Recently, there have been several important discoveries about vitamin E. Last month, we discussed a little about the public’s concept of the health roles of vitamin E and asked leading vitamin E expert Maret Traber, Ph.D., to discuss some basics about our needs for vitamin E. As a starter, Dr. Traber reviewed her scientifically “game-changing” research on the human tocopherol transport protein (TTP) and why the only form of vitamin E with a significant role in human health is alpha-tocopherol. This month we will discuss more of the vitamin E basics including the differences between various forms of vitamin E available in dietary supplements, especially the differences between “natural-source” and synthetic vitamin E. We will also discuss why there is no significant difference in health benefits between the various vitamin E esters (e.g., tocopheryl acetate, tocopherol succinate) in oral vitamin E supplements.

Vitamin E is needed as an essential nutrient, however about 90% of Americans do not even get the recommended dietary allowance (RDA). After reviewing these basic actions of vitamin E, we will then be ready to discuss some of the important new research in subsequent columns including why vitamin E is needed for brain health. Is there a link between vitamin E deficiency and neurological damage? Will supranutritional amounts of vitamin E help Alzheimer’s disease patients? There is also new research on whether metabolic syndrome impairs vitamin E absorption and whether obese people need more vitamin E but actually get less. Do pregnant or lactating women need more vitamin E? Can high blood cholesterol or triglycerides interfere with vitamin E reaching body tissues? Do epidemiological (population) studies show that vitamin E reduces the incidence of heart disease? What about possible interactions between vitamin E and vitamin K? There is so much to learn!

Dr. Maret G. Traber is the Helen P. Rumbel Professor for Micronutrient Research in the Linus Pauling Institute and a professor in the nutrition program, College of Public Health and Human Sciences, at Oregon State University. She received her Ph.D. in nutrition from the University of California, Berkeley, CA. She currently serves on the editorial boards of Free Radical Biology & Medicine and the Journal of Nutrition.

Dr. Traber served on the National Academy of Sciences, Institute of Medicine, Panel on Dietary Antioxidants to develop the 2000 Dietary Requirements for vitamins C and E, selenium and carotenoids. She is considered one of the world’s leading experts on vitamin E. She pioneered the use of deuterium-labeled vitamin E for studies evaluating vitamin E status in humans. Her studies caused a paradigm shift in our understanding of the mechanisms regulating vitamin E availability in humans. Her work has provided the scientific basis for understanding the complex role of vitamin E in human health.

She has published over 190 peer-reviewed and 110 invited papers in highly regarded journals. She pioneered the methodologies for evaluating vitamin E status in humans, and through this, identified key mechanisms for regulating vitamin E bioavailability in humans. In 2013, she received the Pfizer Consumer Healthcare Nutrition Sciences Award, presented by the American Society for Nutrition and the DSM Nutritional Science Award 2013 on Fundamental Research in Human Nutrition. Current research projects include vitamin E bioavailability and requirements in humans using advanced pharmacokinetic methodologies; assessment of the interactions of vitamin E and K; and determination of the mechanisms of vitamin E function during embryogenesis using zebrafish.

Passwater: Dr. Traber, after discussing the human tocopherol transport protein (TTP) and the fact that it transports only the alpha-tocopherol isomer for utilization and storage in humans, it begged the question of just what is the essential nutritional role of vitamin E.

Traber: Well, to help answer that question, we have focused our research on an animal that does not require a placenta to examine the role of vitamin E in fertility. We use the zebrafish, which we make vitamin E deficient by feeding a diet exquisitely low in alpha-tocopherol (and all the other tocopherols). Vitamin E is critical for the embryo. We use the zebrafish embryo because it is a great model for studying embryogenesis. It doesn’t have the ethical and logistic issues common in human and rodent studies.

Zebrafish have many genes in common with human beings and so researchers often study them to gain insights into human disorders and conditions. The NIH video, Zebrafish: A Key to Understanding Human Development, shows the importance of zebrafish in developmental research (1). This video is available for free viewing on the Internet.

In our studies, the zebrafish are spawned; they lay and fertilize their eggs. We know the precise ages of the embryos.

Figure 1: Zebrafish have many genes in common with human beings and so researchers often study them to gain insights into human disorders and conditions. Zebrafish have no placenta and zebrafish embryos are an excellent model for studying embryogenesis.

