Profiling free fatty acids in edible oils via magnetic dispersive extraction and comprehensive two-dimensional gas chromatography-mass spectrometry.

The analysis of free fatty acids (FFAs) in edible oils can provide important information for quality control and oil authentication. Herein, we report the comprehensive profiling of FFAs in edible oils via magnetic dispersive extraction combined with comprehensive two-dimensional gas chromatography-mass spectrometry (GC × GC-MS). A magnetic extractant was designed for dispersive extraction of FFAs. The extraction conditions were carefully optimized. To assess the extraction method, we first use the method for analysis of 7 targeted FFAs. The limits of detection range from 5.6 to 25.8 ng g-1, and the recoveries in oil samples are 81%-107%. We then performed comprehensive profiling of untargeted FFAs in oil by combining the extraction method with GC × GC-MS. A total of 64 FFAs were identified positively or putatively. The proposed method can provide FFA fingerprint data to guide the processing, storage and authentication of edible oils.

Preparation of Pinolenic Acid Concentrates from Pine Nut Oil Fatty Acids by Solvent Fractionation.

Pinolenic acid (PLA), which is a fatty acid (FA) exclusively found in the oils of edible pine nuts, has an appetite-suppression effect, thereby being effective to reduce body weight in humans. PLA concentrates would be suitable for use in functional foods and nutraceuticals due to the health benefits of PLA. PLA concentrates were prepared from free FA (FFA) obtained from pine nut oil using solvent fractionation. Siberian pine nut oil containing 18.3 wt% PLA was used as the starting material for the fractionation. The fractionation was performed in n-hexane at ultra-low temperatures down to -85°C. The PLA concentrates produced under the optimal conditions established in this study (temperature, -85°C; n-hexane-to-FFA ratio (v/w), 30:1; fractionation time, 36 h) contained 69.8 wt% PLA. The yield of PLA was 77.4 wt% of the initial PLA weight in the FFA. These results suggest that solvent fractionation is a more effective approach to prepare PLA concentrates with higher PLA contents at a particular yield of PLA than published methods using urea crystallization (e.g., PLA content = ~47 wt%, yield of PLA = ~77 wt%, Woo et al. (2016)) or lipase-catalyzed reactions (e.g., PLA content = ~30 wt%, yield of PLA = ~61 wt%, Lee et al. (2011)). The resulting PLA concentrates contained 11 of the 12 different species of FA present in the FFA, thereby indicating that the PLA concentrates prepared by solvent fractionation have more diverse FA profiles than those prepared by urea crystallization (e.g., 7 species of FA, Woo et al. (2016)).

Lipidomic profiling reveals free fatty acid alterations in plasma from patients with atrial fibrillation.

Atrial fibrillation (AF) is the most common cardiac arrhythmia, and its incidence is increasing worldwide. One method used to restore sinus rhythm is direct current cardioversion (DCCV). Despite the high success rate of DCCV, AF typically recurs within the first 2 weeks. However, our understanding of the pathophysiology of AF recurrence, incidence, and progression are highly limited. Lipidomic profiling was applied to identify altered lipids in plasma from patients with AF using ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry coupled with multivariate statistical analysis. Partial least-squares discriminant analysis revealed a clear separation between AF patients and healthy controls. The levels of several lipid species, including fatty acids and phospholipids, were different between AF patients and healthy controls, indicating that oxidative stress and inflammation are associated with the pathogenesis of AF. Similar patterns were also detected between recurrent and non-recurrent AF patients. These results suggest that the elevated saturated fatty acid and reduced polyunsaturated fatty acid levels in AF patients may be associated with enhanced inflammation and that free fatty acid levels may play a crucial role in the development and progression of AF.

A New Strategy to Refine Crude Indian Sardine Oil.

