Omega-3 Fatty Acids are Inversely Related to Callous and Unemotional Traits in Adolescent Boys with Attention Deficit Hyperactivity Disorder

A number of research studies have reported abnormal plasma fatty acid profiles in children with ADHD along with some benefit of n−3 to symptoms of ADHD. However, it is currently unclear whether (lower) long chain-polyunsaturated fatty acids (LC-PUFAs) are related to ADHD pathology or to associated behaviours. The aim of this study was to test whether (1) ADHD children have abnormal plasma LC-PUFA levels and (2) ADHD symptoms and associated behaviours are correlated with LC-PUFA levels. Seventy-two, male children with (n=29) and without a clinical diagnosis of ADHD (n=43) were compared in their plasma levels of LC-PUFA. Plasma DHA was higher in the control group prior to statistical correction. Callous–unemotional (CU) traits were found to be significantly negatively related to both eicosapentaenoic acid (EPA), and total omega-3 in the ADHD group. The findings unveil for the first time that CU and anti-social traits in ADHD are associated with lower omega-3 levels.

Deleterious Effects of Lard-enriched Diet on Tissues Fatty Acids Composition and Hypothalamic Insulin Actions

Altered tissue fatty acid (FA) composition may affect mechanisms involved in the control of energy homeostasis, including central insulin actions. In rats fed either standard chow or a lard-enriched chow (high in saturated/low in polyunsaturated FA, HS-LP) for eight weeks, we examined the FA composition of blood, hypothalamus, liver, and retroperitoneal, epididymal and mesenteric adipose tissues. Insulin-induced hypophagia and hypothalamic signaling were evaluated after intracerebroventricular insulin injection. HS-LP feeding increased saturated FA content in adipose tissues and serum while it decreased polyunsaturated FA content of adipose tissues, serum, and liver. Hypothalamic C20:5n-3 and C20:3n-6 contents increased while monounsaturated FA content decreased. HS-LP rats showed hyperglycemia, impaired insulin-induced hypophagia and hypothalamic insulin signaling. The results showed that, upon HS-LP feeding, peripheral tissues underwent potentially deleterious alterations in their FA composition, whist the hypothalamus was relatively preserved. However, hypothalamic insulin signaling and hypophagia were drastically impaired. These findings suggest that impairment of hypothalamic insulin actions by HS-LP feeding was not related to tissue FA composition.

Chromatographic determination of fatty acids profiles in regionally-produced Portuguese goat cheese along curing

The biochemistry of cheese ripening involves mechanisms such as glycolysis, proteolysis and lipolysis. Fatty acids are released by the action of lipases from different sources, milk, rennet, bacteria, moulds included as secondary starters, and other exogenous lipases, during lipolysis [1]. The composition of the lipid fraction contributes positively to the flavour of cheese, for being precursors of more complex aroma compounds responsible for the characteristic “goaty flavour” of goat cheeses [2]. Goat milk is recognized by its easier digestibility, alkalinity, buffering capacity and certain therapeutic values in medicine and human nutrition [3]. A high total content of fatty acids is strongly linked to a rancid and tart off flavour in goat milk and may be considered undesirable in most cheese varieties [4]. In this sense, the purpose of the present study was to examine the composition and changes in fatty acids and saponification value of goat cheese during curing period (2, 7 and 12 months). Goat cheese was made in industrial unit of Cachão - Mirandela (Trás-os- Montes) with raw milk Serrana goats’ race, salt and rennet from animal origin. During the first two months, the samples were stored in a ripening chamber (9.5-11 °C and RH 75-85%). From the second month to one year, the samples were stored in a preservation chamber (10.5-12 °C and RH 75-85%). The fatty acids profile of the inner part of the cheese was analyzed by gas-chromatography coupled to flame ionization detection (GC-FID). The degree of saponification was determined both in the crust and inside the cheese by HCl titration of ethanol KOH solution of the samples. Twenty-six fatty acids (FA) were identified and quantified in the inner part of the cheese (total fat was 45-46 g/100 g during the curing period). Saturated fatty acids (SFA) did not change up to 7 months of curing, increasing only after 12 months, being palmitic (C16:0), stearic (C18:0), myristic (C14:0) and capric (C10:0) acids the most abundant FA in this class. Monounsaturated fatty acids (MUFA) decreased only after 12 months, and oleic acid (C18:1) was the predominant FA. In polyunsaturated fatty acids (PUFA) class, the most abundant were linoleic (C18:2) and linolenic (C18:3) acids, and followed the same tendency of MUFA. This is corroborated by an increase in the degree of saponification, either in the crust as in the inner part of the cheese, after 12 months of curing, probably related with the saturation of the fatty acids [3]. Extra-long curing can be done in cheeses produced with goat milk up to seven months of storage without changing the total fat and individual FA content.

