Omega-3 Fatty Acid Therapy: A Review of Study Design Flaws, Quality, and Composition

Main Article Content

Jacob M. Hands, B.A. Leigh A Frame, Ph.D., M.H.S.

Abstract

There is marked heterogeneity in the clinical response to omega-3 fatty acid therapy with many authors documenting futility in large-scale trials, secondary re-analysis, and meta-analysis. The question of failure in the context of omega-3 therapy is multifactorial and complicated by the observation that fish intake has been broadly linked to significant risk reductions across a range of conditions. The question that remains is how can we resolve the discrepancy between pre-clinical evidence and epidemiology, which dually emphasize the benefit of omega-3 therapy against the limited success of large-scale clinical trials and smaller scale clinical studies that do not consistently report benefit and may even report harm, especially as it pertains to atrial fibrillation. We present three primary considerations that may clarify the supposed failures of omega-3 therapy: 1) correction for omega-3:omega-6 ratio and competition, 2) variation in the fatty acid composition and quality of omega-3 products, and 3) fundamental concerns pertaining to the omega-3 vehicle and its impact on omega-3 metabolism. While the predominant source of omega-3 therapy is supplements, they are typically not regulated prior to market like drugs and have significant variability in fatty acid composition, vehicle, oxidation, and quality control. Further, the individual response to omega-3 therapy is likely variable and dependent on ambient dietary conditions and inherited differences in endogenous desaturase activity, which has infrequently been accounted for in large studies. The net effect of these concerns should engender pharmaceutical and consumer companies alike to consider 1) refining trial design and 2) consider the role of oxidation in the failure of omega-3 products.

Keywords: Omega-3 Fatty Acid Therapy, Omega-3, Fatty Acid

Article Details

How to Cite
HANDS, Jacob M.; FRAME, Leigh A. Omega-3 Fatty Acid Therapy: A Review of Study Design Flaws, Quality, and Composition. Medical Research Archives, [S.l.], v. 12, n. 4, apr. 2024. ISSN 2375-1924. Available at: <https://esmed.org/MRA/mra/article/view/5273>. Date accessed: 27 may 2024. doi: https://doi.org/10.18103/mra.v12i4.5273.
Section
Review Articles

References

1. Bowman L, Mafham M, Stevens W, Haynes R, Aung T, Chen F, Buck G, Collins R, Armitage J; ASCEND Study Collaborative Group. ASCEND: A Study of Cardiovascular Events iN Diabetes: Characteristics of a randomized trial of aspirin and of omega-3 fatty acid supplementation in 15,480 people with diabetes. Am Heart J. 2018 Apr;198:135-144. doi: 10.1016/j.ahj.2017.12.006. Epub 2017 Dec 24. PMID: 29653635; PMCID: PMC5971211.,

2. Bhatt DL, Steg PG, Miller M, Brinton EA, Jacobson TA, Ketchum SB, Doyle RT Jr, Juliano RA, Jiao L, Granowitz C, Tardif JC, Ballantyne CM; REDUCE-IT Investigators. Cardiovascular Risk Reduction with Icosapent Ethyl for Hypertriglyceridemia. N Engl J Med. 2019 Jan 3;380(1):11-22. doi: 10.1056/NEJM oa1812792. Epub 2018 Nov 10. PMID: 30415628.

3. Nicholls SJ, Lincoff AM, Garcia M, Bash D, Ballantyne CM, Barter PJ, Davidson MH, Kastelein JJP, Koenig W, McGuire DK, Mozaffarian D, Ridker PM, Ray KK, Katona BG, Himmelmann A, Loss LE, Rensfeldt M, Lundström T, Agrawal R, Menon V, Wolski K, Manson JE, Cook NR, Lee IM, Christen W, Bassuk SS, Mora S, Gibson H, Gordon D, Copeland T, D'Agostino D, Friedenberg G, Ridge C, Bubes V, Giovannucci EL, Willett WC, Buring JE; VITAL Research Group. Vitamin D Supplements and Prevention of Cancer and Cardiovascular Disease. N Engl J Med. 2019 Jan 3;380(1):33-44. doi: 10.1056/NEJMoa1809944. Epub 2018 Nov 10. PMID: 30415629; PMCID: PMC6425757.,

4. Nissen SE. Effect of High-Dose Omega-3 Fatty Acids vs Corn Oil on Major Adverse Cardiovascular Events in Patients at High Cardiovascular Risk: The STRENGTH Randomized Clinical Trial. JAMA. 2020 Dec 8;324(22):2268-2280. doi: 10.1001/jama.2020 .22258. PMID: 33190147; PMCID: PMC7667577.

