Method For Analysis Of Fatty Acids Research Articles

Sample Preparation Methods for Fatty Acid Analysis in Different Raw Meat Products by GC-FID

Sample Preparation Methods for Fatty Acid Analysis in Different Raw Meat Products by GC-FID

Development and validation of a gas chromatography\u2013mass spectrometry method to analyze octanoate enrichments at low concentrations in human plasma

A new method for accurately analyzing octanoate enrichment in plasma was developed and validated. Samples were derivatized directly in plasma by transesterification with isobutanol and were analyzed by gas chromatography–mass spectrometry (GC-MS). This method was developed to analyze the precursor enrichment in a stable isotope tracer protocol. Glyceryl tri[1,2,3,4-13C4] octanoate, a stable isotope-labeled medium-chain triglyceride (MCT), was orally administered in combination with (1) exclusively MCT or (2) a combination of protein, carbohydrates, and MCT to investigate the metabolic route of oral MCT under various conditions. Accurate analysis of octanoate enrichment in plasma at concentrations as low as 0.43 μM (lower limit of quantification, LLOQ) was performed. This is an improvement of about twenty times for the LLOQ for analysis of the enrichment of octanoate when compared with the gold-standard method for fatty acid analysis (methyl esterification). Moreover, we found that‚ with this gold-standard method, study samples were easily contaminated with (unlabeled) octanoate from other sources, leading to biased, incorrect results. The precision and linearity obtained using the new method were good (coefficient of variation intraday < 9.1%, interday < 9.3%, R2 of the calibration curve > 0.99). The sensitivity was sufficient for analyzing samples obtained using the stable isotope protocol. This new method is more sensitive than methyl esterification and it minimizes the risk of contamination.Graphical abstract

Profile of dry sausages traditionally prepared in Pirot, eastern Serbia

The aims of this study were to determine the nutritional composition (moisture, protein and total fat) of peglana sausages produced in eastern Serbia and to analyze the composition of fatty acids. Determination of fatty acid composition in the sausages was performed after ripening and after 20 days of storage. Also, a sample preparation method for fatty acid analysis after simultaneous microwave-assisted extraction-esterification was implemented and results were compared with conventional extraction. The results obtained show peglana sausages have high contents of proteins and saturated fatty acids, but no nitrite; the lack of nitrite makes these sausages a suitable product for consumers trying to avoid this additive. The good agreement between results provided by both fat extraction methods demonstrates the usefulness of both methods as routine methods for the treatment of meat samples prior to fatty acid analysis.

Comparison of methylation methods for fatty acid analysis of milk fat

Three acid- and alkaline-catalysed transesterification methods were compared with the aim to validate a simple yet reliable protocol for fatty acid (FA) profiling of milk fat. While both the acid- and alkaline-catalysed methods were able to convert completely triglycerides and phospholipids into fatty acid methyl esters (FAMEs), the acid catalyst caused significant degradation of conjugated linoleic acid C18:2c9t11 at high temperature. Although a milder temperature can mitigate this negative impact, a long reaction time (2 h) is required to achieve full methylation. By contrast, despite being unable to methylate free fatty acids (FFA), the alkaline-catalysed transesterification yielded comparable results for all major FA due to the very low level of FFA in milk. The alkaline-catalysed methylation is benign for C18:2c9t11. We recommend here a simple one-step protocol based on 0.2 M methanolic KOH, a short reaction time (20 min) and a mild reaction temperature (50 °C) for milk FAME preparation.

Comparative studies of immobilized lipase- and acid-catalyzed fatty acid methyl ester synthesis for seed lipid analysis.

Fatty acids are one of the most important nutrients in food. Acid- or base-catalyzed transesterification methods are commonly used for the analysis of fatty acids. However, several drawbacks were reported for these methods, including the isomerization and degradation of fatty acids. Lipase-catalyzed reactions are usually undertaken at mild conditions, preventing such problems. In this study, commercial resin-bound lipase from Candida antartica was tested for possible application in fatty acid methyl ester analysis. Experimental parameters, including temperature, reaction time, and re-cycling were evaluated. The optimized condition was (5-10 mg lipid, 0.5 mL of MeOH, and 50 mg Novozyme 435 in 2 mL toluene, 80°C for 1 h). In optimized condition, the lipase-catalyzed methods yielded similar results with the chemical method. In overall, lipase-catalyzed transesterification can be a useful alternative to acid-catalyzed methods for fatty acid analysis.