Enhancing bioactive fatty acids of the meat from lambs reared in intensive systems through nutritional modulation

Tese de Doutoramento em Ciências Veterinárias, especialidade de Produção Animal

Ethyl-eicosapentaenoic acid in first-episode psychosis : a 1H-MRS study

Ethyl-eicosapentaenoic acid (E-EPA) is an omega-3 fatty acid that has been used in a range of neuropsychiatric conditions with some benefits. However, its mechanism of action is unknown. Here, we investigate its effects on in vivo brain metabolism in first-episode psychosis (FEP). Proton magnetic resonance spectroscopy at 3 T was performed in the temporal lobes of 24 FEP patients before and after 12 weeks of treatment in the context of a larger double-blind, placebo-controlled E-EPA augmentation study. Treatment group effects for glutathione (F1,12=6.1, p=0.03), and a hemisphere-by-group interaction for glutamine/glutamate (F1,20=4.4, p=0.049) were found. Glutathione increased bilaterally and glutamate/glutamine increased in the left hemisphere following E-EPA administration. Improvement in negative symptoms correlated with metabolic brain changes, particularly glutathione (r=-0.57). These results suggest that E-EPA augmentation alters glutathione availability and modulates the glutamine/glutamate cycle in early psychosis, with some of the metabolic brain changes being correlated with negative symptom improvement. Larger confirmatory studies of these postulated metabolic brain effects of E-EPA are warranted.

Mass spectrometric study of the dissociation of Group XI metal complexes with fatty acids and glycerolipids : Ag2H+ and Cu2H+ ion formation in the presence of a double bond

Results of mass spectrometric studies are reported for the collisional dissociation of Group XI (Cu, Ag, Au) metal ion complexes with fatty acids (palmitic, oleic, linoleic and a-linolenic) and glycerolipids. Remarkably, the formation of M2H+ ions (M = Cu, Ag) is observed as a dissociation product of the ion complexes containing more than one metal cation and only if the lipid in the complex contains a double bond. Ag2H+ is formed as the main dissociation channel for all three of the fatty acids containing double bonds that were investigated while Cu2H+ is formed with one of the fatty acids and, although abundant, is not the dominant dissociation channel. Also. Cu(I) and Ag(I) ion complexes were observed with glycerolipids (including triacylglycerols and glycerophospholipids) containing either saturated or unsaturated fatty acid substituents. Interestingly. Ag2H+ ion is formed in a major fragmentation channel with the lipids that are able to form the complex with two metal cations (triacylglycerols and glycerophosphoglycerols), while lipids containing a fixed positive charge (glycerophospocholines) complex only with a single metal cation. The formation of Ag2H+ ion is a significant dissociation channel from the complex ion Ag-2(L-H)(+) where L = Glycerophospholipid (GP) (18:1/18:1). Cu(I) also forms complexes of two metal cations with glycerophospholipids but these do not produce Cu2H+ upon dissociation. Rather organic fragments, not containing Cu(I), are formed, perhaps due to different interactions of these metal cations with lipids resulting from the much smaller ionic radius of Cu(I) compared to Ag(I) (C).