5. Kalstad AA, Myhre PL, Laake K, Tveit SH, Schmidt EB, Smith P, Nilsen DWT, Tveit A, Fagerland MW, Solheim S, Seljeflot I, Arnesen H; OMEMI Investigators. Effects of n-3 Fatty Acid Supplements in Elderly Patients After Myocardial Infarction: A Randomized, Controlled Trial. Circulation. 2021 Feb 9;143(6):528-539. doi: 10.1161/CIRCULATIO NAHA.120.052209. Epub 2020 Nov 15. PMID : 33191772.).

6. Shen S, Gong C, Jin K, Zhou L, Xiao Y, Ma L. Omega-3 Fatty Acid Supplementation and Coronary Heart Disease Risks: A Meta-Analysis of Randomized Controlled Clinical Trials. Front Nutr. 2022 Feb 3;9:809311. doi: 10.3389/fnut.2022.809311. PMID: 35187035; PMCID: PMC8850984.

7. Khan SU, Lone AN, Khan MS, Virani SS, Blumenthal RS, Nasir K, Miller M, Michos ED, Ballantyne CM, Boden WE, Bhatt DL. Effect of omega-3 fatty acids on cardiovascular outcomes: A systematic review and meta-analysis. EClinicalMedicine. 2021 Jul 8;38:100 997. doi: 10.1016/j.eclinm.2021.100 997.PMI D: 34505026; PMCID: PMC8413259.

8. Bernasconi AA, Wiest MM, Lavie CJ, Milani RV, Laukkanen JA. Effect of Omega-3 Dosage on Cardiovascular Outcomes: An Updated Meta-Analysis and Meta-Regression of Interventional Trials. Mayo Clin Proc. 2021 Feb;96(2):304-313. doi: 10.1016/j.mayocp.20 20.08.034. Epub 2020 Sep 17. PMID: 32951855.

9. Abdelhamid AS, Brown TJ, Brainard JS, Biswas P, Thorpe GC, Moore HJ, Deane KH, Summerbell CD, Worthington HV, Song F, Hooper L. Omega-3 fatty acids for the primary and secondary prevention of cardiovascular disease. Cochrane Database Syst Rev. 2020 Feb 29;3(3):CD003177. doi: 10.1002/146518 58.CD003177.pub5. PMID: 32114706; PMCI D: PMC7049091.Rizos EC, Ntzani EE, Bika E, Kostapanos MS, Elisaf MS. Association between omega-3 fatty acid supplementation and risk of major cardiovascular disease events: a systematic review and meta-analysis. JAMA. 2012 Sep 12;308(10):1024-33. doi: 10.1001/2012.jama.11374. PMID: 22968891.

10. Maki KC, Palacios OM, Bell M, Toth PP. Use of supplemental long-chain omega-3 fatty acids and risk for cardiac death: An updated meta-analysis and review of research gaps. J Clin Lipidol. 2017 Sep-Oct;11(5):1152-1160.e2. doi: 10.1016/j.jacl.2017.07.010. Epu b 2017 Aug 2. PMID: 28818347.)

11. Curfman G. Omega-3 Fatty Acids and Atrial Fibrillation. JAMA. 2021;325(11):1063. doi:10.1001/jama.2021.2909. Huh JH, Jo SH. Omega-3 fatty acids and atrial fibrillation. Korean J Intern Med. 2023;38(3):282-289. doi:10.3904/kjim.2022.266

12. Lombardi M, Carbone S, Del Buono MG, Chiabrando JG, Vescovo GM, Camilli M, Montone RA, Vergallo R, Abbate A, Biondi-Zoccai G, Dixon DL, Crea F. Omega-3 fatty acids supplementation and risk of atrial fibrillation: an updated meta-analysis of randomized controlled trials. Eur Heart J Cardiovasc Pharmacother. 2021 Jul 23;7(4):e69-e70. doi: 10.1093/ehjcvp/pvab00 8. PMID: 33910233; PMCID: PMC8302253.

13. Gencer B, Djousse L, Al-Ramady OT, Cook NR, Manson JE, Albert CM. Effect of Long-Term Marine ɷ--3 Fatty Acids Supplementation on the Risk of Atrial Fibrillation in Randomized Controlled Trials of Cardiovascular Outcomes: A Systematic Review and Meta-Analysis. Circulation. 2021 Dec 21;144(25):1981-1990. doi: 10.1161/CIR CULATIONAHA.121.055654. Epub 2021 Oct 6. PMID: 34612056; PMCID: PMC9109217.

