When purchasing a fish oil supplement, there are a number of things that should be considered: the amount of EPA+DHA, the source of the fish oil, and whether the fish oil is in triglyceride or ethyl ester form. In this informative and in-depth article, you’ll learn:

  • What triglycerides and ethyl esters are
  • What the differences are between the two
  • How producing fish oils as triglycerides is more costly, but improves the final product
  • How to easily determine whether your fish oil is a triglyceride or ethyl ester
  • And more!

Why are so many people talking about fish oil?

Fish oil supplementation has increased significantly within the past decade as a result of scientific findings, media exposure, and word of mouth. In fact, fish oil supplements are now the third most frequently used supplements following multivitamin-minerals and calcium-containing supplements [1] .

There has been plenty of conversation and debate regarding fish oil quality and bioavailability (which is the ability to be absorbed and used by our bodies)—in particular with respect to the different molecular forms of fish oil supplements that are commercially available.

What are the quality standards for omega-3 fish oil products?

The Global Organization for EPA and DHA (GOED) is an important proactive and accountable association of the finest manufacturers, marketers, and supporters of EPA and DHA omega-3 fatty acids. GOED sets strict quality and purity standards for the omega-3 business sector and the GOED Voluntary Monograph has served as the definition of quality in the industry since 2002.

The Monograph adopts the strictest aspects of quality and safety regulations around the world, including environmental contaminants and oxidation, thus ensuring that products meeting this strict standard are safe and effective.

The GOED Monograph represents an important step forward in the standardization of high quality fish oil but, unfortunately, it does not address all quality-related issues. In particular, the Monograph does not differentiate between lipid classes (or molecular forms) of fish oil products.

Fish oils have traditionally been marketed in a molecular form called triglycerides. Within the past decade, however, “fish oil” supplements are largely marketed in an alternative molecular form called fatty acid ethyl esters.

What are triglycerides?

Triglycerides are made of three fatty acids (e.g. EPA and DHA) attached to a glycerol backbone. This is the molecular form that makes up virtually all fats and oils in both animal and plant species. The omega-3 fats present naturally in fish are almost exclusively triglycerides [2].

What are ethyl esters?

Ethyl esters are made of one fatty acid attached to one ethanol molecule. They are a class of lipids that are well characterized with their own monograph in the British Pharmacopoeia [3]. Generally, ethyl esters are not found in nature, and are only created through chemical synthesis.

How are triglycerides and ethyl esters different?

EPA and DHA in ethyl ester form have different chemical properties than EPA and DHA provided in the natural triglyceride form of fish oil. Both triglycerides and ethyl esters are “esterified forms,” which means that an ester link/bond holds the fatty acids onto their chemical backbone. The fatty acids in triglycerides are esterified to a glycerol backbone, whereas the fatty acids in ethyl esters are esterified to an ethanol (alcohol) backbone. This may seem inconsequential, but it is important.

According to Bailey’s Industrial Oil and Fat Products (6thEd.), “Fatty acids in oils and fats are found esterified to glycerol [2].” Given this definition in a standard reference book on food chemistry and processing technology related to edible oils, ethyl esters are not technically oils.

Triglycerides, on the other hand, which contain three fatty acids attached to a glycerol backbone, meet the above definition and are therefore classified as oils. Labelling ethyl ester forms of EPA and DHA as “fish oil” is legally allowed, but it could be considered a misnomer since ethyl esters are not technically oil.

How are ethyl esters produced?

EPA and DHA in ethyl ester form are highly refined omega-3 fatty acids that are created by reacting free fatty acids (FFA) with ethanol in a process called trans-esterification. This process involves removing the glycerol backbone of triglyceride fish oil, resulting in FFA and a free glycerol molecule. An ethanol molecule is then attached to each of the FFA, creating ethyl esters.

The resulting ethyl esters allow for the concentration of the omega-3 long chain fatty acids at lower temperatures. This process (molecular distillation) allows for the selective concentration of EPA and DHA to levels greater than found naturally in fish [4]. The resulting ethyl ester concentrate of EPA and DHA is subsequently marketed and sold as “fish oil concentrate.”