Evaluation of a pilot-scale oil extraction from microalgae for biodiesel production

Biodiesel derived from microalgae is one of a suite of potential solutions to meet the increasing demand for a renewable, carbon-neutral energy source. However, there are numerous challenges that must be addressed before algae biodiesel can become commercially viable. These challenges include the economic feasibility of harvesting and dewatering the biomass and the extraction of lipids and their conversion into biodiesel. Therefore, it is essential to find a suitable extraction process given these processes presently contribute significantly to the total production costs which, at this stage, inhibit the ability of biodiesel to compete financially with petroleum diesel. This study focuses on pilot-scale (100 kg dried microalgae) solvent extraction of lipids from microalgae and subsequent transesterification to biodiesel. Three different solvents (hexane, isopropanol (IPA) and hexane + IPA (1:1)) were used with two different extraction methods (static and Soxhlet) at bench-scale to find the most suitable solvent extraction process for the pilot-scale. The Soxhlet method extracted only 4.2% more lipid compared to the static method. However, the fatty acid profiles of different extraction methods with different solvents are similar, suggesting that none of the solvents or extraction processes were biased for extraction of particular fatty acids. Considering the cost and availability of the solvents, hexane was chosen for pilot-scale extraction using static extraction. At pilot-scale the lipid yield was found to be 20.3% of total biomass which is 2.5% less than from bench scale. Extracted fatty acids were dominated by polyunsaturated fatty acids (PUFAs) (68.94±0.17%) including 47.7±0.43 and 17.86±0.42% being docosahexaenoic acid (DHA) (C22:6) and docosapentaenoic acid (DPA) (C22:5, ω-3), respectively. These high amounts of long chain poly unsaturated fatty acids are unique to some marine microalgae and protists and vary with environmental conditions, culture age and nutrient status, as well as with cultivation process. Calculated physical and chemical properties of density, viscosity of transesterified fatty acid methyl esters (FAMEs) were within the limits of the biodiesel standard specifications as per ASTM D6751-2012 and EN 14214. The calculated cetane number was, however, significantly lower (17.8~18.6) compared to ASTM D6751-2012 or EN 14214-specified minimal requirements. We conclude that the obtained microalgal biodiesel would likely only be suitable for blending with petroleum diesel to a maximum of 5 to 20%.

The effect of exercise on the skeletal muscle phospholipidome of rats fed a high-fat diet

The aim of this study was to examine the effect of endurance training on skeletal muscle phospholipid molecular species from high-fat fed rats. Twelve female Sprague-Dawley rats were fed a high-fat diet (78.1% energy). The rats were randomly divided into two groups, a sedentary control group and a trained group (125 min of treadmill running at 8 m/min, 4 days/wk for 4 weeks). Forty-eight hours after their last training bout phospholipids were extracted from the red and white vastus lateralis and analyzed by electrospray-ionization mass spectrometry. Exercise training was associated with significant alterations in the relative abundance of a number of phospholipid molecular species. These changes were more prominent in red vastus lateralis than white vastus lateralis. The largest observed change was an increase of similar to 30% in the abundance of 1-palmitoyl-2-linoleoyl phosphatidylcholine ions in oxidative fibers. Reductions in the relative abundance of a number of phospholipids containing long-chain n-3 polyunsaturated fatty acids were also observed. These data suggest a possible reduction in phospholipid remodeling in the trained animals. This results in a decrease in the phospholipid n-3 to n-6 ratio that may in turn influence endurance capacity.

Effect of temperature and moisture on high pressure lipid/oil extraction from microalgae

Commercially viable carbon–neutral biodiesel production from microalgae has potential for replacing depleting petroleum diesel. The process of biodiesel production from microalgae involves harvesting, drying and extraction of lipids which are energy- and cost-intensive processes. The development of effective large-scale lipid extraction processes which overcome the complexity of microalgae cell structure is considered one of the most vital requirements for commercial production. Thus the aim of this work was to investigate suitable extraction methods with optimised conditions to progress opportunities for sustainable microalgal biodiesel production. In this study, the green microalgal species consortium, Tarong polyculture was used to investigate lipid extraction with hexane (solvent) under high pressure and variable temperature and biomass moisture conditions using an Accelerated Solvent Extraction (ASE) method. The performance of high pressure solvent extraction was examined over a range of different process and sample conditions (dry biomass to water ratios (DBWRs): 100%, 75%, 50% and 25% and temperatures from 70 to 120 ºC, process time 5–15 min). Maximum total lipid yields were achieved at 50% and 75% sample dryness at temperatures of 90–120 ºC. We show that individual fatty acids (Palmitic acid C16:0; Stearic acid C18:0; Oleic acid C18:1; Linolenic acid C18:3) extraction optima are influenced by temperature and sample dryness, consequently affecting microalgal biodiesel quality parameters. Higher heating values and kinematic viscosity were compliant with biodiesel quality standards under all extraction conditions used. Our results indicate that biodiesel quality can be positively manipulated by selecting process extraction conditions that favour extraction of saturated and mono-unsaturated fatty acids over optimal extraction conditions for polyunsaturated fatty acids, yielding positive effects on cetane number and iodine values. Exceeding biodiesel standards for these two parameters opens blending opportunities with biodiesels that fall outside the minimal cetane and maximal iodine values.