14. Garg PK, Guan W, Nomura S, et al. Plasma ω-3 and ω-6 PUFA Concentrations and Risk of Atrial Fibrillation: The Multi-Ethnic Study of Atherosclerosis. J Nutr. 2021;151(6): 1479-1486. doi:10.1093/jn/nxab016

15. Macchia A, Grancelli H, Varini S, Nul D, Laffaye N, Mariani J, Ferrante D, Badra R, Figal J, Ramos S, Tognoni G, Doval HC; GESICA Investigators. Omega-3 fatty acids for the prevention of recurrent symptomatic atrial fibrillation: results of the FORWARD (Randomized Trial to Assess Efficacy of PUFA for the Maintenance of Sinus Rhythm in Persistent Atrial Fibrillation) trial. J Am Coll Cardiol. 2013 Jan 29;61(4):463-468. doi: 10.1016/j.jacc.2012.11.021. Epub 2012 Dec 19. PMID: 23265344.

16. Zhang B, Xiong K, Cai J, Ma A. Fish Consumption and Coronary Heart Disease: A Meta-Analysis. Nutrients. 2020;12(8):2278. Published 2020 Jul 29. doi:10.3390/nu12082278

17. Giosuè A, Calabrese I, Lupoli R, Riccardi G, Vaccaro O, Vitale M. Relations between the Consumption of Fatty or Lean Fish and Risk of Cardiovascular Disease and All-Cause Mortality: A Systematic Review and Meta-Analysis. Adv Nutr. 2022 Oct 2;13(5):1554-1565. doi: 10.1093/advances/nmac006. PMID: 35108375; PMCID: PMC9526843.)

18. Harris WS, Tintle NL, Etherton MR, Vasan RS. Erythrocyte long-chain omega-3 fatty acid levels are inversely associated with mortality and with incident cardiovascular disease: The Framingham Heart Study [published correction appears in J Clin Lipidol. 2020 Sep - Oct;14(5):740]. J Clin Lipidol. 2018;12(3):718 -727.e6. doi:10.1016/j.jacl.2018.02.010

19. Kleber ME, Delgado GE, Lorkowski S, März W, von Schacky C. Omega-3 fatty acids and mortality in patients referred for coronary angiography. The Ludwigshafen Risk and Cardiovascular Health Study. Atherosclerosis. 2016 Sep;252:175-181. doi: 10.1016/j.atheros clerosis.2016.06.049. Epub 2016 Jul 1. PMID: 27397734.

20. Chen GC, Yang J, Eggersdorfer M, Zhang W, Qin LQ. N-3 long-chain polyunsaturated fatty acids and risk of all-cause mortality among general populations: a meta-analysis. Sci Rep. 2016;6:28165. Published 2016 Jun 16. doi:10.1038/srep28165

21. Harris WS, Del Gobbo L, Tintle NL. The Omega-3 Index and relative risk for coronary heart disease mortality: Estimation from 10 cohort studies. Atherosclerosis. 2017 Jul;262: 51-54. doi: 10.1016/j.atherosclerosis.2017.05. 007. Epub 2017 May 6. PMID: 28511049

22. Harris WS, Von Schacky C. The Omega-3 Index: a new risk factor for death from coronary heart disease? Prev Med. 2004 Jul;3 9(1):212-20. doi: 10.1016/j.ypmed.2004.02.0 30. PMID: 15208005.

23. Adili R, Hawley M, Holinstat M. Regulation of platelet function and thrombosis by omega-3 and omega-6 polyunsaturated fatty acids. Prostaglandins Other Lipid Mediat. 2018;139:10-18. doi:10.1 016/j.prostaglandins.2018.09.005

24. Bagger H, Hansson M, Kander T, Schött U. Synergistic platelet inhibition between Omega-3 and acetylsalicylic acid dose titration; an observational study. BMC Complement Med Ther. 2020;20(1):204. Published 2020 Jul 2. doi:10.1186/s12906-020-02990-9

25. Wojenski CM, Silver MJ, Walker J. Eicosapentaenoic acid ethyl ester as an antithrombotic agent: comparison to an extract of fish oil. Biochim Biophys Acta. 1991 Jan 4;1081(1):33-8. doi: 10.1016/0005-2760(9 1)90246-e. PMID: 1991153.

26. Natto, Z.S., Yaghmoor, W., Alshaeri, H.K. et al. Omega-3 Fatty Acids Effects on Inflammatory Biomarkers and Lipid Profiles among Diabetic and Cardiovascular Disease Patients: A Systematic Review and Meta-Analysis. Sci Rep 9, 18867 (2019). https://doi.org/10.1038/s41598-019-54535-x).

27. Li J, Guasch-Ferré M, Li Y, Hu FB. Dietary intake and biomarkers of linoleic acid and mortality: systematic review and meta-analysis of prospective cohort studies. Am J Clin Nutr. 2020 Jul 1;112(1):150-167. doi: 10.1093/ajcn/ nqz349. PMID: 32020162; PMCID: PMC7326588.