The vitamin E-deficient embryos appear fine for the first few days. Normally, embryos become larvae and swimming fish within five days. By the 48th hour, however, our vitamin E-deficient embryos begin to develop abnormalities and within five days, about 70% are abnormal or dead. Thus, vitamin E is needed by the developing embryo.

Passwater: For years, vitamin E potency was based on a bioassay (i.e., a study done in animals as opposed to an analytical procedure done in a quality control laboratory) involving rat gestation. It is a difficult bioassay indeed! Does this bioassay have any important relevancy to what we know about vitamin E potency today?

Traber: It is an important assay because it is STILL the assay that is used to define the international unit (IU) for vitamin E that is on every bottle of vitamin E supplements that consumers purchase.
Therefore, I thought it was critical to understand what makes the embryo so sensitive to vitamin E deficiency. We are currently pursuing this line of investigation to answer the question, “Why do we need vitamin E?”

Passwater: In Part One of this series (April 2016), we discussed the various tocopherol isomers—alpha-tocopherol, beta-tocopherol, gamma-tocopherol and delta-tocopherol—as well as their various ester forms used in dietary supplements. Now, let’s consider some of the differences between “natural-source” and synthetic vitamin E.

Humans are designed to use the RRR-form of alpha-tocopherol for their “vitamin E” needs to survive. This is the form of alpha-tocopherol found in nature. At this time, it is also thought that synthetic forms having “2R-“ alpha-configuration can satisfy at least some of the body’s vitamin E needs. The body doesn’t care where the RRR-alpha-tocopherol originated—food or supplement. The body readily extracts RRR-alpha-tocopherol molecules from food and supplements during digestion.

One just does not go into the field and pick “free” RRR-alpha-tocopherol from a tree or dig it up. It must be extracted from the plants. Animals don’t make their own alpha-tocopherol, but they do contain alpha-tocopherol because of the foods they eat. This extraction is easy for the body, but not very economically feasible for commercial production of dietary supplements or enriching foods. It is a rare commercial vitamin E producer that simply extracts and concentrates natural alpha-tocopherol without chemically converting the other tocopherols to increase the yield. Limited commercial success has been obtained with sources such as sunflower seed oil, which is higher in alpha-tocopherol than vegetable oils. Your body doesn’t really care—it just wants the RRR-alpha-tocopherol molecules.

“Natural” vitamin E is actually “natural-source” vitamin E, meaning it starts with a food source containing both alpha-tocopherol and non-alpha-tocopherols (e.g., gamma-tocopherol) which are then “methylated” to alpha-tocopherol. “Synthetic vitamin E” means it is not made from a food source.

Is most natural-source alpha-tocopherol semi-synthetic? Most “natural-source” vitamin E producers have found it competitively economical to take a food source (such as soy oil) having relatively high quantities of gamma-tocopherol and other non-alpha-tocopherols compared to alpha-tocopherol levels and methylate it to produce additional alpha-tocopherol to increase vitamin E yield. Methylation consists of inserting methyl groups in available carbons in the chromal “heads” of the tocopherols. Several natural-source vitamin E producers use such patent-protected technology. As an example, a classically used process (EP0176690 A1) produces RRR-alpha-tocopherol from non-alpha-tocopherol homologues by using an alkylation agent such as methanol for a sufficient length of time at a temperature of about 300–550 °C in the presence of an alkylation catalyst.

Thus, “natural-source” begins with natural RRR-forms of tocopherols, which are converted in manufacturing facilities to mostly RRR-alpha-tocopherol. This can be considered “semi-synthetic,” but it yields true vitamin E (which is RRR-alpha-tocopherol) and not any of the un-natural forms produced by synthetic processes that start with petroleum-derived chemicals. It is less expensive to start with a petroleum-based material and react it to produce eight isomers of alpha-tocopherol.

