, and their 0% alcohol counterparts, in low but bioactive amounts. These lipids are characterized by predominance of saturated fatty acids (SFA) and a relatively high ratio of n-6 to n-3 polyunsaturated fatty acids (PUFA) in the TPL composition. The extracts also contained notable levels of phenolic compounds ranging from 0.05 to 0.31 mg/mL. These data provide evidence for the presence of antiplatelet lipid microconstituents that may contribute to the observed epidemiological cardioprotective effects of moderate beer and wine consumption. Furthermore, 0% alcoholic beverages may also provide comparable antiplatelet cardioprotective microconstituents devoid of the risk of ethanol ingestion.

Characterization of Neutral Lipids of the Oleaginous Alga Micractinum inermum.

An oleaginous microalga Micractinum inermum isolated from Mariana Lake, AB, Canada was cultured in a 1000 L photobioreactor with an f/2 medium to study its lipid content and neutral lipid profile. Algal biomass was collected at the stationary phase contained a significant amount of lipids (44.2%), as determined by Folch's method. The lipid was fractionated into neutral lipid, glycolipid and phospholipid fractions. The neutral lipid constitutes almost 77.3% of the total lipid species and is mainly composed of triacylglycerols (TAGs) determined by a proton NMR study. UHPLC-HRMS analysis allows us for the first time to identify 81 TAGs in the neutral lipid fraction of M. inermum. The fatty acid acyl side chains were identified based on fragment ions observed in MSMS analysis. TAGs with fatty acid acyl chains 18:1/18:1/18:1, 18:1/18:1/16:0, 18:2/18:1/16:0, and 18:2/18:2/18:0 were the major ones among the identified TAGs. Fatty acid analysis further supports the fact that oleic acid was the major fatty acid present in the neutral lipid fraction of M. inermum constituting 41.7%, followed by linoleic acid at 21.5%, and palmitic acid at 21.2%. The saturated and monounsaturated fatty acids were 67.8% or higher in the lipid fraction. Long-chain fatty acids were only present in a minor quantity. The results clearly demonstrate that M. inermum is an excellent source for TAGs.

Metabolomics and lipidomics reveal the effects of the toxic dinoflagellate Alexandrium catenella on immune cells of the blue mussel, Mytilus edulis

The increasing occurrence of harmful algal blooms, mostly of the dinoflagellate Alexandrium catenella in Canada, profoundly disrupts mussel aquaculture. These filter-feeding shellfish feed on A. catenella and accumulate paralytic shellfish toxins, such as saxitoxin, in tissues, making them unsafe for human consumption. Algal toxins also have detrimental effects upon several physiological functions in mussels, but particularly on the activity of hemocytes – the mussel immune cells. The objective of this work was to determine the effects of experimental exposure to A. catenella upon hemocyte metabolism and activity in the blue mussel, Mytilus edulis. To do so, mussels were exposed to cultures of the toxic dinoflagellate A. catenella for 120 h. The resulting mussel saxitoxin load had measurable effects upon survival of hemocytes and induced a stress response measured as increased ROS production. The neutral lipid fraction of mussel hemocytes decreased two-fold, suggesting a differential use of lipids. Metabolomic 1H nuclear magnetic resonance (NMR) analysis showed that A. catenella modified the energy metabolism of hemocytes as well as hemocyte osmolyte composition. The modified energy metabolism was reenforced by contrasting plasma metabolomes between control and exposed mussels, suggesting that the blue mussel may reduce feed assimilation when exposed to A. catenella.