28. Mousavi SM, Jalilpiran Y, Karimi E, Aune D, Larijani B, Mozaffarian D, Willett WC, Esmaillzadeh A. Dietary Intake of Linoleic Acid, Its Concentrations, and the Risk of Type 2 Diabetes: A Systematic Review and Dose-Response Meta-analysis of Prospective Cohort Studies. Diabetes Care. 2021 Sep;44(9 ):2173-2181. doi: 10.2337/dc21-0438. Epub 2021 Aug 20. PMID: 34417277.

29. Marklund M, Wu JHY, Imamura F, et al. Biomarkers of Dietary Omega-6 Fatty Acids and Incident Cardiovascular Disease and Mortality. Circulation. 2019;139(21):2422-2436. doi:10.1161/CIRCULATIONAHA.118.038908.

30. Simopoulos AP. The importance of the ratio of ɷ-6:ɷ-3 essential fatty acids. Biomed Pharmacother. 2002 Oct;56(8):365-79. doi: 10 .1016/s0753-3322(02)00253-6. PMID: 12442909

31. Zhang Y, Sun Y, Yu Q, et al. Higher ratio of plasma omega-6/omega-3 fatty acids is associated with greater risk of all-cause, cancer, and cardiovascular mortality: a population-based cohort study in UK Biobank. Preprint. medRxiv. 2024;2023.01.16.2328463 1. Published 2024 Jan 10. doi:10.1101/2023.0 1.16.23284631

32. DiNicolantonio JJ, O'Keefe J. The Importance of Maintaining a Low Omega-6/Omega-3 Ratio for Reducing the Risk of Autoimmune Diseases, Asthma, and Allergies. Mo Med. 2021;118(5):453-459.

33. Calder PC, Campoy C, Eilander A, Fleith M, Forsyth S, Larsson PO, Schelkle B, Lohner S, Szommer A, van de Heijning BJM, Mensink RP. A systematic review of the effects of increasing arachidonic acid intake on PUFA status, metabolism and health-related outcomes in humans. Br J Nutr. 2019 Jun;121( 11):1201-1214. doi: 10.1017/S000711451900 0692. Epub 2019 May 27. PMID: 31130146.

34. Djuricic I, Calder PC. Beneficial Outcomes of Omega-6 and Omega-3 Polyunsaturated Fatty Acids on Human Health: An Update for 2021. Nutrients. 2021;13(7):2421. Published 2021 Jul 15. doi:1 0.3390/nu13072421)

35. Harris WS. The Omega-6:Omega-3 ratio: A critical appraisal and possible successor. Prostaglandins, Leukotrienes, and Essential Fatty Acids. 2018 May;132:34-40. DOI: 10.10 16/j.plefa.2018.03.003. PMID: 29599053.)

36. Petrović-Oggiano G, Debeljak-Martačić J, Ranković S, et al. The Effect of Walnut Consumption on n-3 Fatty Acid Profile of Healthy People Living in a Non-Mediterranean West Balkan Country, a Small Scale Randomized Study. Nutrients. 2020;12( 1):192. Published 2020 Jan 10. doi:10.3390/n u12010192

37. Kuhnt K, Weiß S, Kiehntopf M, Jahreis G. Consumption of echium oil increases EPA and DPA in blood fractions more efficiently compared to linseed oil in humans. Lipids Health Dis. 2016;15:32. Published 2016 Feb 18.doi:10.1186/s12944-016-0199-2

38. Greupner T , Kutzner L , Pagenkopf S , et al. Effects of a low and a high dietary LA/ALA ratio on long-chain PUFA concentrations in red blood cells. Food Funct. 2018;9(9):4742-4754. doi:10.1039/c8fo00735g

39. Prasad P, Anjali P, Sreedhar RV. Plant-based stearidonic acid as sustainable source of omega-3 fatty acid with functional outcomes on human health. Crit Rev Food Sci Nutr. 2021;61(10):1725-1737. doi:10.1080/10 408398.2020.1765137

40. Blasbalg TL, Hibbeln JR, Ramsden CE, Majchrzak SF, Rawlings RR. Changes in consumption of omega-3 and omega-6 fatty acids in the United States during the 20th century. Am J Clin Nutr. 2011;93(5):950-962. doi:10.3945/ajcn.110.006643)

41. Dayton S, Hashimoto S, Pearce ML. Influence of a diet high in unsaturated fat upon composition of arterial tissue and atheromata in man. Circulation. 1965 Dec;32( 6):911-24. doi: 10.1161/01.cir.32.6.911. PMID : 5845249

42. Munakata M, Nishikawa M, Togashi N, Nio E, Kobayashi Y, Omura K, Haginoya K, Tanaka S, Abe T, Hishinuma T, Chida N, Tsuchiya S, Onuma A. The nutrient formula containing eicosapentaenoic acid and docosahexaenoic acid benefits the fatty acid status of patients receiving long-term enteral nutrition. Tohoku J Exp Med. 2009 Jan;217(1): 23-8. doi: 10.1620/tjem.217.23. PMID: 19155604.).