Straminipilan protists : a study of their taxonomic position, morphology and abiotic stress-mediated gene expression

Thraustochytrids have become of considerable industrial and scientific interest in the past decade due to their health benefits. They have been proven to be the principle source in marine and estuarine fish diets with high percentage of long chain (LC) or polyunsaturated fatty acids (PUFA). Therefore, the oil extracted from fish for human document.forms[0].elements[13].select();consumption is rich in PUFA with high omega-3 fatty acid content. Docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) of all of the omega-3 fatty acids, are considered beneficial essential oils for humans with a wide range of health benefits. These include brain and neural development in infants, general wellbeing of adults and drug delivery through precursor molecules. They have become one of the most extensively studied organisms for industrial oil preparations as PUFA extraction from fish becomes less profitable. Many forms of these Thraustochytrid oils are being trialled for human consumption all over the world. In Australia, there has been little research performed on these organisms in the past ten years. A few Australian studies have been conducted in the form of comparative studies related to PUFA production within the related genera, but not focussed on their identification or cellular and genomic characterisation. Therefore, the main aim of this study was to investigate the morphological and genetic characteristics of Australian Thraustochytrids in order to aid in their identification and characterisation, as well as to better understand the effect of environmental conditions in the regulation of PUFA production. It was also noted that there was a knowledge gap in the preservation and total genomic DNA extraction of these organisms for the purposes of scientific research. The cryopreservation of these organisms for studies around the world follows existing generic methods. However, it is well understood that many of these generic methods attract not only high costs for chemicals, but also uses considerable storage space and other resources, all of which can be improved with new or modified approaches. In this context, a simple and inexpensive bead preservation method is described, without compromising the storage shelf life. We also describe, for the first time, the effects of culture age on the successful cryopreservation of Thraustochytrids. It was evident in the literature that DNA and RNA extractions for molecular and genetic studies of Thraustochytrids follow the classical phenol-chloroform extraction methods. It was also observed that modern protocols failed to avoid the use of phenol-chloroform rather than improving preparation and cell disruption. In order to provide a high quantity and quality DNA extraction, a modified protocol has been introduced that employs the use of modern commercial extraction kits and standard laboratory equipment. Thraustochytrids have been shown to be highly conserved in their 18S rDNA gene sequences, which is used as the current standard for identification. It was demonstrated that the 18S rDNA gene sequence limits the recognition of closely related genera or within the genera from each member. Therefore, it was proposed that another profile, such as a randomly amplified polymorphic DNA (RAPD) based profiling system, be tested for use in the characterisation of Thraustochytrids. The RAPD profiles were shown to provide a unique DNA fingerprint for each isolate and small variations in their genome were able to be detected. This method involved the use of a minimum number of standard arbitrary primers and with an increase in the number of different primers used, a very high discrimination between organisms could be achieved. However, the method was not suitable for taxonomic purposes because the results did not correlate with other taxonomic features such as morphology. Another knowledge gap was found with respect to Australian Thraustochytrid growth characteristics, in that these had not been recorded and published. In order to rectify this, a record of colony and microscopic features of 12 selected isolates was performed. The results of preliminary studies indicated that further microbiological and biochemical studies are needed for full characterisation of these organisms. This information is of great importance to bio-prospecting of new Thraustochytrids from Australian ecosystems and would allow for their accurate identification, and so permit the prediction of their PUFA capability by comparison with related genera/species. It was well recognized that environmental stress plays a role in the PUFA production and is mainly due to the reactive oxygen species as abiotic stress (Chiou et al., 2001; Okuyama et al., 2008; Shabala et al., 2009; Shabala et al., 2001). In this aspect, this study makes the first attempt towards better understanding of this phenomenon by way of the use of real-time PCR for the detection of environmental effects on the regulation of PUFA production. Three main environmental conditions including temperature, pH and oxygen availability were monitored as stress inducers. In summary, this study provides novel approaches for the preservation and handling of Thraustochytrids, their molecular biological features, taxonomy, characterisation and responses to environmental factors with respect to their oil production enzymes. The information produced from this study will prove to be vital for both industrial and scientific investigations in the future.