Beyond PLFA: Concurrent extraction of neutral and glycolipid fatty acids provides new insights into soil microbial communities

The analysis of phospholipid fatty acids (PLFAs) is one of the most common methods used to quantify the abundance, and analyse the community structure, of soil microbes. The PLFA extraction method can yield two additional lipid fractions—neutral lipids and glycolipids—which potentially hold additional, valuable information on soil microbial communities. Yet its quantitative sensitivity on complete neutral lipid (NLFA) and glycolipid fatty acid (GLFA) profiles has never been validated. In this study we tested (i) if the high-throughput PLFA method can be expanded to concurrently extract complete NLFA and GLFA profiles, as well as sterols, (ii) whether taxonomic specificities of signature fatty acids are retained across the three lipid fractions in pure culture strains, and (iii) whether NLFAs and GLFAs allow soil-specific fingerprinting to the same extent as PLFA analysis. By adjusting the polarity of chloroform with 2% ethanol for solid phase extraction, pure lipid standards were fully fractionated into neutral lipids, glycolipids, and phospholipids. Sterols eluted in the neutral lipid fraction, and a betaine lipid co-eluted with phospholipids. We found consistent taxonomic specificities of fatty acid markers across the three lipid fractions by analysing pure culture extracts representative of soil microbes. Fatty acid profiles from soil extracts, however, showed stronger differences between PLFAs, NLFAs, and GLFAs than between soil types. This indicates that PLFAs and NLFAs signify different community properties (biomass vs. carbon storage, putatively), and that GLFAs are sensitive markers for community traits which behave differently than PLFAs. Although we consistently found high abundances of characteristic sterols in fungal extracts, the PLFA extraction method only yielded miniscule amounts of ergosterol from soil extracts. We argue that concomitant measurement of fatty acid profiles from all three lipid fractions is a low-effort and potentially information-rich addition to the PLFA method, and discuss its applicability for soil microbial community analyses.

Reproductive quality evaluation of male Indian white prawn Penaeus indicus broodstock fed diets supplemented with polychaete extracts (Marphysa sp.)

ABSTRACT The present study determined the effect of different polychaete extracts, namely, total soluble fraction (TSF), neutral lipid fraction (NLF) and polar lipid fraction (PLF), in the maturation and sperm quality of male Penaeus indicus. Three levels (0.25, 0.50 and 1.00%) of extracts were included using a 3 × 3 factorial design. Groups fed the basal diet (BD) and fresh-frozen diet served as controls. Extracts in varying doses and control groups did not have a significant effect on broodstock survival (67–87%; p = 0.960), maturation rate (42–68%; p = 0.615), inter-spermatophore period (8–10 days; p = 0.505) or sperm viability (97–100%; p = 0.819). However, sperm counts of broodstock fed BD (11.70 × 106 ±1.05 × 106 per spermatophore) and those fed diets supplemented with polychaete extracts were significantly higher compared to that fed with control fresh diet at 0.73 × 106± .09 × 106 (p = 0.001). Spermatophore crude lipid was highest in groups fed 0.25% TSF and 0.25% PLF of broodstock (p =1.0 x10−6 ). Inclusion of TSF (0.25–1.00%) significantly increased the spermatophore crude protein content of broodstock compared to those fed with other diets (p = 1.20 × 10−5). These results demonstrate that NLF and TSF extracts are bioactive components of polychaete which when fed to male P. indicus, can stimulate aspects of sperm production.

Physiological condition of the warty venus (Venus verrucosa L. 1758) larvae modulates response to pile driving and drilling underwater sounds

Noise is now recognized as a new form of pollution in marine coastal habitats. The development of marine renewable energies has introduced new sonorous perturbations, as the wind farm installation requires pile driving and drilling operations producing low frequency sounds at high sound pressure levels. Exponential expansion of offshore wind farms is occurring worldwide, making impact studies, particularly on benthic species highly abundant and diverse in the coastal area used for wind farming, a necessity. As larval recruitment is the basis for establishing a population, we conducted an experimental study to assess the interactive effects of pile driving or drilling sounds and larval rearing temperature on the endobenthic bivalve Venus verrucosa. In ectothermic animals, temperature modifies the organism’s physiology, resulting in performance variability. We hypothesize that temperature modulation could change larval responses to noise and explore the potential interacting effects of temperature and noise. Using two distinct rearing temperatures, physiologically different batches of larvae were produced with contrasting fatty acid content and composition in the neutral and polar lipid fractions. Without defining any absolute audition threshold for the larvae, we demonstrate that the effects of temperature and noise were ontogenic-dependent and modulated larval performance at the peri-metamorphic stage, acting on the metamorphosis dynamic. At the pediveligers stage, a strong interaction between both factors indicated that the response to noise was highly related to the physiological condition of the larvae. Finally, we suggest that underwater noise reduces the compensatory mechanisms established to balance the temperature increase.

