Human Vitamin E Deficiency, and what is and is NOT vitamin E?

Graphical Abstract

INTRODUCTION

This article is written in response to the discussion forum of this Journal dedicated to a scientific debate on Vitamin E nomenclature [1]. The authors of that article state that vitamin E deficiency was recognized as a human disorder in 1981. However, the literature is replete with examples of human vitamin E deficiency prior to that date. Thus, I have discussed the history of human vitamin E deficiency in the first part of this response. Secondly, the proposal has been made that only RRR-α-tocopherol has been shown to reverse human vitamin E deficiency and therefore should be the only form that should be referred to as vitamin E. However, there are several examples where all rac-α-tocopherol has been used to reverse human vitamin E deficiency symptoms, not the least of which were the studies by Horwitt et al [2]. The second half of this article provides the rationale and evidence as to why vitamin E should be defined as 2R-α-tocopherol.

HISTORY OF HUMAN VITAMIN E DEFICIENCY

Attempts to characterize symptoms of human vitamin E deficiency began over 75 years ago. In the 1950s Horwitt et al [2, 3] induced vitamin E deficiency in men at the Elgin (Illinois) State Hospital by providing them a diet low in vitamin E for 6 years. In addition to low circulating α-tocopherol concentrations, erythrocytes from the men depleted of vitamin E were more susceptible in vitro to hemolysis induced by peroxide. ln the mid-1960’s vitamin E deficiency was described in children with various fat malabsorption syndromes, based on both low serum α-tocopherol and erythrocyte hemolysis tests [4–7]. By the mid 1970s, premature infants [8], as well as children with cystic fibrosis [9], were reported to have vitamin E deficiency, based on circulating α-tocopherol concentrations and susceptibility of erythrocytes to hemolysis. Neurologic symptoms, such as spinocerebellar ataxia, were also reported in persons with various chronic fat malabsorption syndromes [10], especially persons with abetalipoproteinemia [5, 11]. Vitamin E deficiency in persons with these disorders was suspected, but not proven despite beneficial effects of vitamin E supplementation, because the study subjects malabsorbed other nutrients.

In 1981 a person with low serum α-tocopherol and peripheral neuropathy, but without fat malabsorption or lipoprotein disorders, was described [12]. This person responded to vitamin E supplements [12, 13] and has been followed for over 40 years [14]. His serum α-tocopherol concentrations increased promptly to vitamin E supplements; some improvement in neurologic abnormalities was noted and progression of symptoms was prevented [14]. These authors and others [15–17] note that large vitamin E supplements, sufficient to normalize circulating α-tocopherol concentrations, will prevent the progression of neurologic symptoms due to vitamin E deficiency in a variety of disorders.

In the early 1990s, pharmacokinetic studies using stable-isotope labeled α-tocopherols were carried out in persons with progressive, peripheral neurologic disease, low circulating α-tocopherol concentrations, but not fat malabsorption syndromes [18, 19]. This syndrome was labeled Familial Isolated Vitamin E (FIVE) deficiency. These persons were unable to maintain plasma α-tocopherol concentrations, and some were unable to discriminate between natural and synthetic α-tocopherols [18–20]. Data from these studies showed that the liver was responsible for replacing the entire plasma α-tocopherol pool daily [20]. This inability of the body to salvage and retain α-tocopherol led to tissue and peripheral nerve α-tocopherol depletion [21], and thus caused vitamin E deficiency. Also in the early 1990s, several persons with symptoms similar to Friedreich’s ataxia were found to have various defects on chromosome 8 in the gene for the α-tocopherol transfer protein (α-TTP), and to have deficient (<1–2 micromolar) plasma α-tocopherol concentrations [22–24]. This genetic disorder that caused vitamin E deficiency was named Ataxia with Vitamin E Deficiency (AVED) and was determined to be identical to the FIVE disorder.