VALIDASI METODE ANALISIS ASAM LEMAK TRANS DALAM MAKANAN BERDASARKAN AOCS OFFICIAL METHOD Ce 1h-05

Cardiovascular disease is related to the high consumption of trans fatty acid. This issue has driven SEAFAST IPB Laboratory to develop a trans fatty acid analysis method for gas chromatography based on AOCS Official Method Ce 1h-05. This research was aimed to validate the method via three steps: preparation step, procedure orientation test, and method validation . During instrument performance testing in the preparation step, linearity, limit of detection (LoD), limit of quantitation (LoQ), range, and precision parameters were validated for FAME 9t-C18:1 and C18:2 isomer mix ed standard solution. During the method validation step, specificity, precision and accuracy parameters were validated for 9t-C18:1 compound attached to food matrix, i.e. doughnut . The instrument validation result of 9t-C18:1 standard solution had fulfilled the criteria of linearity (r 2 =1, requirement r 2 > 0.990) within the range of 0.0020 to 0.1200 mg/mL; precision (RSD) 0.24%; LoD 0.0007 mg/mL and LoQ 0.0020 mg/mL. The instrument validation result for each isomer in C18:2 standard solution also met the criteria of linearity (r 2 = 0.9999, requirement r 2 > 0 . 990) with precision (RSD), LoD and LoQ respectively; 1) 9t,12t- C18:2 : 1 . 40%; 0 . 0033 mg/mL; 0.0100 mg/mL; 2) 9t,12c- C18:2 : 1.89%; 0.0015 mg/mL; 0.0045 mg/mL; 3) 9c,12t- C18:2 : 2.39%; 0 . 0015 mg/mL; 0 . 0044 mg/mL; 4) C18:2 cis: 3 . 39%; 0 . 0006 mg/mL; 0 . 0019 mg/mL. The validation result for 9t-C18:1 in 100 mg fat extract of doughnut suggested that the modified method is validated in term of specificity (Rs = 1 . 2), precision (RSD fatty acid content (mg/g) = 1.78%; RSD percentage of fatty acid content towards total fatty acid = 0.72%) and accuracy ( as percentage of recovery ) 80.54-94.25%.

Development of a reliable GC-MS method for fatty acid profiling using direct transesterification of minimal quantities of microscopic orchid seeds

AbstractOrchid seeds are among the smallest seeds in nature and they are naturally rich in fatty acids. However, the fatty acid composition of orchid seeds has not been investigated because the sample masses utilized for widely used methods for fatty acid profiling would generally require prohibitively large numbers (i.e. 10,000s) of seeds. The present work aimed to develop a method for fatty acid analysis using gas chromatography–mass spectrometry on small quantities (mg) of seeds. The method was developed using the seeds of two species,Dactylorhiza fuchsii, a temperate terrestrial, andGrammatophyllum speciosum, a tropical epiphyte. A range of sample masses was tested to determine the minimum mass required to achieve reliable fatty acid composition data. A direct transesterification method was used, which did not require extraction of fatty acids from seeds prior to analysis, and the effects of seed processing (crushed versus intact seeds) and incubation time in toluene on fatty acid yield were tested. Stable fatty acid profiles were obtained using as little as 10 mg of seeds. Neither crushing the seeds nor extending the toluene incubation step had much effect on the fatty acid yield. The simple direct transesterification method presented will enable the fatty acid composition of orchid seeds, and possibly other small seeds, to be determined reliably for studies into seed development, storage and germination.