43. Clark KJ, Makrides M, Neumann MA, Gibson RA. Determination of the optimal ratio of linoleic acid to alpha-linolenic acid in infant formulas. J Pediatr. 1992 Apr;120(4 Pt 2):S151 -8. doi: 10.1016/s0022-3476(05)81250-8. PMI D: 1348533)

44. Mathias RA, Fu W, Akey JM, Ainsworth HC, Torgerson DG, Ruczinski I, et al. (2012) Adaptive Evolution of the FADS Gene Cluster within Africa. PLoS ONE 7(9): e44926. https://doi.org/10.1371/journal.pone.0044926

45. Conway MC, McSorley EM, Mulhern MS, Strain JJ, van Wijngaarden E, Yeates AJ. Influence of fatty acid desaturase (FADS) genotype on maternal and child polyunsaturated fatty acids (PUFA) status and child health outcomes: a systematic review. Nutr Rev. 2020 Aug 1;78(8):627-646. doi: 10.1 093/nutrit/nuz086. PMID: 31943072; PMCID: PMC7868964.

46. Mathias, R.A., Sergeant, S., Ruczinski, I. et al. The impact of FADS genetic variants on ω6 polyunsaturated fatty acid metabolism in African Americans. BMC Genet 12, 50 (2011). https://doi.org/10.1186/1471-2156-12-50

47. Chilton FH, Manichaikul A, Yang C, et al. Interpreting Clinical Trials With Omega-3 Supplements in the Context of Ancestry and FADS Genetic Variation. Front Nutr. 2022;8:8 08054. Published 2022 Feb 8. doi:10.3389/fn ut.2021.808054

48. Elagizi A, Lavie CJ, O'Keefe E, Marshall K, O'Keefe JH, Milani RV. An Update on Omega-3 Polyunsaturated Fatty Acids and Cardiovascular Health. Nutrients. 2021;13(1): 204. Published 2021 Jan 12. doi:10.3390/nu1 3010204

49. Musa-Veloso K, Binns MA, Kocenas A, Chung C, Rice H, Oppedal-Olsen H, Lloyd H, Lemke S. Impact of low v. moderate intakes of long-chain n-3 fatty acids on risk of coronary heart disease. Br J Nutr. 2011 Oct;106(8):1129 -41. doi: 10.1017/S0007114511001644. Epub 2011 May 31. PMID: 21736820

50. Sherratt SCR, Libby P, Budoff MJ, Bhatt DL, Mason RP. Role of Omega-3 Fatty Acids in Cardiovascular Disease: the Debate Continues. Curr Atheroscler Rep. 2023;25(1):1 -17. doi:10.1007/s11883-022-01075-x)

51. Dempsey M, Rockwell MS, Wentz LM. The influence of dietary and supplemental omega-3 fatty acids on the omega-3 index: A scoping review. Front Nutr. 2023;10:1072653. Published 2023 Jan 19. doi:10.3389/fnut.202 3.1072653

52. von Schacky C. Omega-3 index and cardiovascular health. Nutrients. 2014;6(2):79 9-814. Published 2014 Feb 21. doi:10.3390/n u6020799

53. von Schacky, C., Kuipers, R.S., Pijl, H. et al. Omega-3 fatty acids in heart disease—why accurately measured levels matter. Neth Heart J 31, 415–423 (2023). https://doi.org/10.1007/s12471-023-01759-2

54. Stanton, A.V., James, K., Brennan, M.M. et al. Omega-3 index and blood pressure responses to eating foods naturally enriched with omega-3 polyunsaturated fatty acids: a randomized controlled trial. Sci Rep 10, 15444 (2020). https://doi.org/10.1038/s41598-020-71801-5

55. Clemens von Schacky, William S. Harris, Cardiovascular benefits of omega-3 fatty acids, Cardiovascular Research, Volume 73, Issue 2, January 2007, Pages 310–315, https://doi.org/10.1016/j.cardiores.2006.08.019)

56. Rittenhouse M, Sambuughin N, Deuster P. Optimization of Omega-3 Index Levels in Athletes at the US Naval Academy: Personalized Omega-3 Fatty Acid Dosage and Molecular Genetic Approaches. Nutrients . 2022;14(14):2966. Published 2022 Jul 20. doi:10.3390/nu14142966)

57. DiNicolantonio J, O'Keefe JH. The Flaws of Recent Omega-3 Clinical Trials Should Not Prevent Their Use. Mo Med. 2021;118(4):322.