In summary, vitamin E deficiency symptoms include low circulating and tissue α-tocopherol concentrations. The neurologic abnormalities are best described as a progressive sensory neuropathy. Specifically, spinocerebellar ataxia, loss of motor control and lesions in muscle and peripheral nerves occur [21, 22, 24–27]. Additionally, cervical dystonia has been reported in some unsupplemented persons with AVED [28]. Eye disorders, such as retinitis pigmentosa [29, 30] and macular degeneration [31] have also been reported in AVED and in abetalipoproteinemia [32, 33]. Vitamin E deficiency can also cause anemia because erythrocytes are more susceptible to hemolysis. (Note: The in vitro erythrocyte hemolysis test is a not reliable biomarker of vitamin E deficiency because erythrocyte sensitivity to peroxide-induced hemolysis occurred in clinical disorders other than vitamin E deficiency [34]).

The spectrum of α-tocopherol deficiency symptoms is broad and takes a prolonged period (>5 years) for symptoms to occur in adults; by contrast, children show deficiency symptoms as early as the first few months to years of life [35]. Once symptoms such as severe peripheral neuropathy occur, they are difficult if not impossible to reverse. Thus, vitamin E deficiency in humans can be devastating due to its neurologic consequences. For this reason, investigators have suggested that asymptomatic persons with conditions known to lead to vitamin E deficiency be treated promptly and continuously with α-tocopherol supplements [36]. Thus, awareness by the medical community of the possible causes for human vitamin E deficiency is critical to prevent such consequences. Such disorders are discussed in the next section.

DISORDERS ASSOCIATED WITH HUMAN VITAMIN E DEFICIENCY

Human vitamin E deficiency can occur as a result of a variety of impairments in α-tocopherol transport and trafficking. α-Tocopherol absorption requires bile and pancreatic secretions, as well as transport in chylomicrons. The hepatic α-TTP is needed to salvage α-tocopherol in the liver and transfer it to plasma lipoproteins to prevent its rapid excretion from the body. α-Tocopherol transport in the circulation is dependent on trafficking and delivery of lipids and lipoproteins to tissues. As discussed below any genetic defects in these processes can impact α-tocopherol adequacy in humans.

Dietary α-tocopherol adequacy is also necessary to prevent human vitamin E deficiency. α-Tocopherol deficiency may also occur as a result of increased use of peroxidized oils [37] and potentially ultra-processed foods with low micronutrient contents [38]. Additionally, frank overall malnutrition causes low α-tocopherol and protein intakes that are associated with typical symptoms of α-tocopherol deficiency [39, 40] and oxidative damage [41]. Vitamin E supplements are also indicated in persons in weight-loss regimens, who use certain drugs to limit fat absorption [42, 43] or undergo bariatric surgery [44].

Ataxia with vitamin E deficiency (AVED)

Symptoms in persons with AVED (OMIM 277460) are solely caused by inadequate α-tocopherol concentrations, resulting from impaired or absent α-TTP function [16]. Early reports suggested that the vitamin E deficiency symptoms are more severe than those observed in malabsorption syndromes [45]. There are at least 25 known mutations to the α-TTP gene, TTPA, that result in varying degrees of AVED severity with more severe mutations truncating or preventing protein function [46, 47]. In the liver of healthy persons, α-TTP salvages newly absorbed α-tocopherol and transfers it to the hepatocyte membrane where α-tocopherol is taken up by nascent lipoproteins (very low or high density lipoproteins, VLDL or HDL, respectively), which carry the α-tocopherol into the circulation for delivery to tissues [48]. In persons with AVED, large α-tocopherol supplements (1000 mg per day) obviate the necessity for the hepatic α-TTP. During the post-prandial period, chylomicrons carry α-tocopherol into the circulation and during their catabolism α-tocopherol is non-specifically incorporated into circulating lipoproteins, which can deliver it to tissues [49].

α-TTP has been detected in extrahepatic tissues such as brain [50], eyes [51, 52], and placenta [53–55]. α-TTP concentrations are lower in these tissues than in the liver and their specific function is under investigation. Likely, α-TTP traffics α-tocopherol to membranes similarly to its function in the liver. It is unclear in AVED where there is a lack or dysfunctional α-TTP, if non-specific α-tocopherol delivery to these tissues is sufficient to prevent deficiency symptoms.

α-Tocopherol supplementation in AVED (approximately 1000 mg/d) prevents or halts progression of the neurologic disorder [14, 27, 46, 47]. Importantly, supplements containing either RRR- and all racemic (all rac)-α-tocopherols (natural or synthetic vitamin E—defined below, Figure) have been used successfully in persons with AVED [15, 29, 30, 56, 57]. Improvement occurred, or lack of progression of symptoms was prevented, so long as serum α-tocopherol concentrations remained normalized by supplement consumption [14, 15]. Further, tissue α-tocopherol concentrations in one person with AVED have been reported upon post-mortem analysis to be normalized [30].