Development and application of a comparative fatty acid analysis method to investigate voriconazole-induced hepatotoxicity

BackgroundAs fatty acids play an important role in biological regulation, the profiling of fatty acid expression has been used to discover various disease markers and to understand disease mechanisms. This study developed an effective and accurate comparative fatty acid analysis method using differential labeling to speed up the metabolic profiling of fatty acids. MethodsFatty acids were derivatized with unlabeled (D0) or deuterated (D3) methanol, followed by GC-MS analysis. The comparative fatty acid analysis method was validated using a series of samples with different ratios of D0/D3-labeled fatty acid standards and with mouse liver extracts. ResultsUsing a lipopolysaccharide (LPS)-treated mouse model, we found that the fatty acid profiles after LPS treatment were similar between the conventional single-sample analysis approach and the proposed comparative approach, with a Pearson's correlation coefficient of approximately 0.96. We applied the comparative method to investigate voriconazole-induced hepatotoxicity and revealed the toxicity mechanism as well as the potential of using fatty acids as toxicity markers. ConclusionsIn conclusion, the comparative fatty acid profiling technique was determined to be fast and accurate and allowed the discovery of potential fatty acid biomarkers in a more economical and efficient manner.

A comparative study: the impact of different lipid extraction methods on current microalgal lipid research

Microalgae cells have the potential to rapidly accumulate lipids, such as triacylglycerides that contain fatty acids important for high value fatty acids (e.g., EPA and DHA) and/or biodiesel production. However, lipid extraction methods for microalgae cells are not well established, and there is currently no standard extraction method for the determination of the fatty acid content of microalgae. This has caused a few problems in microlagal biofuel research due to the bias derived from different extraction methods. Therefore, this study used several extraction methods for fatty acid analysis on marine microalga Tetraselmis sp. M8, aiming to assess the potential impact of different extractions on current microalgal lipid research. These methods included classical Bligh & Dyer lipid extraction, two other chemical extractions using different solvents and sonication, direct saponification and supercritical CO2 extraction. Soxhlet-based extraction was used to weigh out the importance of solvent polarity in the algal oil extraction. Coupled with GC/MS, a Thermogravimetric Analyser was used to improve the quantification of microalgal lipid extractions. Among these extractions, significant differences were observed in both, extract yield and fatty acid composition. The supercritical extraction technique stood out most for effective extraction of microalgal lipids, especially for long chain unsaturated fatty acids. The results highlight the necessity for comparative analyses of microalgae fatty acids and careful choice and validation of analytical methodology in microalgal lipid research.

Analysis of Fatty Acid Content and Composition in Microalgae

A method to determine the content and composition of total fatty acids present in microalgae is described. Fatty acids are a major constituent of microalgal biomass. These fatty acids can be present in different acyl-lipid classes. Especially the fatty acids present in triacylglycerol (TAG) are of commercial interest, because they can be used for production of transportation fuels, bulk chemicals, nutraceuticals (ω-3 fatty acids), and food commodities. To develop commercial applications, reliable analytical methods for quantification of fatty acid content and composition are needed. Microalgae are single cells surrounded by a rigid cell wall. A fatty acid analysis method should provide sufficient cell disruption to liberate all acyl lipids and the extraction procedure used should be able to extract all acyl lipid classes. With the method presented here all fatty acids present in microalgae can be accurately and reproducibly identified and quantified using small amounts of sample (5 mg) independent of their chain length, degree of unsaturation, or the lipid class they are part of. This method does not provide information about the relative abundance of different lipid classes, but can be extended to separate lipid classes from each other. The method is based on a sequence of mechanical cell disruption, solvent based lipid extraction, transesterification of fatty acids to fatty acid methyl esters (FAMEs), and quantification and identification of FAMEs using gas chromatography (GC-FID). A TAG internal standard (tripentadecanoin) is added prior to the analytical procedure to correct for losses during extraction and incomplete transesterification.