Yarrowia lipolytica as a Platform for Punicic Acid Production.

Punicic acid (PuA) is a polyunsaturated fatty acid with significant medical, biological, and nutraceutical properties. The primary source of punicic acid is the pomegranate seed oil obtained from fruits of trees that are mainly cultivated in subtropical and tropical climates. To establish sustainable production of PuA, various recombinant microorganisms and plants have been explored as platforms with limited efficiencies. In this study, the oleaginous yeast Yarrowia lipolytica was employed as a host for PuA production. First, growth and lipid accumulation of Y. lipolytica were evaluated in medium supplemented with pomegranate seed oil, resulting in the accumulation of lipids up to 31.2%, consisting of 22% PuA esterified in the fraction of glycerolipids. In addition, lipid-engineered Y. lipolytica strains, transformed with the bifunctional fatty acid conjugase/desaturase from Punica granatum (PgFADX), showed the ability to accumulate PuA de novo. PuA was detected in both polar and neutral lipid fractions, especially in phosphatidylcholine and triacylglycerols. Promoter optimization for PgFADX expression resulted in improved accumulation of PuA from 0.9 to 1.8 mg/g of dry cell weight. The best-producing strain expressing PgFADX under the control of a strong erythritol-inducible promoter produced 36.6 mg/L PuA. These results demonstrate that the yeast Y. lipolytica is a promising host for PuA production.

Developing advanced models of biological membranes with hydrogenous and deuterated natural glycerophospholipid mixtures

Cellular membranes are complex systems that consist of hundreds of different lipid species. Their investigation often relies on simple bilayer models including few synthetic lipid species. Glycerophospholipids (GPLs) extracted from cells are a valuable resource to produce advanced models of biological membranes. Here, we present the optimisation of a method previously reported by our team for the extraction and purification of various GPL mixtures from Pichia pastoris. The implementation of an additional purification step by High Performance Liquid Chromatography-Evaporative Light Scattering Detector (HPLC-ELSD) enabled for a better separation of the GPL mixtures from the neutral lipid fraction that includes sterols, and also allowed for the GPLs to be purified according to their different polar headgroups. Pure GPL mixtures at significantly high yields were produced through this approach. For this study, we utilised phoshatidylcholine (PC), phosphatidylserine (PS) and phosphatidylglycerol (PG) mixtures. These exhibit a single composition of the polar head, i.e., PC, PS or PG, but contain several molecular species consisting of acyl chains of varying length and unsaturation, which were determined by Gas Chromatography (GC). The lipid mixtures were produced both in their hydrogenous (H) and deuterated (D) versions and were used to form lipid bilayers both on solid substrates and as vesicles in solution. The supported lipid bilayers were characterised by quartz crystal microbalance with dissipation monitoring (QCM-D) and neutron reflectometry (NR), whereas the vesicles by small angle X-ray (SAXS) and neutron scattering (SANS). Our results show that despite differences in the acyl chain composition, the hydrogenous and deuterated extracts produced bilayers with very comparable structures, which makes them valuable to design experiments involving selective deuteration with techniques such as NMR, neutron scattering or infrared spectroscopy.

MICROWAVE TREATMENT FOR HIGH LIPID PRODUCTION IN SCENEDESMUS OBLIQUUS

Microalgae with high oil content are only alternative for decreasing fossil fuel supplies, but more work remains to be done to improve the lipid content of microalgae strains. In this study, strain improvement is done using microwave radiation in Scenedesmus obliquus to increase the production of triacylglycerol, which is the main source of biodiesel. Microalgal culture were exposure to varied microwave irradiation over different time periods. Maximum increase of 2.22-fold in biomass and 2.5-fold in triacylglycerol was observed for microwave irradiation of 25mins and 20mins respectively. The percentage of some monounsaturated fatty acids increased in gas chromatographic examination of neutral lipid fractions from total lipids of microwave irradiated samples, which is considered as one of the preferable properties of biodiesel.