Current work aims to develop a refining process for removing phospholipids, free fatty acids (FFA), and metal ions without affecting n-3 polyunsaturated fatty acid (n-3 PUFA) esters present in the crude Indian sardine oil. Sardine oil was subjected to degumming with various acids (orthophosphoric acid, acetic acid, and lactic acid), conventional and membrane assisted deacidification using various solvents (methanol, ethanol, propanol and butanol) and bleaching with bleaching agents (GAC, activated earth and bentonite) and all the process parameters were further optimized. Degumming with 5%(w/w) ortho phosphoric acid, two stage solvent extraction with methanol at 1:1 (w/w) in each stage and bleaching with 3% (w/w) activated charcoal loading, at 80ºC for 10 minutes resulted in the reduction of phospholipid content to 5.66 ppm from 612.66 ppm, FFA to 0.56% from 5.64% with the complete removal of iron and mercury. Under these conditions, the obtained bleached oil showed an enhancement of n-3 PUFA from 16.39 % (11.19 Eicosapentaenoic acid (EPA) + 5.20 Docosahexaenoic acid (DHA)) to 17.91 % (11.81 EPA + 6.1 DHA). Replacing conventional solvent extraction with membrane deacidification using microporous, hydrophobic polytetrafluoroethylene membrane (PTFE), resulted in a lesser solvent residue (0.25% (w/w)) in the deacidified oil. In view of lack of reports on refining of n-3 PUFA rich marine oils without concomitant loss of n-3 PUFA, this report is significant.

Characterization of Palm Fatty Acid Distillate of Different Oil Processing Industries of Pakistan.

Palm fatty acid distillate (PFAD) is cheap and valuable byproduct of edible oil processing industries. This study was designed to characterize PFAD collected from different local oil industries. AOCS methods were used for the determination of physicochemical parameters such as free fatty acid (FFA), saponification value (SV), iodine value (IV), peroxide value (PV) and moisture content. Fatty acid composition was analyzed using GC-MS. Moisture content of samples was found to be in the range between 0.06-7.50%, while FFA, SV, IV and PV were found to be 65.70-94.68%, 195.23-219.64 mg KOH/g, 38.49- 63.10 g I2/100 g, 1.09-16.50 meq/kg, respectively. Mean value of fatty acids in PFAD was found as 0.04% lauric, 0.42% myristic, 41.25% palmitic, 7.29% stearic, 41.58% oleic, 8.95 % linoleic, 0.04% eicosenoic, 0.27% arachidic, 0.07% docosanoic, and 0.05% tetracosanoic acid, respectively. Palmitic acid was found as dominant saturated fatty acid 38.63-45.30%, whereas oleic acid C18:1 n9 was major unsaturated fatty acid 33.54-44.05 % in PFAD.

Production of biodiesel from vegetable oil and microalgae by fatty acid extraction and enzymatic esterification.

The aim of this work was to obtain biodiesel (methyl esters) from the saponifiable lipids (SLs) fraction of the microalga Nannochloropsis gaditana, whose biomass dry weight contains 12.1 wt% of these lipids. SLs were extracted from the microalga as free fatty acids (FFAs) for subsequent transformation to methyl esters (biodiesel) by enzymatic esterification. Extraction as FFAs rather than as SLs allows them to be obtained with higher purity. Microalgal FFAs were obtained by direct saponification of lipids in the biomass and subsequent extraction-purification with hexane. Esterification of FFAs with methanol was catalysed by lipase Novozym 435 from Candida antarctica. Stability studies of this lipase in the operational conditions showed that the esterification degree (ED) attained with the same batch of lipase remained constant over six reaction cycles (36 h total reaction time). The optimal conditions attained for 4 g of FFAs were 25°C, 200 rpm, methanol/FFA molar ratio of 1.5:1, Novozym 435/FFA ratio of 0.025:1 w/w and 4 h reaction time. In these conditions the ED attained was 92.6%, producing a biodiesel with 83 wt% purity from microalgal FFAs. Several experimental scales were tested (from 4 to 40 g FFAs), and in all cases similar EDs were obtained.

[Biological activity of lipids and photosynthetic pigments of Sargassum pallidum C. Agardh].

The biological activity of lipids and photosynthetic pigments of the kelp Sargassum pallidum (Turner) C. Agardh has been studied. Free fatty acids and their esters demonstrated considerable antimicrobial activity against bacteria (Staphylococcus aureus[ital] and Escherichia coli), yeast-like fungi (Candida albicans), and opportunistic pathogenic (Aspergilius niger) and phytopathogenic (Fusarium oxysporum, and Septoria glycines) fungi. Glyceroglycolipids and neutral lipids demonstrated moderate activity. Fucoxanthin and chlorophylls weakly suppressed the growth of microorganisms. None of the studied substances demonstrated activity against Ehrlich's carcinoma. It was shown that the season of weed harvesting affected both antimicrobial and hemolytic activities of different lipids due to changes in their fatty acid composition.