Analysis of Fatty Acid Content and Composition in Microalgae

A method to determine the content and composition of total fatty acids present in microalgae is described. Fatty acids are a major constituent of microalgal biomass. These fatty acids can be present in different acyl-lipid classes. Especially the fatty acids present in triacylglycerol (TAG) are of commercial interest, because they can be used for production of transportation fuels, bulk chemicals, nutraceuticals (ω-3 fatty acids), and food commodities. To develop commercial applications, reliable analytical methods for quantification of fatty acid content and composition are needed. Microalgae are single cells surrounded by a rigid cell wall. A fatty acid analysis method should provide sufficient cell disruption to liberate all acyl lipids and the extraction procedure used should be able to extract all acyl lipid classes. With the method presented here all fatty acids present in microalgae can be accurately and reproducibly identified and quantified using small amounts of sample (5 mg) independent of their chain length, degree of unsaturation, or the lipid class they are part of. This method does not provide information about the relative abundance of different lipid classes, but can be extended to separate lipid classes from each other. The method is based on a sequence of mechanical cell disruption, solvent based lipid extraction, transesterification of fatty acids to fatty acid methyl esters (FAMEs), and quantification and identification of FAMEs using gas chromatography (GC-FID). A TAG internal standard (tripentadecanoin) is added prior to the analytical procedure to correct for losses during extraction and incomplete transesterification.

Report on the distribution of essential and non essential fatty acids in common edible fishes of Porto-Novo coastal waters, southeast coast of India

ObjectiveThe objective of the study was to evaluate the essential and non essential fatty acids and the distribution of Omega-3 and Omega-6 fatty acids in twenty commonly consumed edible fishes of parangipettai coastal waters. MethodsFor fatty acid analysis, each fish specimens were beheaded, eviscerated and filleted manually. The tissue samples were oven dried at 67°C for 24hrs. After that the samples ware grounded finely with pestle and mortar. The saponified samples were cooled at room temperature for 25 min, they were acidified and methylated by adding 2 ml 54% 6 N Hcl in 46% aqueous methanol and incubated at 80°C for 10 min in water bath. Following the base wash step, the FAMEs were cleaned in anhydrous sodium sulphate and then transferred in to GC sample vial for analysis. FAMEs were separated by gas chromatograph. ResultsThe results of the present study revealed that the most abundant individual FAs were Palmitic acid, Oleic acid, Arachidonic acid (AA), Docosahexaenoic acid (DHA) in most the tissues. The total Arachidonic acid (C20:4ω−6) was found to be higher proportion (0.17−4.86%), when compared with other Omega-6 fatty acids. The values found for Linoleic acid (C18:2 ω−6) ranging from 0–7.23%. Siganus javus has 7.23% of Linoleic acid. ConclusionFatty acids are the principle components in lipids. The nutritional importance of fish consumption is in great extent associated with the content of omega-3 fatty acids. Sea food is an important dietary food for human beings. It constitute higher amount of protein, lipids, vitamins and essential and nonessential metals and low concentration of carbohydrates.

Comparison of Transesterification Methods for Fatty Acid Analysis in Higher Fungi: Application to Mushrooms

This study evaluated the effectiveness of two commonly used transesterification methods for the preparation of fatty acid methyl esters (FAMEs) for gas chromatographic analysis. A combination of base followed by acid catalysis [potassium hydroxide/sulfuric acid/methanol (KOH–H2SO4–MeOH)] was compared to an acid-only catalyst [methanolic–boron trifluoride (BF3–MeOH)] for the quantification of the fatty acid content in mushrooms after preextraction of total lipids with chloroform/methanol (2:1 v/v). The mushrooms studied were commonly cultivated species, Agaricus bisporus (brown) and A. bisporus (white), and three wild medicinal mushrooms, turkey tail (Trametes versicolor), artist conk (Ganoderma applanatum), and tinder polypore (Fomes fomentarius). The major fatty acids identified were palmitic (C16:0), stearic (C18:0), oleic (C18:1), and linoleic acid (18:2n6) which constituted 80% to 90% of the total amount. Variation due to the FAME preparation methods was observed in both minor and major fatty acids. Another major observation made between the two methods was the total number of individual fatty acids (FAs) methylated. The base and acid transesterification procedure resulted in the identification of 42 FAMEs, whereas with the acid 38 FAMEs were observed. Therefore, the research recommends the use of a base–acid combination for preparation of FAMEs in mushroom lipids.