Figure. Tocochromanols that are and are not vitamin E.

Of the tocochromanols synthesized by plants, only α-tocopherol meets the requirements for vitamin E because it is the only form shown to reverse human vitamin E deficiency symptoms. Of the natural (RRR-α-tocopherol, left column) or synthetic (right column, all rac-α-tocopherols) forms, only the α-tocopherols with the phytyl tail in the 2R-conformation are recognized by the α-TTP and therefore only 2R-α-tocopherols meet the requirements for vitamin E.

Genetic defects in lipoprotein metabolism

α-Tocopherol deficiency can occur as a result of a number of lipoprotein disorders that impact dietary fat absorption or normal lipid trafficking in the circulation [17]. Persons with abetalipoproteinemia (OMIM 200100) lack lipoproteins containing apolipoprotein B (apoB), e.g. chylomicrons, VLDL and low-density lipoproteins (LDL) [58–60]. Abetalipoproteinemia is caused by genetic defects in the microsomal triglyceride transfer protein (MTTP) [61]. Additionally, defects in apoB can result in hypobetalipoproteinemia (OMIM 615558) [62] or normotriglyceridemic abetalipoproteinemia can result if apoB is truncated [63]. Chylomicron retention disease [64] with SAR1B (Secretion Associated Ras-Related GTPase 1B) mutations, which block apoB48 and chylomicron formation, similarly cause α-tocopherol deficiency [65]

Vitamin E deficiency due to the lack of apoB-lipoproteins can be overcome by the long-term, oral administration of enormous daily α-tocopherol supplements (>100 mg/kg body weight) [66]. Possibly, circulating HDL fulfills the gap in apoB-containing lipoproteins and transports α-tocopherol to tissues [65, 67].

Fat malabsorption and cholestatic liver disease

Bile is needed for micellarization of α-tocopherol to promote its uptake into enterocytes for intestinal absorption [68]. Children with cholestatic liver disease [69] and those with cystic fibrosis [9] who secrete bile poorly, become α-tocopherol deficient due to its malabsorption. The necessity for bile secretion to promote α-tocopherol absorption was demonstrated in children with cholestasis, who upon liver transplant had normal serum α-tocopherol concentrations [70]. α-Tocopherol supplements can improve plasma α-tocopherol concentrations in persons with cystic fibrosis [71]. Persons with cholestatic liver disease need α-tocopherol supplements that self-micellarize, such as tocopherol polyetheylene glycol succinate (TPGS) [72, 73]. TPGS was effective in treating vitamin E deficiency in persons with cholestatic liver disease [74].

Premature Infants

α-Tocopherol concentrations in fetal blood and tissues in normal and premature infants [75] are considerably lower than those of the mother, apparently because of low fetal lipoprotein concentrations and poor placental α-tocopherol transfer to the fetus. Premature infants with very low birth weights (<1500 g) are α-tocopherol-deficient based on their susceptibility to hemolytic anemia, lung abnormalities and eye damage (retinopathy of prematurity) [76]. These infants require a variety of nutrients in addition to α-tocopherol. The complexity of α-tocopherol absorption, its fat-solubility and its potential for oxidation by administered iron supplements, makes α-tocopherol hard to administer and increases likelihood of high amounts remaining in the gut for increased susceptibility to sepsis and bacterial overgrowth [77]. More research is needed to evaluate the best practices to provide optimal nutrition to premature infants.

PROPOSED VITAMIN E AND TOCOCHROMANOL DEFINITIONS

As a rubric “Tocochromanols” encompasses the 4 tocopherols and 4 tocotrienols synthesized by plants, as well as numerous other similar molecules [78]. Tocopherols and tocotrienols have similar ring structures: trimethyl (α-), dimethyl (β- or γ-), and monomethyl (δ-) chromanols; however, tocopherols have a hydrophobic 16-carbon phytyl side chain, while tocotrienols have an unsaturated 16-carbon side chain. The chromanol ring forms the basis for the high antioxidant functions of the tocochromanols, as peroxyl radical scavengers.

