Simultaneous quantification of total eicosapentaenoic acid, docosahexaenoic acid and arachidonic acid in plasma by high‐performance liquid chromatography–tandem mass spectrometry

Abstract

A method for the simultaneous quantification of eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and arachidonic acid (AA) in human plasma by HPLC-tandem mass spectrometry (HPLC-MS/MS) was developed and validated. Free and esterified forms of fatty acids were hydrolysed from plasma samples in the presence of an internal standard and subjected to liquid-liquid extraction. The chromatographic run time was 3.5 min per sample. The assay was linear from 0.5 to 300 mg/L (r(2) > 0.997, n = 18). Based on matrix addition, accuracy deviation was <15%, except for AA at 10 mg/L (30-90%), whereas precision was <8% for all fatty acids studied. The method was applied to the measurement of these omega-3 fatty acids in a fish oil supplement study with healthy volunteers. Healthy males (n = 4) were administered a supplement containing 465 mg EPA and 375 mg DHA per capsule (Omacor®). A dose of two capsules was given daily over a 4 week period. Pre-treatment concentrations varied between subjects for EPA (17-68 mg/L), DHA (36-63 mg/L) and AA (121-248 mg/L). During the dosing period EPA increased 460-480% from the baseline concentration, while DHA increased 150-160%. The EPA-AA ratio increased from 0.07-0.56 to 0.3-3.1 after 4 weeks of dosing. In conclusion, the method described could be suitable for monitoring EPA, DHA and AA in clinical studies that may aid in achieving optimal concentrations of these fatty acids in patients who could be at risk of sudden cardiac death.

Omega-3 Fatty Acids Modulate Weibel-Palade Body Degranulation and Actin Cytoskeleton Rearrangement in PMA-Stimulated Human Umbilical Vein Endothelial Cells

“…However, a greater number of cells stained positively for vWF when pre-treated with DHA or EPA prior to stimulation with PMA, compared to cells that were incubated with PMA alone (Figure 3a,c; paired t -test, p < 0.05, n = 4). The concentrations of LC n -3 PUFAs used in this study (75 and 120 μM) were within the physiological plasma concentration range for DHA (110–192 μM) and EPA (56–225 μM) in healthy individuals [29]. Interestingly, the n -6 PUFA, arachidonic acid (AA) attenuated WPB degranulation to a similar level to that observed for EPA and DHA, whereas shorter-chain fatty acids, oleic acid (C18:1 n -9) and linoleic acid (C18:2 n -6) were not different to PMA-stimulated cells (data not shown).…”

Evidence of altered phosphatidylcholine metabolism in Alzheimer's disease

“…To investigate whether the observations in individual PC species were linked to fatty acid side chains, total plasma ω-fatty acid profiles for arachidonic acid (20:4n6, AA), docosahexaenoic acid (22:6n3, DHA), and eicosapentaenoic acid (20:5n3, EPA) were obtained using a previously validated method (Salm et al, 2011). Analysis was performed on a group of 30 patients (10 per group; control, AD, and MCI subjects) that was age and gender matched across all 3 groups.…”

“…Analysis of total AA, DHA, and EPA was completed using a validated extraction and LC-MS method (Salm et al, 2011). Adjustments to the method are elaborated upon in the Supplementary Information.…”

A liquid chromatography–tandem mass spectrometry method to measure fatty acids in biological samples

“…Several LC-MS/MS protocols have previously been published for FA analysis [23,[25][26][27], some of which use linear ion trap quadrupole tandem mass spectrometers, often reporting relatively high LOD (up to 1ug/ml, [26])…”

“…while others have restricted methodology to EPA only [25] or EPA, DHA and AA [27]. Of note is the method of…”