GTRS and 2D-NMR studies of alpha and gamma linolenic acids each containing the same H2C14-(H–CC–H)–C11H2–(H–CC–H)–C8H2 moiety

Authors: Schmidt, Walter; CHEN, FU - University Of Maryland; BROADHURST, LEIGH - University Of Maryland; Nguyen, Julie; Kim, Moon; Qin, Jianwei - Tony Qin; Chao, Kuanglin - Kevin Chao

Submitted to: Journal of Molecular Structure
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/11/2019
Publication Date: 6/19/2019
Citation: Schmidt, W.F., Chen, F., Broadhurst, L.C., Nguyen, J.K., Kim, M.S., Qin, J., Chao, K. 2019. GTRS and 2D-NMR studies of alpha and gamma linolenic acids each containing the same H2C14-(H–CC–H)–C11H2–(H–CC–H)–C8H2 moiety. Journal of Molecular Structure. 1196(11):258-270. https://doi.org/10.1016/j.molstruc.2019.06.046.
DOI: https://doi.org/10.1016/j.molstruc.2019.06.046

Interpretive Summary: Alpha-linolenic acid (ALA) and gamma-linolenic acid (GLA), two polyunsaturated fatty acids found in plant-based foods such as seeds, nuts, grains, and vegetables, are essential fats important to healthy body function and known for their anti-inflammatory properties. Public interest and consumption of these two fats as added (omega-3 and omega-6) food ingredients are at an all-time high due to health supplement trends and the growing popularity of products for “paleolithic” diets. As more products containing these two fats appear in the marketplace, faster and more accurate ingredient authentication is needed. Both ALA and GLA each have 18 carbons and 3 double bonds; their molecular structures differ only in where the first double bond is located. However, GLA is a less common and more expensive ingredient; substitution using ALA is easy to do, while detecting the substitution requires complex, labor-intensive gas chromatography analysis. Through a comprehensive analysis of the molecular structures of ALA and GLA by combining gradient temperature Raman spectroscopy (GTRS) and nuclear magnetic resonance spectroscopy, ARS scientists have provided a GTRS method that can rapidly identify the fatty acid contents in a mixed oil. This rapid, accurate, and cost-effective technology for identifying and authenticating fatty acid ingredients will help food and supplement processors ensure the safety and quality of their products and support customer confidence in those products.

Technical Abstract: The polyunsaturated fatty acids alpha linolenic acid ('-LN; n-3) and gamma-linolenic acid ('-LN; n-6) are structural analogs and contain the moiety [(H-C=C-H)-CH2]3. A combination of gradient temperature Raman (GTRS) and nuclear magnetic resonance (NMR) analysis shows the spectral signature of the H-C= and CH sites in this moiety are very clearly different. GTRS showed the C-C asymmetrical stretching vibrational mode is different for each double bond, plus '-LN had a unique fourth mode in the same spectral region. High field NMR experiments confirmed that the three C=C sites are not identical. The C=C spectra region has six spectral lines in two sets of three, with none of the two sets aligned between '-LN and''-LN. Two-dimensional NMR experiments (INADEQUATE, HSQC, HMBC) enabled full and unambiguous assignment of every chemical shift to individual proton and carbon sites. Long-range coupling between C11H2 and C6H2 confirmed the '-LN large-scale carbon backbone is quite rigid. In contrast, long-range coupling in '-LN was between C11H2 and C4H2. At C14 and C8 sites, each CH2 group is twisted +90° and -90° respectively (relative to C11Hb); down and up from the plane in '-LN, and up and down from the plane in '-LN. This structural difference between is fully sufficient to explain why the lipid metabolism of '-LN and '-LN differs biochemically. It also helps explains variations in enzymatic oxidation efficiencies carbon number preferences for n-3 vs. n-6 fatty acids.