Nigella sativa L. oil: Study of its toxicity, antiradical activity, and effect on circulating xanthine oxidoreductase

Introduction: Nigella sativa L. is a widely used medicinal plant throughout the world. The low toxic effects and low price of this plant make it an excellent treatment choice for many diseases. The present study aims to investigate the hepatoprotective effect of N. sativa L. total oil (TO) and its neutral lipid fraction (NLF) via the estimation of circulating xanthine oxidoreductase (XOR) level and anti-XOR antibodies titer. Methods: Antiradical activities of TO and NLF, in vitro, were carried out using three reactive oxygen species (ROS), superoxide anion, hydroxyl radical, and hydrogen peroxide. In vivo study was conducted to determine the possible protective effects of TO and NLF against ethanol-induced hepatotoxicity in rats feeding Lieber-DeCarli liquid diet in both sera and liver homogenate. Before conducting the hepatoprotective effects, we assessed the toxicity of our extracts using the same animal model. The administrated doses of 400 mg/kg for TO and 300 mg/kg for NLF showed no toxic effects. Results: ELISA assay indicated a significant increase (P<0.001) in the level of XOR and the titer of anti-XOR antibodies in rats treated with ethanol compared to the control group. After treatment with TO and NLF, the titer of anti-XOR antibodies and the level of XOR decreased significantly compared to the control group. Conclusion: XOR plays an important role in alcohol liver pathologies as a major source of free radicals. In addition, TO and NLF have significant potential as liver protective agents and might be utilized as new antioxidant therapeutics.

HIGH OLEIC SUNFLOWER OIL DECREASES ENDOGENOUS BIOSYNTHESIS OF ENERGY FATTY ACIDS AND INCREASES ENDOGENOUS BIOSYNTHESIS OF ω-3 LONG-CHAIN PUFA

The work shows that Fatty acids of dietary fats provide two main functions in the human and animal body: energy and structural-regulatory, Polyunsaturated fatty acids (PUFA) are the basis of membrane lipids of all cells of the body The structure and functional activity of cells, their resistance to pathogenic factors depends on the ratio of ω-6 / ω-3 PUFA. Аdherence to recommended metabolic energy reserves in poultry feed is essential to optimize feed costs. The use of oils or fats is a common economic practice in modern poultry production. Energy functions are carried out due to the oxidation of energy fatty acids in mitochondria, which include, first of all, palmitic (С16:0), palmitooleic (С16:0), stearic (С18:1) and oleic (С18:1). In addition to providing energy, edible oil can also enhance dietary palatability, reduce dustiness, and increase lipoprotein hydrolysis to promote the production of essential fatty acids. Adipose tissue should be considered not only as a source of various fatty acids, but also as an important endocrine organ that takes an active part in the activity of the immune system. To determine the effect of a diet with high oleic sunflower oil on the content of energy fatty acids (EFA) and long-chain PUFA (LCPUFA) in rat liver lipids. We used high oleic sunflower oil (HOSO) containing 85.5% oleic acid. The rats were fed a fat-free diet (FFD) and diets with 5 or 15% HOSO for 35 days. Lipids were extracted from the liver and separated into three fractions: neutral lipids (NL), phospholipids (PL) and free fatty acids (FFA). The fatty acid composition of lipid fractions was determined by gas chromatography. FFA are the sum of the following acids:С16:0, С16:1, С18:0, С18:1andС18:2. LCPUFA are presented С20:4 ω-6, С20:5 ω-3, С22:5 ω-3and С22:6 ω-3. Most of all, EFA is contained in the NL fraction (89%), then in the PL fraction (79%), and least of all in the FFA fraction (68%). LCPUFA is found most of all in the FFA fraction (20%), then in the PL fraction (13%), and least of all in the NL fraction (2%). In rats that received fat diets, the content of EFA increased in the NL fraction by 2-3%, in the FFA fraction by 5-8%, and did not change in the PL fraction. The content of LCPUFA ω-6 (arachidonic acid) with fat nutrition dose-dependently decreases in the fraction of NL and FFA and does not change in the fraction of PL. On the contrary, the content of ω-3 LCPUFA increases in rats treated with HOSO in all lipid fractions. Also, the ω-6/ω-3 LCPUFA ratio is significantly reduced in rats treated with HOSO. Consumption of HOSO stimulates endogenous biosynthesis of ω-3 LCPUFA.