The human dietary requirement for vitamin E, as set in the United States by the Institute of Medicine (IOM) [79], is met only by α-tocopherol (IUPAC Name: (2R)-2,5,7,8-tetramethyl-2-[(4R,8R)-4,8,12-trimethyltridecyl]-3,4-dihydrochromen-6-ol). The specific α-tocopherol features for its vitamin E activity include: the hydroxyl group at the 6 position (necessary for antioxidant activity) and three methyl groups (at positions 5, 7, 8) on the chromanol head and the phytyl tail in the 2R conformation. α-Tocopherol chemical synthesis results in an equimolar mixture of all eight stereoisomers, but only half of the stereoisomers of all rac-α-tocopherol are in the 2R-conformation (RRR-, RSR-, RRS- and RSS-). Thus, on a weight for weight basis, the ratio of the activity of supplements containing only natural RRR-α-tocopherol to those containing all rac-α-tocopherol (RRR-, RSR-, RRS-, RSS-, SRR-, SSR-, SRS- and SSS-) is 2:1. The US Food & Drug Administration defined vitamin E as 2R-α-tocopherol for labeling of fortified food and vitamin E supplements [79].

Defining vitamin E as 2R-α-tocopherol is supported by a variety of experimental and clinical data (Table). Perhaps the most convincing data is from control subjects given of deuterium (d_n)-labeled d₃-RRR- and d₆-all rac α-tocopherols, then monitoring plasma d₃:d₆ ratios, which increased from 1 in the administered dose to 2 in the plasma [80]. As discussed above, α-tocopherol concentrations are maintained in the circulation by the α-TTP and defects in this protein, or its absence, cause severe human vitamin E deficiency. Studies using deuterium-labeled RRR- and SRR-α-tocopherols in controls and persons with AVED showed that controls preferentially retained RRR-α-tocopherol, while SRR-α-tocopherol was rapidly lost from the circulation. Thus, half of the all rac α-tocopherol stereoisomers are retained in the plasma, similarly to RRR-α-tocopherol. These data suggest that the 4’ and 8’ positions in the phytyl tail are unimportant to discriminate between RRR- and all rac α-tocopherols, as was also shown experimentally in rats [81].

Table.

Tocochromanol Properties—Evidence for Vitamin E Activity of 2R-α-Tocopherols^*

Parameter	α-Tocopherol			Non-α-Tocopherols
	RRR-	2R-	2S-	ß-, γ-, δ-Tocopherols	Tocotrienols
Biokinetics	Slow [20, 80, 91–93]	Slow [80]	Fast [93, 94]	Fast [93, 95]	Very fast [96–98]
(half-lives)	~30–50 h	~30–50 h	~15 h	~15 h for γ-tocopherol	~4 h
α-TTP interactions
Ligand binding	100% [85, 99–101]		SRR-α-tocopherol,	ß-tocopherol, 38%	α-tocotrienol, 12% [85]
			11% [85]	γ-tocopherol, 9%
				δ-tocopherol, 2% [85, 101]
Ligand specificity (dissociation constant, nM)	High [102]		Low [102]	Low [102, 103]	Low [102]
	25.0 ± 2.8		545 ± 62	ß-tocopherol, 124 ± 4.7	214 ± 13
				γ-tocopherol, 266 ± 9
				δ-tocopherol, 586 ± 75
Transfer function (relative to defective α-TTP)	High [99, 104, 105]			Low [103]
Catabolism
ω-hydroxylation tested in human microsomes expressing CYP4F2, data below from [106]	Low [106–108]	Low [106]		High [106, 107]	High [106, 108]
(Vmax × 10−3)	1.1 ± 0.1	0.9 ± 0.1		ß-tocopherol, 1.4 ± 0.2	ß-tocotrienol, 7.4 ± 0.8
				γ-tocopherol, 3.8 ± 0.4	γ- tocotrienol, 21.5 ± 8.4
				δ-tocopherol, 5.4 ± 0.5	δ- tocotrienol, 13.1 ± 4.6
(Km)	306 ± 70	318 ± 74		ß-tocopherol, 134 ± 45	ß- tocotrienol, 67 ± 9
				γ-tocopherol, 155 ± 24	γ- tocotrienol, 64 ± 29
				δ-tocopherol, 56 ± 8	δ- tocotrienol, 34 ± 16
Excretion as CEHC	Slow [57, 83, 109]	Slow [82]	Fast [80, 82, 110–112]	Fast [109, 113–119]	Fast [109, 120, 121]
Rats (n = 5/group) gavaged 10 mg α-tocopherol, 10 mg γ-tocopherol, or 29.5 mg tocotrienol mixture; from [109]	α-CEHC nmol/12 h			γ-CEHC nmol/12 h	γ-CEHC nmol/12 h
	119 ± 35			1787 ± 83	2033 ± 383