Effects of three planting patterns on soil microbial community composition

Aims Planting patterns have direct effects on soil microorganisms.Because of different plants and human activities,planting patterns change soil fertility.Our objective is to understand the effects of different planting patterns on soil microbial community structure.Methods We carried out a long-term field experiment at Jilin Agriculture University that used phospholipid fatty acid analysis method to study the effects of three planting patterns of corn:continuous cropping,non-continuous cropping and uncultivated.Important findings Different planting patterns affected soil microbial community structure.Continuous cropping of corn had the lowest total phospholipid fatty acids and bacterial phospholipid fatty acids(33.12 nmol·g-1 and 18.09 nmol·g-1,respectively).Fungal phospholipid fatty acids and fungi/bacteria with non-continuous cropping were 0.61 nmol.g-1 and 3.06%,respectively,and were significantly lower than with uncultivated and continuous cropping(p 0.05).But the ratio of gram positive phospholipid fatty acids/gram negative phospholipid fatty acids was greater with non-continuous cropping.The uncultivated treatment had greater total and bacterial phospholipid fatty acids(42.98 nmol·g-1 and 24.68 nmol·g-1,respectively),higher contents of gram positive phospholipid fatty acids and gram negative phospholipid fatty acids,and a lower ratio of gram positive phospholipid fatty acids/gram negative phospholipid fatty acids.Principal component analysis showed that soil microbial community structure with cropping was significant different(p 0.05) with that of uncultured in first principal component.Non-continuous cropping and continuous cropping were negatively positioned in first principal component with principal component scores of-2.48 and-1.84,respectively.Uncultivated was in a positive position in first principal component with a principal component score of 2.31.Redundancy analysis of soil microbial community structure and environmental variables showed that soil pH and soil-water stable aggragates had the strongest positive correlation with phospholipid fatty acids and that total nitrogen and available phosphorus were also positively correlated with phospholipid fatty acids.

Fatty acid analysis of Erwinia amylovora from Serbia and Montenegro

Automated method of fatty acid analysis was used to identify and study heterogeneity of 41 Erwinia amylovora strains, originating from 8 plant species grown in 13 locations in Serbia and one in Montenegro. All strains contained 14:0 3OH fatty acid, characteristic for the ?amylovora? group. According to fatty acid composition 39 strains were identified as E. amylovora as the first choice from the database. Due to their specific fatty acid composition, two strains were identified as E. amylovora, but as a second choice. Fatty acid analysis also showed that E. amylovora population from Serbia could be differentiated in three groups, designated in this study as ?, ? and ?. All strains originating from central or south Serbia, as well as four strains from north Serbia clustered into group ?. Group ? and ? contained only strains isolated in northern Serbia (Vojvodina). The results show that E. amylovora population in this area is heterogeneous and indicate pathogen introduction from different directions. Fatty acid analysis enabled identification at species level, as well as new insights of heterogeneity of E. amylovora population.

Recent Advances in Fast Gas-Chromatography: Application to the Separation of Fatty Acid Methyl Esters

Gas-chromatography (GC) is a powerful separation technique for resolving and quantifying a wide range of compounds, such as fatty acid methyl esters (FAME) derivatives. Separation of common FAME is typically achieved on highly polar or polyethylene glycol stationary phases that can separate fatty acids according to chain length, number and geometry of ethylenic double bonds. GC is one of the most commonly available methods for fatty acid analysis with constant improvements for different applications. One such improvement is faster GC analysis, which has long been the focus of investigators researching the application and benefits of this technique. A greater speed of analysis offers many key benefits, such as increased sample throughput, reduced analytical expenses, and increased laboratory productivity. Fast GC analysis can be accomplished by using a short column with reduced column film thickness, high carrier gas velocity, and fast program temperatures. A summary of options for fast GC analysis with emphasis on FAME analysis is discussed in the present review.

Improved method for fatty acid analysis in herbage based on direct transesterification followed by solid-phase extraction

Direct transesterification (DT) and solvent extraction with acid or basic derivatization procedure for fatty acid (FA) analysis in herbage were compared. The highest total FA, α-linolenic and linoleic acid contents were obtained with DT. However, DT also produced the highest amount of interfering compounds, identified as phytadienes and sugar derivative products, which may co-elute with FA. An additional step based on solid-phase extraction to produce clean samples was proposed. This procedure is simple and gives good recoveries for the FA fortified samples. Additionally, structural characterization of 16:1 trans-3 was conducted by covalent adduct chemical ionization tandem mass spectrometry.