Are all fish oil concentrates ethyl esters?

Manufactured ethyl ester concentrates can be converted back to the natural triglyceride form using enzymes in a process called glycerolysis. Food-grade enzymes separate the ethanol molecule from the fatty acid, creating a FFA and a free ethanol molecule.

When glycerol is reintroduced to the solution, the enzymes then re-esterify the fatty acids back onto a glycerol backbone, creating triglyceride oil. These oils are commonly referred to as re-esterfied (or reformed) triglycerides, which have identical structures to natural triglycerides but with higher concentrations of the desired fatty acids, EPA and DHA.

The process of converting ethyl esters back to triglycerides is costly, and is therefore bypassed by many fish oil manufacturers. In fact, the vast majority of fish oil concentrate softgels sold globally—including those sold in North America—are ethyl ester concentrates. Only a small percentage of fish oil concentrate softgels on the market are true triglyceride oils.

While fish oil softgels are commonly found in ethyl ester form, the majority of liquid fish oil products sold in North America are in triglyceride form. Liquid ethyl ester products are not widely used because they degrade faster and are characterized by a more intense fishy flavour, making them less palatable for consumers.

How do triglycerides work in my body, compared to ethyl esters?

While converting concentrated ethyl ester fish oil back to its natural triglyceride form increases manufacturing costs, it improves metabolism and bioavailability.

All dietary fats (triglycerides) are digested in the small intestine by the action of bile salts and pancreatic lipase. Bile salts break up fat globules into much smaller emulsion droplets, which increase the surface area where lipase can work to liberate two of the three fatty acids from the triglyceride, resulting in two FFA and a monoglyceride (one fatty acid attached to glycerol).

FFA and monoglycerides then form micelles, which are absorbed by intestinal enterocytes, the absorptive cells lining the intestines.  Once inside the enterocyte, the FFA and monoglycerides are reassembled back into triglycerides. Carrier molecules called chylomicrons then transport the triglycerides into the lymphatic channel and finally into the blood (reviewed in [5]).

The digestion of ethyl ester is slightly different, because they lack a glycerol backbone. In the small intestine, ethyl esters are emulsified by bile salts and hydrolyzed by pancreatic lipase. This hydrolysis releases the fatty acid from the ethanol backbone resulting in a FFA and an ethanol molecule.

While the release of ethanol in the intestine has been expressed as a potential concern regarding the safety of ethyl esters, the ethanol release in this process is considered insignificant and ethyl esters are safe for consumption in humans [6].

Similar to triglycerides, the FFA liberated from ethyl esters are absorbed by enterocytes where they are converted to triglycerides so they can be transported in the blood. This step is straightforward with triglycerides, since they already contain a glycerol molecule that can be used to re-esterify the FFA back to triglycerides within intestinal enterocytes.

Ethyl esters contain ethanol and not glycerol, which means that the FFA must obtain a glycerol molecule from another source (such as dietary fat) within the enterocyte to become transformed into a triglycerides.

Once transformed, triglycerides are packaged into chylomicrons that transport the TG into the lymphatic channel and subsequently into the blood.

Metabolism of ethyl esters is less efficient than triglycerides. Pancreatic lipase hydrolyses ethyl esters to a lesser degree and at a slower rate than triglycerides [7-10]. In fact, it takes pancreatic lipase 10 to 50 times longer to break the fatty acid-ethanol bond found in ethyl esters compared to breaking the fatty acid-glycerol bond found in triglycerides [11].

To complicate matters, EPA and DHA hydrolysis may be further compromised in individuals with digestive disorders, such as pancreatic insufficiency. This has been demonstrated in a cystic fibrosis population whereby supplementation with ethyl ester fish oil increased the EPA/arachidonic acid ratio 9.8-fold, falling short of the 23-fold increase in healthy subjects [12].

Furthermore, since a glycerol or monoglyceride substrate is absent in ethyl esters, triglyceride re-synthesis in the enterocyte is delayed and subsequent transport of EPA and DHA to the blood may be less efficient in ethyl ester fish oils than triglyceride fish oils [13].