Vitamin E is a commodity, but making it is not an easy undertaking. Veteran readers of this column likely remember the days when just three companies—Henkel, Eastman and Tama—successfully produced natural-source vitamin E commercially. Today, there are more, but still relatively few, producers of vitamin E in the world. Archer Daniels Midland Company (ADM) claims to be the world’s largest manufacturer of natural-source vitamin E. DSM claims to be the world’s largest manufacturer of vitamin E and also distributes natural-source vitamin E produced by Cargill. There are many brands that sell vitamin E supplements. They all must purchase bulk vitamin E from a producer. Few brands—even those that manufacture their own tablets and hardshell capsules—make their own soft gels. Many brands claim to be most natural or superior in some way. If this is of interest to you, ask to see evidence of what company they buy their vitamin E from and the manufacturing process the producer uses.

There are important differences in the vitamin E activity of equal weights of natural and synthetic vitamin E (2). Consumers should consider vitamin E activity in terms of IUs rather than the weight. Many consumers have long been aware of the former differences in designation of natural and synthetic vitamin E. The older designation was made in terms of the ability of natural vitamin E to rotate plane-polarized light clockwise (to the right) (dextro), whereas synthetic vitamin E does not rotate the light. This difference is caused by the arrangement of atoms in the two forms. An “epimer” is a stereoisomer that rotates polarized light because it differs in confirmation at only one of several chiral centers.

The challenge was how to show the true vitamin E activity of the various forms of vitamin E correctly. The bioassay forced vitamin E potency to be expressed in terms of “units.” Next, an arbitrary definition of an IU was established.

Many years ago, the IU for vitamin E activity was set based on the first commercially available synthetic form. At that time, this synthetic vitamin E was an equal amount of two tocopherol forms: dextro-alpha-tocopheryl acetate (d-alpha-tocopheryl acetate) and levo-alpha-tocopheryl acetate (l-alpha-tocopheryl acetate). “Levo” means the molecules rotate polarized light to the left. This synthetic form was called dl-alpha-tocopheryl acetate. One milligram of dl-alpha-tocopheryl acetate was arbitrarily set as one IU of vitamin E.

This first commercially available synthetically produced vitamin E was called dl-alpha-tocopherol and the natural form found in nature was called d-alpha-tocopherol. This terminology is incorrect, but it is still in wide use today on food and supplements labels. It is not the current terminology in use by scientists. The current commercially available synthetic vitamin E is a mixture of eight stereoisomers rather than just two.

The acetate ester form of alpha-tocopherol is a stable form of vitamin E that is most frequently used in supplements and to fortify foods. It is produced by reacting acetic acid with vitamin E to add the acetate radical to the hydroxyl portion in the “head” of the vitamin E molecule. Depending on the form of alpha-tocopherol used, this ester form is called either d-alpha-tocopheryl acetate or dl-alpha-tocopheryl acetate. The dl-alpha-tocopheryl acetate form has been arbitrarily set as the standard for measuring vitamin E activity. One milligram of dl-alpha-tocopheryl acetate is considered to be 1 IU. One milligram of dl-alpha-tocopherol contains 1.1 IU, 1 mg of d-alpha-tocopheryl acetate has 1.36 IU and 1 mg of d-alpha-tocopherol has 1.49 IU of activity.

Decades ago, synthetic vitamin E was produced from natural phytol, which is a plant sterol intermediate with the correct chirality in the tail, and thus resulted in a racemic mixture having only one chiral center. This manufacturing process resulted in just the two stereoisomers: d-alpha-tocopherol and l-alpha-tocopherol. Scientists call this form 2-ambo-alpha-tocopherol, as it has only one asymmetrical carbon, which is at the 2-position on the chromal ring (head). It is no longer commercially available.

Currently, synthetic vitamin E is produced from synthetic isophytol, which produces equal amounts (12.5%) of eight stereoisomers because there are three asymmetrical carbon atoms at the 2, 4’ and 8’ positions in the phytyl “tail.” These stereoisomers produced in the configuration of the “tail” are not the same as speaking of the isomers produced by different amounts of methyl groups in the “head.”
It is still labeled as dl-alpha-tocopherol, which is incorrect. It is more accurately called all-rac-alpha-tocopherol. Seven of the stereoisomers are not known to be found anywhere in nature. The body circulates and maintains four of the eight stereoisomers (2R).

When the manufacturing process for synthetic vitamin E changed, the description of the vitamin E stereoisomers changed from the Latin “dextro” and “levo” to the German “rectus” and “sinister” at each of the three molecular centers of asymmetry in the vitamin E molecule. (Please see Figure 2.)