58. Elagizi A, Lavie CJ, O'Keefe E, Marshall K, O'Keefe JH, Milani RV. An Update on Omega-3 Polyunsaturated Fatty Acids and Cardiovascular Health. Nutrients. 2021;13(1): 204. Published 2021 Jan 12.) doi:10.3390/nu13010204).

59. Albert BB, Derraik JG, Cameron-Smith D, et al. Fish oil supplements in New Zealand are highly oxidized and do not meet label content of n-3 PUFA [published correction appears in Sci Rep. 2016 Nov 07;6:35092]. Sci Rep. 2015;5:7928. Published 2015 Jan 21. doi:10.1 038/srep07928.

60. Hands JM, Anderson ML, Cooperman T, Frame LA. A Multi-Year Rancidity Analysis of 72 Marine and Microalgal Oil Omega-3 Supplements. J Diet Suppl. 2023 Sep 15:1-12. doi: 10.1080/19390211.2023.2252064. Epub ahead of print. PMID: 37712532.

61. Lovaza (Omacor) capsules, prescribing information. Liberty Corner, NJ: Reliant; 2005. Revised August 2007.)

62. Shramko VS, Polonskaya YV, Kashtanova EV, Stakhneva EM, Ragino YI. The Short Overview on the Relevance of Fatty Acids for Human Cardiovascular Disorders. Biomolecules. 2020;10(8):1127. Published 2020 Jul 30. doi:1 0.3390/biom10081127

63. van Rooijen MA, Mensink RP. Palmitic Acid Versus Stearic Acid: Effects of Interesterification and Intakes on Cardiometabolic Risk Markers—A Systematic Review. Nutrients. 2020; 12(3):615. https://doi.org/10.3390/nu12030615

64. Donis N, Jiang Z, D'Emal C, et al. Regular Dietary Intake of Palmitate Causes Vascular and Valvular Calcification in a Rabbit Model. Front Cardiovasc Med. 2021;8:692184. Publis hed 2021 Jun 23. doi:10.3389/fcvm.2021.692184

65. Wang, Y., Qian, Y., Fang, Q. et al. Saturated palmitic acid induces myocardial inflammatory injuries through direct binding to TLR4 accessory protein MD2. Nat Commun 8, 13997 (2017). https://doi.org/10.1038/ncomms13997,

66. Diet-derived and diet-related endogenously produced palmitic acid: Effects on metabolic regulation and cardiovascular disease risk Palm oil and palmitic acid: a review on cardiovascular effects and carcinogenicity

67. Zong G, Li Y, Wanders A J, Alssema M, Zock P L, Willett W C et al. Intake of individual saturated fatty acids and risk of coronary heart disease in US men and women: two prospective longitudinal cohort studies BMJ 2016; 355 :i5796 doi:10.1136/bmj.i5796)

68. Kanner J. Dietary advanced lipid oxidation endproducts are risk factors to human health. Mol Nutr Food Res. 2007 Sep;51(9):1094-101. doi: 10.1002/mnfr.20060 0303. PMID: 17854006. Esterbauer H, Schaur RJ, Zollner H. Chemistry and biochemistry of 4-hydroxynonenal, malonaldehyde and related aldehydes. Free Radic Biol Med. 1991; 11(1):81-128. doi: 10.1016/0891-5849(91)901 92-6. PMID: 1937131.

69. Albert BB, Cameron-Smith D, Hofman PL, Cutfield WS. Oxidation of marine omega-3 supplements and human health. Biomed Res Int. 2013;2013:464921. doi:10.1155/2013/464921

70. Ottestad I, Nordvi B, Vogt G, Holck M, Halvorsen B, Brønner KW, Retterstøl K, Holven KB, Nilsson A, Ulven SM. Bioavailability of n-3 fatty acids from n-3-enriched foods and fish oil with different oxidative quality in healthy human subjects: a randomised single-meal cross-over study. J Nutr Sci. 2016 Oct 28;5:e43. doi: 10.1017/jns. 2016.34. PMID: 28620470; PMCID: PMC5465811.

71. Thiery J, Seidel D. Fish oil feeding results in an enhancement of cholesterol-induced atherosclerosis in rabbits. Atherosclerosis. 1987 Jan;63(1):53-6. doi: 10.1016/0021-9150( 87)90081-5. PMID: 3827970.

