Quantitation of Underivatized Omega-3 and Omega-6 Fatty Acids in Foods by HPLC and Charged Aerosol Detection

Abstract

The omega fatty acids are a group of compounds that include essential n-3 (omega-3, e.g., a-linolenic acid [ALA]), n-6 (omega-6, e.g., linoleic and arachidonic acids), and nonessential n-9, (omega-9, e.g., oleic and erucic acids) analytes. The omega-3 fatty acids, which also include eicosapentanoic acid (EPA) and docosahexanoic acid (DHA), are required for normal growth. Their consumption is purported to have a number of health benefits: e.g., cancer prevention, cardiovascular disease prevention, and improved immune function. Although both omega-3 and -6 fatty acids can give rise to eicosanoid-signaling molecules (prostaglandins, prostacyclins, thromboxanes, and leukotrienes), the omega-6 eicosanoids are generally pro-inflammatory and may play a role in cardiovascular disease, high blood pressure, and arthritis. It appears that the amounts and balance of omega fatty acids in a person’s diet affect their eicosanoid-controlled functions. A proper balance of omega fatty acids in the diet is important. Traditionally, omega fatty acids are measured using gas chromatography (GC). For foods, analytes are extracted from the samples prior to hydrolysis to release the fatty acids from their triglycerides, then converted to their volatile methyl esters prior to analysis by GC. This approach is tedious, time-consuming, and the high temperatures can affect polyunsaturated fatty acid stability. The Thermo Scientific Dionex Corona charged aerosol detector provides a univeral massbased approach that is sensitive, has a wide dynamic range, and has a major advantage in that all nonvolative analytes give similar response independent of chemical structure. No derivatization is required, and unlike UV detection, the analyte does not need to contain a chromophore. Presented here is a simple and direct high-performance liquid chromatography and charged aerosol detection (HPLC-charged-aerosol-detector) method that can be used to measure omega-3, -6 and -9 fatty acids in traditional and commercially produced meat, fish, and oils, as well as over-the-counter supplements. Quantitation of Underivatized Omega-3 and Omega-6 Fatty Acids in Foods by HPLC and Charged Aerosol Detection Ian Acworth, Marc Plante, Bruce Bailey, and Christopher Crafts Thermo Fisher Scientific, Chelmsford, MA, USA Introduction The common determination of omega lipids in foods comprises several steps, including extraction, hydroysis, and derivitization for measurement by GC. GC works well as a standard analytical tool for these determinations but requires alteration of the sample, and it can also adversely affect temperature-sensitive functional groups on specialized lipids. HPLC with ultraviolet detection requires use of low wavelengths, which limits solvent selection and increases the likelihood of matrix interference. Shown here is a reversed-phase HPLC method for the determination of omega fatty acids in oil/food samples using a dual-gradient method and charged aerosol detection. This combination enables determination of many fatty acids in a single analysis, and without the sample derivatization that is required for GC analysis. Several fatty acids were analyzed, including six omega-3 fatty acids (stearidonic acid [SDA], eicosapentanoic acid [EPA], a-linolenic acid [ALA], docosahexanoic acid [DHA], docosapentanoic acid [DPA], eicosatrienoic acid [ETA]), five omega-6 fatty acids (γ-linolenic acid [GLA], arachidonic acid [Arach.], linoleic acid [LLA], adrenic acid, and eicosadienioic acid [EDA]), and two omega-9 fatty acids (oleic and erucic acids). An omega-7 fatty acid, 9E,14Z-conjugated linoleic acid (CLA), was also included due to its cited importance as a nutrient.1,2 Charged aerosol detection is mass sensitive and can be added to HPLC or ultra HPLC (UHPLC) platforms. The detector provides the most consistent response for all nonvolatile and some semivolatile analytes of all HPLC detection techniques.3 It works by charging particles as shown in Figure 1, and is not dependent on light scattering, which has large variability and generally lower sensitivity. Charged aerosol detection has been successfully used to characterize lipids of all classification,4 including phospholipids (reversed phase5 and normal phase6,7), acylglycerides, phytosterols, free fatty acids, and free fatty alcohols. This method complements the free fatty acids method, using higher specificity for these analytes and a Thermo Scientific Acclaim C30 reversed-phase column.