Figure 2: Commercial production of alpha-tocopherol is either “natural-source” or “synthetic.” Synthetic production yields equal amounts of eight stereoisomers of alpha-tocopherol. Only four of the stereoisomers—the “2R-” stereoisomers—are believed to have human vitamin E activity (2000 DRI). The difference in activity is taken into account by assigning different IU activity for the two forms. Drawing courtesy of ADM.

The serious vitamin E student may wish to read the sidebar on p.40 for help in understanding the importance of the various stereoisomers of alpha-tocopherol.

Dr. Traber, plants synthesize the alpha-tocopherol structure in the “RRR” form, whereas today’s manufacture of synthetic alpha-tocopherol also produces seven other forms (RSR-, RRS-, RSS-, SRR-, SSR-, SRS- and SSS-). Please see Figures 2 and 3. Are all forms of alpha-tocopherol used by humans?

Traber: The 2000 DRI committee decided that 2R-alpha-tocopherols (RRR, RSR, RRS and RSS) were the forms of vitamin E recognized by the alpha-TTP. (3) They recommended that vitamin E be reported as milligrams of 2R-alpha-tocopherol. Thus, half of the alpha-tocopherol in the synthetic is useful as vitamin E in the body.

Passwater: Or, to put it another way, the “2S” stereoisomers are not maintained in the body. Thus, half of the synthetic alpha-tocopherol is not used in the body and therefore is not included in the determination of the strength or activity of the product. This is taken into account by the use of IU system in the labeling.

Traber: The basis for this recommendation is the human disorder ataxia with vitamin E deficiency. This disorder is rare, but is important because individuals can absorb vitamin E, but quickly metabolize and excrete it because they are lacking the alpha-TTP. This protein preferentially recognizes alpha-tocopherol. It’s kind of like your car keys; they won’t unlock the front door of your house. Maybe your neighbor’s keys look like yours, but they won’t work either. Readers can download the DRI book free from the link given in Reference 3.

	Figure 3: The chemical structural differences in the stereoisomers produced commercially in the “natural-source” and “synthetic” manufacturing processes.

Figure 4 shows the stereochemical differences between the natural RRR-alpha-tocopherol and one of the synthetic forms, SRR-alpha-tocopherol. Note that the difference in the vitamin E molecule’s tail (side chain) aligning with the vitamin E molecule’s “head” ring structure is like the fingers on a glove not lining up with the fingers on the hand. The “S” configuration sticks out more like an opposing thumb than a normal “finger.” As I mentioned in Part One, the ability of TTP to transport vitamin E depends on it fitting hand and glove with the molecule.

Passwater: Does the designation “2R-alpha-tocopherol” include the RRR-, RSR-, RSS and RSS- stereoisomers? Is this designation essentially the same now as the old designations of dextro-alpha-tocopherol (d-alpha-tocopherol)? Is this group of four stereoisomers biochemically different from just the single stereoisomer RRR-alpha-tocopherol produced in nature?

Traber: These four “R” stereoisomers have not been tested directly with the TTP, but we tested them in mice. We compared radio-labeled RRR with all-rac-alpha-tocopherol and found the ratio was two (4).
 

Figure 4: There are significant differences in the stereochemical forms between vitamin E (RRR-alpha-tocopherol) and the “S”-variations. The “S” variations do not fit well stereochemically into the vitamin E tocopherol transport protein (TTP) and thus disappear rapidly from the body. This figure shows the stereochemical structural differences between vitamin E and one of the “S” stereoisomers (SRR-alpha-tocopherol). Chemically synthesized alpha-tocopherol, known as all-rac-alpha-tocopherol, contains a mixture of eight stereoisomers. Only the vitamin E stereoisomers having an R-configuration in Position Two of alpha-tocopherol meet human vitamin E requirements. Thus, half of the stereoisomers present in all-rac-alpha-tocopherol are considered to be biologically active forms of vitamin E.

Passwater: Okay, we’re done with the biochemistry! Now back to the general questions. Do some of the “non-alpha” tocopherols have specific actions that alpha-tocopherol does not have? If tocotrienols and non-alpha-tocopherols have specific actions that alpha-tocopherol does not have, is it time to separate them as nutrients other than alpha-tocopherol vitamin E?