Given the metabolism differences between triglycerides and ethyl esters, one could conclude that there would be differences in EPA and DHA absorption as well. A comprehensive review of the scientific literature does indeed provide evidence suggesting that fish oil in triglyceride form is more efficiently absorbed than fish oil in ethyl ester form [7, 8, 14-16].

Which form of omega-3 fish oil is better absorbed by my body?

There is considerable debate about whether different molecular forms of EPA and DHA are similarly absorbed. Of course, fish contains omega-3 fatty acids in the natural triglyceride form—however, the vast majority of studies demonstrating the clinical efficacy of fish oil have used ethyl ester fish oil. Importantly, most of these studies have not directly compared ethyl esters versus triglycerides in terms of clinical efficacy or absorption.

There have been a number of studies that have directly compared the absorption or bioavailability of ethyl ester and triglyceride oils. Although a small number of these studies have found that absorption is similar between triglyceride and ethyl ester oils [9, 17], the majority of research supports significantly increased absorption and bioavailability of EPA and DHA from triglycerides.

For example, a small study found that plasma EPA and DHA levels were 50% higher after supplementation with triglyceride fish oil than with ethyl ester fish oil [14].

A different study found more dramatic differences, with EPA and DHA from triglycerides being 340% and 271% better absorbed, respectively, than those consumed as ethyl esters. The authors of that study went so far as concluding that ethyl esters “are poorly absorbed in man” due to decreased pancreatic lipase activity [7].

An additional study involving supplementation of fasting volunteers with 1,000 mg of EPA as either triglycerides or ethyl esters reported that EPA incorporation into plasma lipids was considerably higher for the triglyceride group; incorporation of EPA into plasma lipids was also much slower for the ethyl ester group. The researchers suggested that this could be a result of low pancreatic lipase during ethyl ester digestion, coupled with reduced incorporation of EPA into micelles [8].

While these smaller studies provide support that EPA and DHA are better absorbed from triglyceride oils than ethyl ester oils, some more recent well-designed studies provide strong evidence that triglyceride oils have superior bioavailability.

A two-week double-blind placebo-controlled study compared the absorption of different oil preparations in 72 subjects: a triglyceride concentrate, an ethyl ester concentrate, a FFA preparation, fish body oil that was obtained from pressing the bodies of fish (considered natural triglycerides fish oil), cod liver oil obtained from pressing the livers of cod fish (considered natural triglycerides fish oil), and a corn placebo oil.

Increases in absolute amounts of EPA and DHA in blood lipids were examined and results revealed that bioavailability of EPA and DHA from the triglyceride concentrate was 24% greater than natural fish oil, while the bioavailability from the ethyl ester concentrate was 27% less vs. natural fish oil.

In other words, EPA and DHA were significantly better absorbed in the triglyceride concentrate and natural triglyceride fish oil than they were in the ethyl ester concentrate. The researchers stated that these results were likely a result of pancreatic lipase activity, which limited the absorption of EPA and DHA from the ethyl ester oil [15].

Another six-month double-blind placebo-controlled trial involving 150 subjects investigated the effect of EPA and DHA given as a triglyceride concentrate, ethyl ester concentrate, or a corn placebo oil on the omega-3 index, a measure of red blood cell EPA and DHA levels.

After three months, the omega-3 index increased by 186% in the triglyceride group compared to 161% in the ethyl ester group, representing a 25% difference.

After six months, the omega-3 index increased by 197% in the triglyceride group compared to 171% in the ethyl ester group, representing a 26% difference [16].

Furthermore, in a subset of these subjects, fasting serum triacylglycerol levels were found to be significantly reduced from baseline in the triglyceride group (but not the ethyl ester group) after three and six months of supplementation [18]. These results are extremely valuable, because they suggest that not only are cellular levels of EPA and DHA raised about 25% more effectively when consumed as a triglyceride concentrate versus an ethyl ester concentrate, but also that the bioavailability difference translates into more desirable biological effects.