Rapid separation of free fatty acids in vegetable oils by capillary zone electrophoresis.

INTRODUCTION: Olive oil is a very important product to human health since it inhibits formation of free radicals, tumour growth, lesions and inflammatory substances. High concentrations of free fatty acids in olive oils results in lipid deterioration due to oxidative or hydrolytic rancidity. OBJECTIVE: To optimise an alternative capillary zone electrophoresis methodology, under ultraviolet indirect detection and to determine free fatty acids in edible vegetable oils without derivatisation steps in sample preparation. METHODS: The condition used consisted of 15 mm NaH2 PO4 -Na2 HPO4 at pH ~6.86, 4.0 mm of sodium dodecybenzenesulphonate, 8.3 mm of Brij 35®, 45% v/v of acetonitrile and 2.1% of 1-octanol, injection at 12.0 mbar of pressure for 4 s, +19 kV of applied voltage and indirect detection at 224 nm, within an analysis time of 11 min. RESULTS: The capillary zone electrophoresis method was successfully applied to determination of free fatty acids in extra virgin olive oil, virgin olive oil and soybean oil samples. The comparison with the official volumetric titration method showed no significant difference within the 95% confidence interval. CONCLUSION: The main advantage to the proposed method is the possibility to observe the individual amount of the free fatty acids, which would be useful for researchers interested in studying the effect of the free fatty acids profile on oxidative process in food.

Inactivation of hepatitis C virus infectivity by human breast milk.

BACKGROUND: Hepatitis C virus (HCV) is spread through direct contact with blood, although alternative routes of transmission may contribute to the global burden. Perinatal infection occurs in up to 5% of HCV-infected mothers, and presence of HCV RNA in breast milk has been reported. We investigated the influence of breast milk on HCV infectiousness. METHODS/RESULTS: Human breast milk reduced HCV infectivity in a dose-dependent manner. This effect was species-specific because milk from various animals did not inhibit HCV infection. Treatment of HCV with human breast milk did not compromise integrity of viral RNA or capsids but destroyed the lipid envelope. Fractionation of breast milk revealed that the antiviral activity is present in the cream fraction containing the fat. Proteolytic digestion of milk proteins had no influence on its antiviral activity, whereas prolonged storage at 4°C increased antiviral activity. Notably, pretreatment with a lipase inhibitor ablated the antiviral activity and specific free fatty acids of breast milk were antiviral. CONCLUSIONS: The antiviral activity of breast milk is linked to endogenous lipase-dependent generation of free fatty acids, which destroy the viral lipid envelope. Therefore, nursing by HCV-positive mothers is unlikely to play a major role in vertical transmission.

Removal of free fatty acid in Azadirachta indica (Neem) seed oil using phosphoric acid modified mordenite for biodiesel production.

In this study free fatty acids present in Azadirachta indica (Neem) oil were esterified with our synthesized phosphoric acid modified catalyst. During the esterification, the acid value was reduced from 24.4 to 1.8 mg KOH/g oil. Synthesized catalyst was characterized by NH(3) TPD, XRD, SEM, FTIR and TGA analysis. During phosphoric acid modification hydrophobic character and weak acid sites of the mordenite were increased, which lead to better esterification when compared to H-mordenite. A kinetic study demonstrates that the esterification reaction followed pseudo-first order kinetics. Thermodynamic studies were also done based on the Arrhenius model.

Removal of free fatty acid in waste frying oil by esterification with methanol on zeolite catalysts.

The removal of free fatty acid (FFA) in waste frying oil by esterification with methanol was conducted using various zeolite catalysts. The ZSM-5 (MFI), mordenite (MOR), faujasite (FAU), beta (BEA) zeolites, and silicalite were employed with different Si/Al molar ratio in the reaction. The effects of acidic properties and pore structure of the zeolite catalysts were discussed relating to the conversion of the FFA. The MFI zeolite induced an improvement of the removal efficiency of FFA by cracking to the FFA in its pore structure due to its narrow pore mouth. The catalytic activity for FFA removal was lowered with decreasing of acid strength of the zeolites. The strong acid sites of zeolites induced the high conversion of FFA comparatively. The acid strength and pore structure of acidic zeolites affected the catalytic activity in FFA removal.

Supercritical fluid extraction of grape seed oil and subsequent separation of free fatty acids by high-speed counter-current chromatography.