Efficient method for the determination of the neutral lipid content of oil-producing microalgae strains required for biodiesel

The continued use of petroleum-derived fuels is unsustainable, as these fuels contribute to the environmental accumulation of carbon dioxide. Renewable fuels are necessary for environmental and economic sustainability. Microalgae appear to be the source of renewable biofuel that can meet the global demand for transport fuels. Like plants, microalgae use sunlight to produce oils but they do so more efficiently than crop plants. Oil productivity of many microalgae greatly exceeds the oil productivity of the best producing oil crops. Using microalgae to produce biofuel does not occupy agricultural land will not compromise production of food and other products derived from crops.During the growth of microalgae, the quantitative and qualitative values of produced lipids are constantly changing. Therefore, a laboratory lipid determination method had to be developed. Among the various industrial lipid extraction and purification methods, we only dealt with methods that can also be used in the laboratory. Because only neutral lipid fraction can be used to produce biofuels, an optimized method had to be developed to separate and determine the amount of neutral lipids. Each step was optimized in terms of time, volume, amount of chemical, simplicity and accuracy, these were the main aspects so that the method could be used for a wide range of lipid content and composition. The developed microextraction method enables the examination of a small amount of sample (100 mg) in a small volume (5 mL), in that way many samples can be processed simultaneously. This method saves time, money and chemicals in the laboratory while improving the accuracy of the lipid content determination.

Enhancement of Lipid and Biomass Production in Microalgae Scenedesmus abundans by Microwave Irradiation

Due to the rising depletion of fossil fuels and increased energy demands, human society is looking for clean sustainable energy. Commercially produced algal biodiesel is limited by the expense and difficulty of oil extraction and subsequent biodiesel conversion technologies. Microalgae with high oil content are only alternatives for decreasing fossil fuel supplies, but more work remains to be done to improve the lipid content of microalgae strains. In this study, strain improvement is done using microwave radiation in Scenedesmus abundans to increase the production of triacylglycerol, which is the main source of biodiesel. Microalgal cultures were exposed to varied microwave irradiation over different time periods. Under microwave irradiation, 20-25 mins reaction time seems suitable for the complete in situ transesterification reaction. Microwave heating transesterification has been shown to be more effective for adequate biodiesel yield compared to the conventional transesterification process. Maximum increase of 2.22-fold in biomass, and 2.5-fold in triacylglycerol was observed for microwave irradiation of 25 mins and 20 mins intervals respectively. The percentage of some monounsaturated fatty acids increased in gas chromatographic examination of neutral lipid fractions from total lipids of microwave irradiated samples, is considered as one of the preferable properties of biodiesel. According to our study findings, Scenedesmus abundans qualifies as the most efficient feedstock for biodiesel production, and microwave-assisted in situ transesterification reduces the requirement for a large amount of solvents, longer reaction times, and high reaction temperatures and pressures.

Enhancement of Lipid and Biomass Production in Microalgae Scenedesmus abundans by Microwave Irradiation