Traber: Non-alpha-tocotrienols and tocotrienols are interesting; plants make them as fat-soluble antioxidants. But, people actively metabolize them and they disappear from the circulation very quickly.

Passwater: Alpha-tocopherol is an antioxidant. This means that it protects other compounds and body components from oxidation and/or reactive oxygen species (ROS). As we discussed in Part One, tocopherol does this by donating hydrogen from its hydroxyl group. In plants and animals, tocopherol is rarely found in an unprotected form. It is generally protected in a storage or transport form until it is needed to react with a free radical or oxygen. Similarly, if tocopherol is allowed to sit in air or a pill, it will quickly react with oxygen and no longer have antioxidant ability. It is spent. In the body, such spent tocopherol can be regenerated by other antioxidants such as vitamin C or a selenium enzyme.

Since tocopherol is so reactive with oxygen, even within a pill, it has a short shelf life. Manufacturers normally combine “free” tocopherols with an organic acid such as acetic acid or succinic acid to increase their shelf-life and preserve their antioxidant potential. As mentioned earlier, this combination is called an “ester.”

Many supplements list vitamin E as “alpha-tocopherol” while in reality, many are a more stable ester form of vitamin E such as alpha-tocopheryl acetate or alpha-tocopheryl succinate. The alpha-tocopherol portion is released during digestion and uptake into the bloodstream. Alpha-tocopheryl succinate is a solid at room temperature making it very attractive to use in solid dose forms. Alpha-tocopheryl acetate is a free-flowing liquid at room temperate and can easily be used in softgels. Or, it can be sprayed onto a food-grade excipient for use in solid dosage forms. Manufacturers normally choose the form that is best for the particular supplement depending on whether it is a powder or an oil.

Yet a couple of “test tube” studies suggest that one form or the other may be better for a specific condition. As an example, a study or two suggest that in in vitro (cell cultures), the succinate form is better at reducing cancer cell growth in Petri dishes than the acetate form. Does it make any real difference in the body as to which ester of vitamin E is ingested?

Traber: All of the esters are hydrolyzed (i.e., undergo ester hydrolysis) and are split into a “free” unesterified tocopherol and an acid by a pancreatic esterase enzyme in the middle portion of the small intestine (jejunum), and only tocopherol—not esterified tocopherol—is absorbed (See Figure 5). This process requires fat digestion, so it is important to consume supplements with a meal.

It is also very important to understand how the studies are carried out. Tocopheryl succinate has very interesting effects in cell culture, but the material would have to be given intravenously (IV) or injected to achieve similar results in the body. Moreover, even if given by IV, I am not sure, how long or effective tocopheryl succinate would be in the body. Injections or IV are not the same as oral supplements, which are processed in the digestive tract.

	Figure 5: Vitamin E esters in supplements are hydrolyzed by esterase enzymes to “free” vitamin E in the intestine and then the vitamin E is absorbed via bile-dependent incorporation into intraluminal micelles.

Passwater: My advice to readers is that the next time you read that vitamin E succinate or tocopheryl succinate is more effective than alpha-tocopherol against cancer—and there are a host of such articles on the Internet repeating the same studies over and over—inspect the reference to see if the study is a test tube or cell culture study or if the vitamin E was administered via injection or intravenously (IV). The oral form of vitamin E succinate is broken apart (hydrolyzed) in the intestine to yield the “free” form of tocopherol (just as vitamin E acetate would be) plus succinic acid and is not absorbed as intact vitamin E succinate.

Dr. Traber, you have served on the scientific panel that sets the official RDA and DRI. It has been difficult to assess just how much vitamin E people should consume when we may not fully understand exactly all of the biochemical functions of vitamin E in the body. For decades, we have relied on an assumption or the need for vitamin E to prevent undesirable oxidation and free radical formation from polyunsaturated fats. The assumption was that since we don’t recognize any overt vitamin E deficiency diseases, then the population therefore must be consuming adequate amounts of vitamin E. So, as the thinking went, all that needs to be done is to calculate how much vitamin E is in the American diet and divide that by the population to get the number. This is not very scientific, but a reasonable first estimate in the absence of scientific data. The Elgin Project, headed by Dr. Max Horwitt, was the first study to approximate vitamin E needs indirectly by measuring the oxidation of polyunsaturated fats. Dr. Horwitt chatted with us in 1992 about vitamin E and the RDA (5). Can vitamin E adequacy be assessed by measuring blood levels of vitamin E?