Comparative analysis and distribution of Omega-3 lcPUFA Biosynthesis genes in marine molluscs

Recent research has identified marine molluscs as an excellent source of omega-3 long-chain polyunsaturated fatty acids (lcPUFAs), based on their potential for endogenous synthesis of lcPUFAs. In this study we generated a representative list of fatty acyl desaturase (Fad) and elongation of very long-chain fatty acid (Elovl) genes from major orders of Phylum Mollusca, through the interrogation of transcriptome and genome sequences, and various publicly available databases. We have identified novel and uncharacterised Fad and Elovl sequences in the following species: Anadara trapezia, Nerita albicilla, Nerita melanotragus, Crassostrea gigas, Lottia gigantea, Aplysia californica, Loligo pealeii and Chlamys farreri. Based on alignments of translated protein sequences of Fad and Elovl genes, the haeme binding motif and histidine boxes of Fad proteins, and the histidine box and seventeen important amino acids in Elovl proteins, were highly conserved. Phylogenetic analysis of aligned reference sequences was used to reconstruct the evolutionary relationships for Fad and Elovl genes separately. Multiple, well resolved clades for both the Fad and Elovl sequences were observed, suggesting that repeated rounds of gene duplication best explain the distribution of Fad and Elovl proteins across the major orders of molluscs. For Elovl sequences, one clade contained the functionally characterised Elovl5 proteins, while another clade contained proteins hypothesised to have Elovl4 function. Additional well resolved clades consisted only of uncharacterised Elovl sequences. One clade from the Fad phylogeny contained only uncharacterised proteins, while the other clade contained functionally characterised delta-5 desaturase proteins. The discovery of an uncharacterised Fad clade is particularly interesting as these divergent proteins may have novel functions. Overall, this paper presents a number of novel Fad and Elovl genes suggesting that many mollusc groups possess most of the required enzymes for the synthesis of lcPUFAs.

A soluble preparation from developing groundnut seeds (Arachis hypogaea) catalyzes de novo synthesis of long chain fatty acids

A 100,000 x g supernatant fraction prepared from developing groundnut seeds (30-35 days after flowering) catalyzed the synthesis of fatty acids from [l-14C]acetate at a rate of 120nmoles of acetate incorporated per hr per gram fresh weight of tissue. 90% of this incorporated label was associated with fatty acids. The major fatty acids formed were stearic- (77%) and palmitic acids (14%) with 4% of oleic acid. The fatty acid synthetase activity was stable when stored at 0-4 degrees C for at least fifteen days. It is concluded from these results that acetyl-coA carboxylase and all the enzymes of fatty acid synthetase from developing groundnut seeds are soluble.

Frictional response of fatty acids on steel

Self-assembled monolayers of fatty acids were formed on stainless steel by room-temperature solution deposition. The acids are covalently bound to the Surface as carboxylate in a bidentate manner. To explore the effect Of Saturation in the carbon backbone on friction in sliding tribology, we Study the response of saturated stearic acid (SA) and unsaturated linoleic acid (LA) as self-assembled monolayers using lateral force microscopy and nanotribometry and when the molecules are dispersed in hexadecane, using pin-on-disc tribometry. Over a very wide range (10 MPa-2.5 GPa) of contact pressures it is consistently demonstrated that the unsaturated linoleic acid molecules yield friction which is significantly lower than that of the saturated stearic acid. it is argued, using density functional theory predictions and XPS of slid track, that when the molecular backbone of unsaturated fatty acids are tilted and pressed strongly by a probe, in tribological contact, the high charge density of the double bond region of the backbone allows coupling with the steel Substrate. The interaction yields a low friction carboxylate soap film on the substrate. The saturated fatty acid does not show this effect.