Due to the rising depletion of fossil fuels and increased energy demands, human society is looking for clean sustainable energy. Commercially produced algal biodiesel is limited by the expense and difficulty of oil extraction and subsequent biodiesel conversion technologies. Microalgae with high oil content are only alternatives for decreasing fossil fuel supplies, but more work remains to be done to improve the lipid content of microalgae strains. In this study, strain improvement is done using microwave radiation in Scenedesmus abundans to increase the production of triacylglycerol, which is the main source of biodiesel. Microalgal cultures were exposed to varied microwave irradiation over different time periods. Under microwave irradiation, 20-25 mins reaction time seems suitable for the complete in situ transesterification reaction. Microwave heating transesterification has been shown to be more effective for adequate biodiesel yield compared to the conventional transesterification process. Maximum increase of 2.22-fold in biomass, and 2.5-fold in triacylglycerol was observed for microwave irradiation of 25 mins and 20 mins intervals respectively. The percentage of some monounsaturated fatty acids increased in gas chromatographic examination of neutral lipid fractions from total lipids of microwave irradiated samples, is considered as one of the preferable properties of biodiesel. According to our study findings, Scenedesmus abundans qualifies as the most efficient feedstock for biodiesel production, and microwave-assisted in situ transesterification reduces the requirement for a large amount of solvents, longer reaction times, and high reaction temperatures and pressures.

Evaluation of maturation promoting factor in polychaete ( Marphysa sp.) on Indian White Prawn , Penaeus indicus female broodstock

Polychaete is considered the best maturation diet for penaeids; however, fluctuating supply and quality warrant detailed studies to understand the specific maturation-promoting factors present in polychaete. Indian white prawn, Penaeus indicus, was fed diets supplemented with different fractions of Marphysa sp. extracts. Fractions, such as the total soluble fraction (TSF), neutral lipid fraction (NLF), and polar lipid fraction (PLF), were incorporated in the maturation diet at 0.25, 0.50, and 1.00% following a 3 × 3 factorial design. One group was fed with basal diet (BD), and another was fed with fresh squid, mussel, and polychaete, serving as the control. After a 30-day feeding trial, results showed that the inclusion of polychaete extracts in the diet significantly improved P. indicus maturation compared to groups fed BD and control with 40% maturation rates (MR) (p = 3.4 × 10−4). MR was optimum in groups receiving diets supplemented with ≥0.5% TSF (70.00% ± 0.00) and NLF (60.00% ± 5.77). Similar improved MR was achieved in treatments receiving ≥0.25% PLF supplementation (60% ± 0.00). Accordingly, relative expression of ovarian vitellogenin mRNA of broodstock fed under 0.25 and 0.50% PLF group was 4.44 and 3.96 folds higher than BD, respectively (p = 0.003). No significant differences were detected in the broodstock survival, latency period, hepatosomatic, and gonadosomatic indices. Biochemical content analyses showed no significant differences among the nine treatments except for broodstock's higher ovary protein content in the TSF group (p = 0.037). This study highlights PLF as the most potent component of the polychaete extract in promoting gonad maturation in P. indicus maturation supplemented at a 0.25% optimum inclusion level.

Intravital Subcellular Microscopy of the Mammary Gland.

The mammary gland constitutes a model par excellence for investigating epithelial functions, including tissue remodeling, cell polarity, and secretory mechanisms. During pregnancy, the gland expands from a primitive ductal tree embedded in a fat pad to a highly branched alveolar network primed for the formation and secretion of colostrum and milk. Post-partum, the gland supplies all the nutrients required for neonatal survival, including membrane-coated lipid droplets (LDs), proteins, carbohydrates, ions, and water. Various milk components, including lactose, casein micelles, and skim-milk proteins, are synthesized within the alveolar cells and secreted from vesicles by exocytosis at the apical surface. LDs are transported from sites of synthesis in the rough endoplasmic reticulum to the cell apex, coated with cellular membranes, and secreted by a unique apocrine mechanism. Other preformed constituents, including antibodies and hormones, are transported from the serosal side of the epithelium into milk by transcytosis. These processes are amenable to intravital microscopy because the mammary gland is a skin gland and, therefore, directly accessible to experimental manipulation. In this paper, a facile procedure is described to investigate the kinetics of LD secretion in situ, in real-time, in live anesthetized mice. Boron-dipyrromethene (BODIPY)665/676or monodansylpentane are used to label the neutral lipid fraction of transgenic mice, which either express soluble EGFP (enhanced green fluorescent protein) in the cytoplasm, or a membrane-targeted peptide fused to either EGFP or tdTomato. The membrane-tagged fusion proteins serve as markers of cell surfaces, and the lipid dyes resolve LDs ≥ 0.7 µm. Time-lapse images can be recorded by standard laser scanning confocal microscopy down to a depth of 15-25 µm or by multiphoton microscopy for imaging deeper in the tissue. The mammary gland may be bathed with pharmacological agents or fluorescent dyes throughout the surgery, providing a platform for acute experimental manipulations as required.