Traber: I am currently working on a biomarker of vitamin E status. It is a metabolite of vitamin E called carboxy ethyl hydroxy chromanol (CEHC). These studies are in progress, but the general idea is that the body makes CEHC when it has sufficient vitamin E.

Passwater: Now that is very interesting and so is your recent research concerning vitamin E and vitamin K interactions. Let’s discuss the vitamin E RDA and more of your recent research on vitamin E and brain health in another column. For now, let’s sum up this column on vitamin E basics with the following thoughts.

Only the alpha-tocopherol form of vitamin E is nutritionally essential for humans, but research is still examining possible nonessential and clinical roles for vitamin E beyond its essential nutritional roles. As far as oral supplements are concerned, there is no significant difference in health benefit between the various esters of vitamin E (e.g., alpha-tocopheryl acetate, alpha-tocopheryl succinate) as they are all hydrolyzed to vitamin E (alpha-tocopherol) in the digestive tract.

The known nutritional requirement for vitamin E is 15 mg (22.5 IU), yet about 90% of the American population does not even reach 12 mg in their diet (6, 7). In the United States, the average intake of alpha-tocopherol from food (including enriched and fortified sources) for adults is 7.2 mg/day; this level is short of being just half of the RDA of 15 mg/day of alpha-tocopherol. Supranutritional intakes of vitamin E—some in the gram (1,000 mg) range—appear to have beneficial health benefits according to clinical studies.

Dr. Traber, your recent research concerning vitamin E is drawing wide interest in the scientific community and we thank you for sharing this information with us. Let’s chat about the vitamin E DRI next time and then we can update our readers about your exciting new research. WF

For more, see WholeFoodsMagazine.com/Columns/Vitamin-Connection

Dr. Richard Passwater is the author of more than 45 books and 500 articles on nutrition. Dr. Passwater has been WholeFoods Magazine’s science editor and author of this column since 1984. More information is available on his Web site, www.drpasswater.com.

References
1. National Institutes of Medicine, “New Video Highlights NIH Investment in Zebrafish Research,” www.nichd.nih.gov/news/releases/Pages/072512-zebrafish.aspx, accessed Apr. 27, 2016.
2. S.K. Jensen, J.V. Nørgaard and C. Lauridsen, “Bioavailability of Alpha-Tocopherol Stereoisomers In Rats Depends On Dietary Doses Of All-Rac- or RRR-Alpha-Tocopheryl Acetate,” Br. J. Nutr. 95 (3), 477–487 (2006).
3. Food and Nutrition Board, Institute of Medicine, Dietary Reference Intakes for Vitamin C, Vitamin E, Selenium, and Carotenoids, National Academy Press, Washington, D.C., 2000. http://www.ncbi.nlm.nih.gov/books/NBK225483.
4. S.W. Leonard, et al., “Incorporation Of Deuterated RRR- Or All Rac A-Tocopherol Into Plasma And Tissues Of A-Tocopherol Transfer Protein Null Mice,” Am. J. Clin. Nutr. 75: 555–560 (2002).
5. R.A. Passwater, “Vitamin E and the RDA,” WholeFoods Mag. (1992).
6. V.L. Fulgoni et al., “Foods, Fortificants, and Supplements: Where Do Americans Get Their Nutrients?” J. Nutr. 141 (10), 1847–1854 (2011). doi: 10.3945/jn.111.142257.
7. A. Moshfegh, J. Goldman and L. Cleveland, “What We Eat In America, NHANES 2001–2002: Usual Nutrient Intakes From Food Compared To Dietary Reference Intakes. USDA, Agricultural Research Service [cited 2014 Apr 8],” www.ars.usda.gov/SP2UserFiles/Place/12355000/pdf/0102/usualintaketables2001–02.pdf.­­­

Published in WholeFoods Magazine June 2016