Which form of omega-3 fish oil has a better shelf life?

Few studies directly compare oxidation rates of triglyceride and ethyl ester fish oil products with similar levels of EPA and DHA. The studies that have been conducted demonstrated that fatty acids in the form of ethyl ester oxidize faster than those in triglyceride form. For example, when triglyceride and ethyl ester fish oils were incubated at 80°C while bubbling air through them, ethyl esters oxidized more rapidly than triglycerides [19].

Additionally, in another study where oxygen concentration was monitored in the headspace of DHA-enriched fish oil stored at various temperatures, results revealed that DHA in ethyl ester form oxidized faster than triglycerides [20].  These results were supported by another clinical trial that found oxidation proceeded more rapidly in ethyl esters than triglycerides [21].

Most relevant is a recently conducted study that is currently under review for peer-reviewed publication. This trial assessed oxidation of triglyceride and ethyl ester fish oil that had similar amounts of EPA and DHA at 5-60°C.  The rate of oxidation, as measured by change in peroxide value and Anisidine value, was found to be lower in triglyceride fish oil than in ethyl ester [22].

How do I know if my omega-3 fish oil is an ethyl ester or a triglyceride?

Fish oil products available in North America do not require labels to state whether they are in triglyceride or ethyl ester form. Since this information is not easily accessible, there is a simple, inexpensive, and rapid method to determine if a fish oil supplement is in the triglyceride or ethyl ester form by using polystyrene (Styrofoam) cups.

Measure and place 20 ml of fish oil in a polystyrene cup, place the cup on a plate to avoid any mess, and observe the cup after 10 minutes. Due to their chemical composition, ethyl ester oils will actually dissolve the polystyrene cup. While this effect becomes evident after just a few minutes, significant leakage is seen after 10 minutes.

Ethyl ester oils dissolve polystyrene by dissolving the chemical bonds used to keep the polystyrene from collapsing. When these bonds are broken, the air trapped in the polystyrene escapes causing the structure to collapse.

Natural triglyceride fish oils placed in the same cup, on the other hand, will not show leakage after 10 minutes—but they may show leakage through the cup in very small amounts after 2-3 hours.

Conclusion

Natural triglyceride fish oil supplements offer numerous advantages over ethyl ester fish oil supplements.  First, triglyceride oils are the molecular form found naturally in fish, and they are more resistant to oxidation than ethyl ester oils.

Most important, however, is the fact that triglyceride fish oils are metabolized and absorbed more efficiently and completely than ethyl ester oils, which may increase overall health benefits for consumers.

References

1.         Bailey, R.L., et al., Why US adults use dietary supplements. JAMA Intern Med, 2013. 173(5): p. 355-61.

2.         Shahidi, F., ed. Bailey’s Industrial Oil and Fat Products, 6th edition 2005, John Wiley  and Sons: Hoboken, USA.

3.         Omega-3-Acid Ethyl Esters 90, in British Pharmacopoeia 2012 Online. 2011, TSO Information & Publishing Solutions: Norwich, England.

4.         Breivik, H., H. G.G., and B. Kristinsson, Preparation of highly purified concentrates of eicosapentaenoic acid and docosahexaenoic acid. JAOCS, 1997. 74(11): p. 1425-29.

5.         Carlier, H., A. Bernard, and C. Caselli, Digestion and absorption of polyunsaturated fatty acids. Reprod Nutr Dev, 1991. 31(5): p. 475-500.

6.         Bookstaff, R.C., et al., The safety of the use of ethyl oleate in food is supported by metabolism data in rats and clinical safety data in humans. Regul Toxicol Pharmacol, 2003. 37(1): p. 133-48.

7.         Lawson, L.D. and B.G. Hughes, Human absorption of fish oil fatty acids as triacylglycerols, free acids, or ethyl esters. Biochem Biophys Res Commun, 1988. 152(1): p. 328-35.

8.         el Boustani, S., et al., Enteral absorption in man of eicosapentaenoic acid in different chemical forms. Lipids, 1987. 22(10): p. 711-4.