72. Nakagawa F, Morino K, Ugi S, Ishikado A, Kondo K, Sato D, Konno S, Nemoto K, Kusunoki C, Sekine O, Sunagawa A, Kawamura M, Inoue N, Nishio Y, Maegawa H. 4-Hydroxy hexenal derived from dietary n-3 polyunsaturated fatty acids induces anti-oxidative enzyme heme oxygenase-1 in multiple organs. Biochem Biophys Res Commun. 2014 Jan 17;443(3):991-6. doi: 10.1016/j.bbrc.2013.12.085. Epub 2013 Dec 19. PMID: 24361890.

73. Ishikado A, Morino K, Nishio Y, et al. 4-Hydroxy hexenal derived from docosahexaenoic acid protects endothelial cells via Nrf2 activation. PLoS One. 2013;8(7): e69415. Published 2013 Jul 23. doi:10.1371/j ournal.pone.0069415

74. Mackness B, Durrington P, McElduff P, Yarnell J, Azam N, Watt M & Mackness M (2003) Low paraoxonase activity predicts coronary events in the Caerphilly prospective study. Circulation 107, 2775– 2779.

75. Naruszewicz M, Wozny E, Mirkiewicz E, Nowicka G & Szostak WB (1987) The effect of thermally oxidized soya bean oil on metabolism of chylomicrons – increased uptake and degradation of oxidized chylomicrons in cultured mouse macrophages. Atherosclerosi s 66, 45 – 53.

76. Turner R., McLean C. H. & Silvers K. M. Are the health benefits of fish oils limited by products of oxidation? Nutr Res Rev 19, 53–62, 10.1079/nrr2006117 (2006).

77. Mackness MI, Arrol S, Abbott C & Durrington PN (1993) Protection of low-density lipoprotein against oxidative modification by high-density lipoprotein associated paraoxonase. Atherosclerosis 104, 129– 135.

78. Kota SK, Meher LK, Kota SK, Jammula S, Krishna SV, Modi KD. Implications of serum paraoxonase activity in obesity, diabetes mellitus, and dyslipidemia. Indian J Endocrinol Metab. 2013;17(3):402-412. doi:1 0.4103/2230-8210.111618).

79. Mason RP, Sherratt SCR. Omega-3 fatty acid fish oil dietary supplements contain saturated fats and oxidized lipids that may interfere with their intended biological benefits. Biochem Biophys Res Commun. 2017 Jan 29;483(1):425-429. doi: 10.1016/j.b brc.2016.12.127. Epub 2016 Dec 21. PMID: 28011269.

80. Rupp, T. P., Rupp, K. G., Alter, P., & Rupp, H. (2013). Replacement of Reduced Highly Unsaturated Fatty Acids (HUFA Deficiency) in Dilative Heart Failure: Dosage of EPA/DHA and Variability of Adverse Peroxides and Aldehydes in Dietary Supplement Fish Oils. Cardiology, 125(4), 223–231. doi:10.1159/00 0350656.

81. https://www.regulations.gov/document/FDA-2019-P-3424-0003

82. Huston J, Schaffner H, Cox A, et al. A Critical Review of Icosapent Ethyl in Cardiovascular Risk Reduction. Am J Cardiovasc Drugs. 2023;23(4):393-406. doi:10 .1007/s40256-023-00583-8 ).

83. Burri L, Hoem N, Banni S, Berge K. Marine omega-3 phospholipids: metabolism and biological activities. Int J Mol Sci. 2012;13(11):15401-15419. Published 2012 Nov 21. doi:10.3390/ijms131115401

84. Polvi, Sherilyn M., and Robert G. Ackman. "Atlantic salmon (Salmo salar) muscle lipids and their response to alternative dietary fatty acid sources." Journal of Agricultural and Food Chemistry 40.6 (1992): 1001-1007.

85. Chevalier L, Vachon A, Plourde M. Pharmacokinetics of Supplemental Omega-3 Fatty Acids Esterified in Monoglycerides, Ethyl Esters, or Triglycerides in Adults in a Randomized Crossover Trial. J Nutr. 2021;151 (5):1111-1118. doi:10.1093/jn/nxaa458

86. Yang LY, Kuksis A, Myher JJ. Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination. J Lipid Res. 1990 Jan;31(1):137-47. PMID: 2313198.

87. R. G. Ackman (1992). The absorption of fish oils and concentrates. , 27(11), 858–862. doi:10.1007/bf02535864,Yang, L.-Y., Kuksis, A., and Myher, J. (1990) J. Lipid Res. 31, 137-148.