Hydrolysis of sphingomyelin to ceramide with hydrofluoric acid

Sphingomyelin was hydrolyzed to ceramide near quantitatively on treatment with 40% HF at 40°C for 72 hr. The reaction of sphingomyelin with HF is much slower than phosphoglycerides. HF treatment did not alter either the fatty acid composition or the stereochemical configuration of the sphingosine moiety of ceramide formed.

Spectroscopy of tissue triglyceride composition : In vitro and in vivo studies

Lipid analysis is commonly performed by gas chromatography (GC) in laboratory conditions. Spectroscopic techniques, however, are non-destructive and can be implemented noninvasively in vivo. Excess fat (triglycerides) in visceral adipose tissue and liver is known predispose to metabolic abnormalities, collectively known as the metabolic syndrome. Insulin resistance is the likely cause with diets high in saturated fat known to impair insulin sensitivity. Tissue triglyceride composition has been used as marker of dietary intake but it can also be influenced by tissue specific handling of fatty acids. Recent studies have shown that adipocyte insulin sensitivity correlates positively with their saturated fat content, contradicting the common view of dietary effects. A better understanding of factors affecting tissue triglyceride composition is needed to provide further insights into tissue function in lipid metabolism. In this thesis two spectroscopic techniques were developed for in vitro and in vivo analysis of tissue triglyceride composition. In vitro studies (Study I) used infrared spectroscopy (FTIR), a fast and cost effective analytical technique well suited for multivariate analysis. Infrared spectra are characterized by peak overlap leading to poorly resolved absorbances and limited analytical performance. In vivo studies (Studies II, III and IV) used proton magnetic resonance spectroscopy (1H-MRS), an established non-invasive clinical method for measuring metabolites in vivo. 1H-MRS has been limited in its ability to analyze triglyceride composition due to poorly resolved resonances. Using an attenuated total reflection accessory, we were able to obtain pure triglyceride infrared spectra from adipose tissue biopsies. Using multivariate curve resolution (MCR), we were able to resolve the overlapping double bond absorbances of monounsaturated fat and polyunsaturated fat. MCR also resolved the isolated trans double bond and conjugated linoleic acids from an overlapping background absorbance. Using oil phantoms to study the effects of different fatty acid compositions on the echo time behaviour of triglycerides, it was concluded that the use of long echo times improved peak separation with T2 weighting having a negligible impact. It was also discovered that the echo time behaviour of the methyl resonance of omega-3 fats differed from other fats due to characteristic J-coupling. This novel insight could be used to detect omega-3 fats in human adipose tissue in vivo at very long echo times (TE = 470 and 540 ms). A comparison of 1H-MRS of adipose tissue in vivo and GC of adipose tissue biopsies in humans showed that long TE spectra resulted in improved peak fitting and better correlations with GC data. The study also showed that calculation of fatty acid fractions from 1H-MRS data is unreliable and should not be used. Omega-3 fatty acid content derived from long TE in vivo spectra (TE = 540 ms) correlated with total omega-3 fatty acid concentration measured by GC. The long TE protocol used for adipose tissue studies was subsequently extended to the analysis of liver fat composition. Respiratory triggering and long TE resulted in spectra with the olefinic and tissue water resonances resolved. Conversion of the derived unsaturation to double bond content per fatty acid showed that the results were in accordance with previously published gas chromatography data on liver fat composition. In patients with metabolic syndrome, liver fat was found to be more saturated than subcutaneous or visceral adipose tissue. The higher saturation observed in liver fat may be a result of a higher rate of de-novo-lipogenesis in liver than in adipose tissue. This thesis has introduced the first non-invasive method for determining adipose tissue omega-3 fatty acid content in humans in vivo. The methods introduced here have also shown that liver fat is more saturated than adipose tissue fat.