9.         Krokan, H.E., K.S. Bjerve, and E. Mork, The enteral bioavailability of eicosapentaenoic acid and docosahexaenoic acid is as good from ethyl esters as from glyceryl esters in spite of lower hydrolytic rates by pancreatic lipase in vitro. Biochim Biophys Acta, 1993. 1168(1): p. 59-67.

10.       Ikeda, I., et al., Digestion and lymphatic transport of eicosapentaenoic and docosahexaenoic acids given in the form of triacylglycerol, free acid and ethyl ester in rats. Biochim Biophys Acta, 1995. 1259(3): p. 297-304.

11.       Yang, L.Y., A. Kuksis, and J.J. Myher, Lipolysis of menhaden oil triacylglycerols and the corresponding fatty acid alkyl esters by pancreatic lipase in vitro: a reexamination. J Lipid Res, 1990. 31(1): p. 137-47.

12.       Henderson, W.R., Jr., et al., Oral absorption of omega-3 fatty acids in patients with cystic fibrosis who have pancreatic insufficiency and in healthy control subjects. J Pediatr, 1994. 124(3): p. 400-8.

13.       Yang, L.Y., A. Kuksis, and J.J. Myher, Intestinal absorption of menhaden and rapeseed oils and their fatty acid methyl and ethyl esters in the rat. Biochem Cell Biol, 1990. 68(2): p. 480-91.

14.       Beckermann, B., M. Beneke, and I. Seitz, [Comparative bioavailability of eicosapentaenoic acid and docasahexaenoic acid from triglycerides, free fatty acids and ethyl esters in volunteers]. Arzneimittelforschung, 1990. 40(6): p. 700-4.

15.       Dyerberg, J., et al., Bioavailability of marine n-3 fatty acid formulations. Prostaglandins Leukot Essent Fatty Acids, 2010. 83(3): p. 137-41.

16.       Neubronner, J., et al., Enhanced increase of omega-3 index in response to long-term n-3 fatty acid supplementation from triacylglycerides versus ethyl esters. Eur J Clin Nutr, 2011. 65(2): p. 247-54.

17.       Nordoy, A., et al., Absorption of the n-3 eicosapentaenoic and docosahexaenoic acids as ethyl esters and triglycerides by humans. Am J Clin Nutr, 1991. 53(5): p. 1185-90.

18.       Schuchardt, J.P., et al., Moderate doses of EPA and DHA from re-esterified triacylglycerols but not from ethyl-esters lower fasting serum triacylglycerols in statin-treated dyslipidemic subjects: Results from a six month randomized controlled trial. Prostaglandins Leukot Essent Fatty Acids, 2011. 85(6): p. 381-6.

19.       Lee, H., et al., Analysis of headspace volatile and oxidized volatile compounds in DHA-enriched fish oil on accelerated oxidative storage. J Food Sci, 2003. 68(7): p. 2169-77.

20.       Yoshii, H., et al., Autoxidation kinetic analysis of docosahexaenoic acid ethyl ester and docosahexaenoic triglyceride with oxygen sensor. Biosci Biotechnol Biochem, 2002. 66(4): p. 749-53.

21.       Litiwinienko, G., Daniluk, A., & Kasprzycka-Guttman, T. , Study on autoxidation kinetics of fats by differential scanning calorimetry. 1. Saturated C12-C18 fatty acids and their esters. . Ind Eng Chem Res 2000. 39(1): p. 7-12.

22.       Sullivan Ritter, J.C., S.M. Budge, and F. Jovica, Oxidation rates of triglyceride and ethyl ester fish oils. Submitted to Food Chem (in review), 2014.

 

Download the Natural Health Enthusiast's Guide to Omega-3 Fish Oil

Omega-3 fish oils are essential for your overall health and wellness. When you download the FREE Natural Health Enthusiast’s Guide to Omega-3 Fish Oil, you’ll learn what look for when choosing an omega-3 fish oil supplement, the potential health benefits of omega-3 fish oil supplements, and more!

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