88. Mahesar, S. A., Sherazi, S. T. H., Khaskheli, A. R., Kandhro, A. A., & Uddin, S. (2014). Analytical approaches for the assessment of free fatty acids in oils and fats. Anal. Methods, 6(14), 4956–4963. doi: 10.103 9/C4AY00344F)

89. Tullberg C, Vegarud G, Undeland I. Oxidation of marine oils during in vitro gastrointestinal digestion with human digestive fluids - Role of oil origin, added tocopherols and lipolytic activity. Food Chem. 2019 Jan 1;270:527-537. doi: 10.1016/j.foodc hem.2018.07.049. Epub 2018 Jul 10. PMID: 30174082.

90. Saw CL, Yang AY, Guo Y, Kong AN. Astaxanthin and omega-3 fatty acids individually and in combination protect against oxidative stress via the Nrf2-ARE pathway. Food Chem Toxicol. 2013 Dec;62:869-75. doi: 10.1016/j.fct.2013.10.02 3. Epub 2013 Oct 21. PMID: 24157545.

91. Piche, L.A., Draper, H.H. and Cole, P.D. (1988), Malondialdehyde excretion by subjects consuming cod liver oil vs a concentrate of n-3 fatty acids. Lipids, 23: 370-371. https://doi.org/10.1007/BF02537352

92. Tu T, Li B, Li X, Zhang B, Xiao Y, Li J, Qin F, Liu N, Sun C, Liu Q, Zhou S. Dietary ω-3 fatty acids reduced atrial fibrillation vulnerability via attenuating myocardial endoplasmic reticulum stress and inflammation in a canine model of atrial fibrillation. J Cardiol. 2022 Feb;79(2):194-201. doi: 10.1016/j.jjcc.2021.08.012. Epub 2021 Oct 23. PMID: 34702603.

93. Driscoll DF, Welty FK, Bistrian BR. Omega-3 Fatty Acids as Antiarrhythmic Drugs: Upstream Target Modulators Affecting Acute and Long-Term Pathological Alterations in Cardiac Structure and Function. Crit Care Explor. 2023;5(10):e0977. Published 2023 Sep 22. doi:10.1097/CCE.0000000000000977

94. Sakabe M, Shiroshita-Takeshita A, Maguy A, Dumesnil C, Nigam A, Leung TK, Nattel S. Omega-3 polyunsaturated fatty acids prevent atrial fibrillation associated with heart failure but not atrial tachycardia remodeling. Circulation. 2007 Nov 6;116(19):2101-9. doi: 10.1161/CIRCULATIONAHA.107.704759. Epub 2007 Oct 22. PMID: 17967774.

95. Mozaffarian D, Psaty BM, Rimm EB, et al. Fish intake and risk of incident atrial fibrillation. Circulation. 2004;110(4):368-373. doi:10.1161/01.CIR.0000138154.00779.A5

96. Zhang B, Xiong K, Cai J, Ma A. Fish Consumption and Coronary Heart Disease: A Meta-Analysis. Nutrients. 2020;12(8):2278. Published 2020 Jul 29. doi:10.3390/nu12082278

97. Zhao, LG., Sun, JW., Yang, Y. et al. Fish consumption and all-cause mortality: a meta-analysis of cohort studies. Eur J Clin Nutr 70, 155–161 (2016). https://doi.org/10.1038/ejcn.2015.72.

98. Yokoyama M, Origasa H, Matsuzaki M, et al. Effects of eicosapentaenoic acid on major coronary events in hypercholesterolaemic patients (JELIS): a randomised open-label, blinded endpoint analysis [published correction appears in Lancet. 2007 Jul 21;370(9583):220]. Lancet. 2007;369(9567):10 90-1098. doi:10.1016/S0140-6736(07)60527-3

99. Saito Y, Yokoyama M, Origasa H, et al. Effects of EPA on coronary artery disease in hypercholesterolemic patients with multiple risk factors: sub-analysis of primary prevention cases from the Japan EPA Lipid Intervention Study (JELIS) [published correction appears in Atherosclerosis. 2009 May;204(1):233]. Atherosclerosis. 2008;200(1):135-140. doi:10 .1016/j.atherosclerosis.2008.06.003

100. Manson JE, Cook NR, Lee IM, et al. Marine n-3 Fatty Acids and Prevention of Cardiovascular Disease and Cancer. N Engl J Med. 2019;380(1):23-32. doi:10.1056/NEJMo a1811403

101. Sweeney TE, Gaine SP, Michos ED. Eicosapentaenoic acid vs. docosahexaenoic acid for the prevention of cardiovascular disease. Curr Opin Endocrinol Diabetes Obes. 2023;30(2):87-93. doi:10.1097/MED.0000000000000796