^*Data using α-tocotrienol generally represents most findings. Quantitative data given as examples from representative studies.

Importantly, in persons with AVED both deuterium-labeled RRR- and SRR-α-tocopherols were lost at rates similar to those of SRR in the controls [19, 20]. These data emphasize that in AVED both RRR- and SRR-α-tocopherols are equally well absorbed and lost equally well. Further in control subjects the data shows that the α-TTP function results in the recycling of the entire plasma pool of α-tocopherol daily [20]!

Data from catabolism studies following oral administration to controls with d₃-RRR- and d₆-all rac-α-tocopherols (150 mg each) show that ratio of the isotopes in the plasma was 2:1, d3:d6, while the ratio in the urine was 1:3, excreted as α-carboxyethyl hydroxychromanol (α-CEHC) [82]. However, since α-CEHC production is only a small fraction (~1%) of the α-tocopherol absorbed [83], catabolism is not the only fate of tocochromanols. Nonetheless, studies in persons with AVED on long-term supplementation with either RRR- and all rac-α-tocopherol showed that catabolism to α-CEHC was not impaired and, in fact, increased with a test dose of all rac-α-tocopherol supplement [57]. Additionally, 2R-α-tocopherol is retained in the plasma of control subjects and is catabolized more slowly than is 2S-α-tocopherol [82]. Similar findings were reported using radioactive RRR- and SRR-α-tocopherols in rats [84].

In vitro studies of α-TTP characteristics of binding and transfer have also been examined (Table). The α-TTP ligand binding studies show that 2R-α-tocopherol is strongly preferred relative to other tocochromanols, including 2S-α-tocopherol [85]. By contrast, non-α-tocopherols and 2S-α-tocopherol are preferentially omega-hydroxylated by cytochrome P450–4F2. Non-2R-α-tocopherols are also preferentially catabolized to CEHCs and excreted.

The importance of synthetic α-tocopherol as a source of vitamin E is highlighted by studies of α-tocopherol availability in the world human food supply [86], which found that there is a shortage of adequate α-tocopherol with a nutrient gap of 31% when comparing requirements with availability. This gap could be filled by synthetic α-tocopherols because daily supplements (100 IU or more) containing natural or synthetic (RRR- or all rac-) α-tocopherol raise circulating α-tocopherol concentrations to similar levels [87]. Thus, synthetic vitamin E can be an important dietary vitamin E source globally [88]. Notably, typical multivitamins contain synthetic vitamin E.

Based on their increased rates of catabolism and excretion, non-α-tocopherols should not be called vitamin E and should be referred to by the specific name describing the structure, or as an overall rubric, “tocochromanols”. Supplements containing either natural or synthetic (RRR- or all rac-) α-tocopherol have been used to prevent or halt the progression of vitamin E deficiency symptoms in persons with AVED [15, 29, 30, 56, 57]. Moreover, half of the stereoisomers in all rac-α-tocopherol are 2R-α-tocopherols which are biologically indistinguishable from RRR-α-tocopherol. Thus, both RRR- or all rac-α-tocopherols can serve as sources of vitamin E. Therefore, I propose that vitamin E should be defined as 2R-α-tocopherol, a definition used by the US Institute of Medicine [79], the US Food & Drug Administration [89], and the European Food Safety Authority [90].

Highlights.

Human vitamin E deficiency was recognized in the 1950s

Deficiency symptoms include anemia and peripheral neuropathy

Vitamin E is defined as 2R-alpha-tocopherols (RRR-, RSR-, RRS-, RSS-)

2R-alpha-tocopherols are preferentially retained by the body

2S-alpha-tocopherols (SRR-, SSR-, SRS-, SSS-) are catabolized and excreted