Supercritical fluid extraction (SFE) of grape seed oil was performed to study the effect of various parameters such as pressure, temperature and the particle size of the sample on the yield and composition of oil using an analytical-scale SFE system. Then the extraction was scaled up by 125 times using a preparative SFE system under the optimized conditions of high pressure (30-40 MPa) and low temperature (35-40 degrees C) with medium particle size (20-40 mesh). The maximum yield of the oil can reach 6.2% with pure supercritical CO2 and 4.0% more oil can be obtained by adding 10% of ethanol as modifier. The unsaturated fatty acids (UFSs) make up about 70% in the oil on the basis of free fatty acids. The grape seed oil was then subjected to separation and purification for free fatty acids after saponification by high-speed counter-current chromatography coupled with evaporative light scattering detection (ELSD). The separation of 1.0 g of oil can yield about 430 mg pure linoleic acid at 99% purity. The fatty acids were analyzed by HPLC-ELSD.

[Preparation of capillary gas chromatographic columns for separation of polar organic compounds by sol-gel method].

A sol-gel method was used in preparing capillary gas chromatographic columns with simplicity and rapidity. Due to the formation of the chemical bonds between the coating layer and the inner wall of the capillary, the thermal stability of the column was greatly increased. The sol-gel poly(dimethyl-siloxane) (PDMS) column was able to withstand temperatures as high as 400 degrees C. This column showed excellent separation of free fatty acids, amines, and other polar compounds. The sol-gel column also showed good repeatability of retention time on a single column as well as good repeatability of capacity factor, column efficiency, symmetry and McReynold's constant on several columns of the same type.

Identification of free fatty acids in maize protein bodies and purified alpha zeins by (13)C and (1)H nuclear magnetic resonance.

Zeins, the maize storage proteins, are the most abundant proteins in the corn endosperm, and are synthesized on the rough endoplasmatic reticulum and deposited in discrete organelles called protein bodies. Several authors, using circular dichroism and optical rotatory dispersion, have concluded that these proteins have a high alpha-helical content in alcoholic solution. In this work we have studied these proteins, within the protein bodies themselves and after extraction from the corn grains with 70% ethanol, using NMR (nuclear magnetic resonance) spectroscopy. We conclusively demonstrate the presence of free fatty acids within both the protein bodies and also in the alcohol extracted alpha zeins. We present evidence for a direct interaction between the free fatty acids and the alpha zein proteins within the protein body and suggest possible mechanisms by which such an association has arisen during the evolution of the maize endosperm.

Changes in the lipid content and fatty acid composition of 2nd-stage juveniles of Globodera rostochiensis after rehydration, exposure to the hatching stimulus and hatch.

The total lipid content of the dry weight of whole cysts and 2nd-stage juveniles (J2) of Globodera rostochiensis was 17.1% in dry cysts, 20.9% in cysts soaked in distilled water (DW), 20.3% in cysts that had been in potato root diffusate (PRD) for 7 days, 7.3% in cysts that had been in PRD for 28 days and 29.2% for hatched J2. The fatty acid composition of the total lipid did not differ between dry cysts, cysts in DW and cysts in PRD for 7 days. However, major differences in the fatty acid composition of all lipid classes were found between rehydrated cysts in PRD and freshly hatched J2. After hatching, the degree of saturation and the percentage of monounsaturated fatty acids decreased and the percentage of polyunsaturated fatty acids increased considerably, especially in the free fatty acid fraction, where C20:1 showed an 8-fold decrease and C20:4 a 33-fold increase. There was a difference in the fatty acid composition of cysts in PRD for 7 days and cysts in PRD for 28 days (after most of the J2 had hatched); with increased time in PRD the percentage of polyunsaturated fatty acids increased and the percentage of monounsaturated fatty acids decreased in all lipid classes. Differences in the fatty acid profiles between cysts in PRD for 28 days and hatched J2 were found mainly in the free fatty acid and the non-acidic phospholipid fractions. The free fatty acid fraction of the cysts was dominated by monounsaturated fatty acids (52%), whereas the same fraction of hatched J2 contained large amounts of polyunsaturated fatty acids (60%). These polyunsaturated fatty acids, especially C20:4, might play an important part in nematode-plant interactions during infection.

Effects of free fatty acids on the binding of bovine and human serum albumin with steroid hormones.