Obtaining EPA-rich polar lipids from microalga Nannochloropsis sp. by silica-gel chromatography using non-toxic solvents

Current research indicates that n-3 polyunsaturated fatty acids (PUFAs) bind to polar lipids (phospholipids and glycolipids) seem to exert a greater bioavailability compared to their neutral forms. The aim of this work was to obtain eicosapentaenoic acid (EPA) rich polar lipids from the saponifiable lipids (SLs) extracted from the microalga Nannochloropsis sp. (33.4 ± 0.1% of EPA; 60 ± 0.6% polar lipids) by fractionation using silica-gel columns and importantly, non-polar and polar (ethanol) non-toxic solvents. Nowadays, few studies have been conducted towards the extraction and purification of polar lipids. Firstly, the solvent type for obtaining the neutral saponifiable lipid (NSL) fraction (ethyl acetate, EA, butyl acetate, BA) and the SL/silica-gel, SL/BA, and SL/ethanol ratios were optimized in a small silica-gel cartridge (0.69 g silica gel). The optimized conditions were an SL/silica-gel ratio of 22.6 mg/g, an SL/BA ratio of 1.56 mg/mL and an SL/ethanol ratio of 0.312 mg/mL. Next, the fractionation scale was increased to a column containing 10 g of silica-gel. At this scale, a BA SL fraction was obtained with 96.2 ± 0.5% of NSLs, and an ethanol SL fraction containing 97.7 ± 0.3% of polar lipids and 44.9 ± 0.2% of EPA. In the ethanol fraction, 86.6 ± 0.2% of the polar lipids and 71.5 ± 0.4% of the EPA from the SL microalgal extract were recovered. Consequently, EPA-rich polar lipids were obtained at high yields and purities, which could be used as a source of n-3 PUFAs with greater bioavailability than those based on neutral lipids.

Effect of dietary fats on endogenous oleic acid biosynthesis in rat liver

Aim: Determine the effect of dietary fats with different fatty acid composition on the biosynthesis of oleic acid and its metabolic precursors in the liver .
 Methods: High linoleic sunflower oil (HLSO), high oleic sunflower oil (HOSO) and palm oil (PO) were used. Rats were fed a semi-synthetic fat-free diet (FFD) and fat diets containing 5 % of the above oils (instead of starch) for 30 days. Liver lipids were divided into 3 fractions: neutral lipids (NL), phospholipids (PL) and free fatty acids (FFA). The fatty acid composition of the fractions was determined by gas chromatography. The “activity” of fatty acid synthase was determined from the total content of the products of this reaction (C16:0 and C16:1). The “activity” of palmitic acid elongase was determined by the ratio С18:0/С16:0, as well as by the formula (С18:0+С18:1)/(С16:0–С16:1). The “activity” of stearic acid desaturase (SCD1) was determined by the ratio C16:1/C16:0 (SCD16) and by the ratio C18:1/C18:0 (SCD18).
 Results: In rats treated with fat diets, the content of palmitic and oleic acids is reduced only in the NL fraction, and to the greatest extent when consuming the diet with HLSO. The “activity” of palmitic acid elongase increases significantly with the consumption of a diet with HLSO. SCD16 desaturase “activity” decreases with fat diet, while SCD18 desaturase “activity” increases. The level of SCD18 is significantly higher than the level of SCD16. Consumption of HLSO reduces the content of ω-3 PUFA in rat liver lipids, while the intake of HOSO increases it.
 Conclusions: HLSO diet reduces the endogenous biosynthesis of oleic and palmitic acids, as determined by the analysis of the rat liver NL fraction. A fat diet reduces SCD16 “activity” but increases SCD18 “activity”, especially when fed a diet with HOSO. The diet with HLSO reduces the content of ω-3 PUFA in liver lipids.