Comparison of extraction and derivatization methods for fatty acid analysis in solid environmental matrixes

This study involved comparison of different extraction and derivatization methods for determining FAs in soil and in four highly organic matrixes (cattle manure, pig slurry, compost, and vermicompost), by application of a multifactor categorical design. Although some studies have been carried out regarding the application of FA analysis to highly organic matrixes, comparison and verification are still required to test which methods of extraction and derivatization of FAs function best for these matrixes. We compared three extraction methods (one in which the same extraction mixture as used in the Folch method was employed, a modification of the Bligh and Dyer method, and a microwave-assisted extraction) and two derivatization procedures (alkaline methanolysis and derivatization with trimethylsulfonium hydroxide (TMSH)). The highest yields of FAs belonging to different structural classes, and of individual FAs used as microbial biomarkers were obtained by application of the same extraction mixture as in the Folch method and use of TMSH as derivatization agent. These methods also involved a significant reduction in the complexity and time involved in sample preparation.

Comparison of three solid-phase extraction methods for fatty acid analysis of lipid fractions in tissues of marine bivalves

Objective of this study was to investigate the effect of using pre-packed Si (Si), manually packed silica hydrated with water (Si–H 2O) and pre-packed aminopropyl-bonded silica (NH 2), at various mass ratios of lipid to sorbent, on the recovery of polar lipids following the solid-phase extraction (SPE) of a standard mixture of lipids. We also applied SPE using these sorbents to the separation of lipids from oyster tissues and compared the fatty acid (FA) composition of each fraction. Recoveries of phospholipids after SPE using Si increased with an increasing ratio of lipid to sorbent. Although the use of Si–H 2O improved the recovery of polar lipid compared to that obtained on Si, the neutral lipid from gills and muscles of oyster showed distorted FA compositions presumably due to a leakage of polar lipids. Finally, NH 2 eluted with methanol provided good recoveries of phospholipids from the standard mixture; although polar lipids of oyster tissues showed a reduction in 20:4 n − 6 and MUFA likely due to the selective retention of acidic phospholipids.

1 Fatty Acid Profile in A Blood Drop Collected in 3- Day Old Infants: Comparisons with Adult Subjects and Correlations with Physiologic Parameters

Background: Long Chain Polyunsaturated (LCP) Fatty Acid (FA) status in the newborn may affect post natal growth and development, but data in newborns are limited due to difficulties in sample collection.Methods: A new method for FA analysis in a drop of whole blood absorbed on a strip of Chromatography Paper (Marangoni et al, Anal Biochem 2004;326:267) was applied to a population of 110 infants, by analyzing blood samples collected from the heel within 72 h after delivery (37–41 wks post-conceptional age).Results: Comparisons with data from an unrelated, healthy adult population (100 subjects), analyzed with the same technique, showed lower levels of linoleic acid (LA) and alpha-linolenic acid (ALA) together with higher LCP (mainly arachidonic acid, AA, and docosahexaenoic acid, DHA, 22:6 n-3) levels, and markedly higher proportions of > 22 C FA of all FA families in the newborns, revealing major differences in FA intake, metabolism and incorporation in lipid pools between the two groups. Differences in FA profiles occurred also within the newborns, in relation with 1. gender (higher LA in females) 2. gestational age, with lower AA and DHA levels in the highest decile (10 s) for post-conceptional age at birth (41.2 weeks, SD 0.1) compared to the others 3. birth weight, with higher DHA levels in the lowest (10 s) vs the highest (12 s) decile (%: 4.2, SD 0.4, vs 3.4, SD 1.0, Mann-Whitney U test: P = 0.002) and 4. maternal life style (higher 22:5 n-6/22:6 n-3 ratio in smoking vs non-smoking mothers).Conclusion: The new method of FA analysis provides valuable information on the FA status and biochemical features related to FA at very early stages of post natal development, an age that has not been adequately investigated so far.