Recent studies have shown that, in addition to free steroid hormones, those bound to albumin in plasma may also be available to tissues. In this report, the effects of free fatty acids (FFA) on the binding of steroids to albumin were compared for the cases of bovine serum albumin (BSA) and human serum albumin (HSA). The apparent association constant, Ka, was estimated from the changes in the equilibrium partition coefficient of steroids between the aqueous/hexane phases caused by the addition of albumin to the aqueous phase. In the case of BSA, Ka for progesterone and testosterone increased upon binding of FFA (myristic, palmitic and stearic acid) to BSA and the maximum value of Ka for these steroids could be attained by 3--4 mol of FFA bound per mol BSA. Furthermore, the elution profiles of gel-filtration chromatography clearly showed that progesterone and testosterone are easily liberated from the steroid/BSA complexes and that FFA potentiates the binding of these steroids to BSA. In the case of HSA, the binding affinities of progesterone and testosterone were not greatly affected by bound FFA. On the other hand, the affinities of ethynylestradiol to both BSA and HSA were unaffected below 4 mol of FFA binding per mol.

Determination of plasma non-esterified fatty acids and triglyceride fatty acids by gas chromatography of their methyl esters after isolation by column chromatography on silica gel.

Non-esterified fatty acids (NEFA) and triglycerides were isolated from human plasma by column chromatography on silica gel. Eight principal fatty acids of each of these lipid classes were determined by gas chromatography of their methyl ester derivatives and quantified relative to multipoint standard curves. Within-day relative standard deviations for plasma non-esterified fatty acid and triglyceride fatty acid determinations were 2.4 and 3.2%, respectively. Day-to-day relative standard deviations for plasma non-esterified fatty acid and triglyceride fatty acid determinations were 1.4 and 1.1%, respectively. The total plasma concentration and the relative proportions of the eight non-esterified fatty acids determined by this method were significantly different from results obtained according to two generally accepted methods for direct plasma non-esterified fatty acid determination without a specific isolation step. These comparisons suggested that considerable fatty acid ester lipid hydrolysis occurred during these direct determination procedures, and that this hydrolysis resulted in 3-fold overestimation of plasma NEFA content by those methods. Measured levels of arachidonic acid are substantially overestimated by these direct determination methods in which non-esterified fatty acids are not isolated before derivatization.

Antitumor components isolated from the Chinese herbal medicine Coix lachryma-jobi.

The seeds of the gramineous plant Coix lachryma-jobi L. var. ma-yuen Stapf are a traditional Chinese medicine possessing antitumor activity. An acetone extract of the seeds was fractionated by the aqueous alkali method and by silica gel column chromatography in order to identify the antitumor components. Antitumor activity, as assayed by an in vivo growth inhibition test on a transplantable mouse tumor, was attributed to an acidic fraction. Infrared spectroscopy and gas-liquid chromatography showed that this acidic fraction was composed of four free fatty acids: palmitic, stearic, oleic, and linoleic acids.

Abnormalities in epidermal lipid metabolism in patients with atopic dermatitis.

Atopic dermatitis is an inflammatory skin disease characterized by dryness and itch of the skin. In this study, we measured the phospholipid content and the fatty acid pattern of lesional and lesion-free epidermal keratome biopsies on 15 patients. For comparison, epidermal biopsies were obtained from healthy individuals undergoing plastic surgery. The phospholipid content of atopic epidermis was nearly twice as high as in healthy epidermis. Monounsaturated fatty acids in the phosphoglycerides were significantly increased (p less than 0.001) and n-6 fatty acids were significantly decreased (p less than 0.001) in lesional atopic epidermis compared to lesion-free epidermis. The content of esterified arachidonic acid in phosphatidylcholine from lesional epidermis was only 49% of that found in healthy epidermis (p less than 0.001). The content of free arachidonic acid was 47% higher (p less than 0.05), whereas the content of free long-chain saturated fatty acids was decreased by 29% (p less than 0.01), in lesional compared to lesion-free atopic epidermis. The disease severity, calculated as an arbitrary index, correlated inversely with the n-6 fatty acid content of lesion-free atopic epidermis (r = -0.89, p less than 0.001). Our findings suggest that atopic epidermis is characterized by an increased activity of phospholipase A2 and an incomplete transformation of phospholipids into other lipid classes.