Choline: The forgotten essential nutrient

By:
Anthea Van Parys

Centre for Nutrition, Department of Clinical Science, University of Bergen

Introduction

Choline (2-hydroxyethyl-trimethyl-ammonium salt) is an essential nutrient needed for the regular operation of all cells and has a wide variety of functions including roles in cholinergic neurotransmission, lipid transport, membrane synthesis, and one-carbon metabolism (1). Choline was first isolated from ox bile in 1862, hence the name choline which is derived from the Greek term for bile (i.e. chole) (2). Humans can obtain choline through endogenous synthesis; however, this route is not sufficient to support biological requirements. In the 1930s, its nutritional importance was first recognized, however, it was only in 1998 that choline was first recognized as an essential nutrient by the US Institute of Medicine (currently known as the National Academies of Medicine, NAM) (3).

In Norway, dietary recommendations for choline are currently unavailable (4) and choline is not included in the Norwegian food database (www.matvaretabellen.no). Fortunately, choline will be included in the new Nordic Nutrition Recommendations of 2022. Therefore, this manuscript aims to provide an introduction and general overview of this essential nutrient.

Individual choline forms

In the body and food items, choline occurs in both water- (free choline, glycerophosphocholine (GPC), and phosphocholine) and lipid-soluble forms (lysophosphatidylcholine (lysoPC), phosphatidylcholine (PC), and sphingomyelin) (Figure 1) (5). Free choline is a precursor for betaine, which is a methyl donor and links choline to the one-carbon metabolism. It is also a precursor for the neurotransmitter acetylcholine, which is important for brain functions such as memory, mood, sleep, learning, and muscle control. Finally, free choline can form PC via the cytidine diphosphate (CDP)-choline pathway. Another physiologically interesting choline form is PC (or lecithin). PC is an essential membrane phospholipid and accounts for >50% of phospholipids in most mammalian membranes. It is required for VLDL synthesis and secretion in the liver as well as for bile secretion.

Figure 1 Structures of water- and lipid-soluble choline forms. The dashed box indicates free choline. R indicates a fatty acid chain. (Adapted from Wallace et al. (2))

Absorption and transport

After digestion by pancreatic and mucosal enzymes, dietary choline is absorbed in the small intestine in the form of lysoPC or choline. The latter is transported into the enterocyte by sodium-independent carrier-mediated transport or passive diffusion when intestinal choline concentrations are high. Afterward, free choline enters the portal circulation and is taken up by the liver. Here, the majority enters the CDP-pathway for PC formation via phosphorylation to phosphocholine. The absorption of lipid-soluble choline forms differs from that of the water soluble-forms. PC is hydrolyzed by phospholipase A2 to lysoPC before absorption in the enterocyte while sphingomyelin is taken up intact. After uptake, lysopPC can be reacetylated to PC or can be broken down to GPC. The lipid-soluble choline forms enter the lymphatic system packed in chylomicrons. Hereby, they reach peripheral organs such as adipose and muscle tissue before entering the liver (1,6,7).

Figure 2 Simplified overview of the four main fates of choline. PC indicates phosphatidylcholine; PE, phosphatidylethanolamine; PEMT, phosphatidylethanolamine-N-methyltransferase; SAM, S-adenosylmethionine; TMAO, trimethylamine N-oxide

Choline metabolism and metabolic implications

In general, choline has four main fates, which include the synthesis of PC, betaine, acetylcholine, and trimethylamine N-oxide (TMAO). A simplified overview of the choline metabolism is provided in Figure 2.

CDP-choline and PEMT pathway

Figure 3 The CDP- choline and PEMT pathway. The enzymes indicated are (1) choline kinase, (2) CTP:phosphocholine cytidylyltransferase, (3) CDP-choline:1,2-diacylglycerol cholinephosphotransferase, (4) phosphatidylethanolamine-N-methyltransferase, (5) various phospholipase and lysophospholipases.

CDP indicates cytidine diphosphate; PC, phosphatidylcholine; PDME, phosphatidyldimethylethanolamine; PE, phosphatidylethanolamine; PEMT, phosphatidylethanolamine N-methyltransferase; PMME, phosphatidylmonoethylethanolamine; SAH, S-adenosylhomocysteine; SAM, S-adenosylmethionine. Adapted from Cole et al. (8).

The major fate of both endogenous and exogenous choline is conversion to PC. This process takes place in all mammalian, nucleated cells. It is therefore not surprising that PC accounts for ~95% of the total choline pool in most animal tissues. The major pathway for PC synthesis is the CDP-choline or Kennedy pathway where PC is formed from free choline in three enzymatic steps. PC can also be formed endogenously via the phosphatidylethanolamine-N-methyltransferase (PEMT) pathway which is only quantitatively significant in the liver and accounts for ~30% of total hepatic PC synthesis. This PC is secreted into bile, secreted into VLDL, or used for the formation of high-density lipoproteins in plasma. The PEMT pathway is the only way, apart from the diet, to derive new choline molecules. The CDP-choline and PEMT are interrelated and highly regulated to maintain optimal choline homeostasis (1,7,8). An overview of both pathways is provided in Figure 3.

One-carbon metabolism

In the liver and kidneys, choline can be converted to betaine in a two-step reaction which links choline to the folate-dependent one-carbon metabolism. Betaine serves as a methyl donor in the betaine-homocysteine methyltransferase reaction which catalyzes the conversion of homocysteine to methionine (Figure 4). Methyl tetrahydrofolate (mTHF) is another methyl donor for this conversion and this reaction is catalyzed by methionine synthase. The metabolism of choline, betaine, folate, methionine, and also vitamin B6, and vitamin B12 which act as cofactors (not shown), is interrelated meaning that disturbances in one will lead to changes in the other. A folate-deficient diet increases dietary choline requirements and vice versa. The remethylation of homocysteine to methionine, and thus choline and betaine concentrations, also influence the concentration of the universal methyl donor S-adenosylmethionine (SAM). Changes in SAM may lead to altered DNA methylation which in turn affects gene transcription, genomic imprinting, and genomic stability.

Other metabolic fates

In selected tissues, such as the placenta and neurons, choline can be acetylated to acetylcholine. This neurotransmitter is important for brain functions such as memory, mood, sleep, learning, and muscle control (1,9).

Finally, choline and choline-containing compounds can undergo bacterial conversion in the human intestine to form trimethylamine (TMA). This reaction is catalyzed by TMA-lyase and requires the presence of microbiota. In the liver, TMA is converted to TMAO by flavin monooxygenase (FMO) 1 and 3. TMAO acts as an organic osmolyte and promotes protein folding in the endoplasmatic reticulum by acting as a chemical chaperone. Most importantly, TMAO has recently been associated with cardiovascular disease (CVD), kidney disease, type 2 diabetes mellitus, and certain types of cancer. Whether TMAO is a cause or rather a mediating factor of these adverse health outcomes remains unclear (10,11).

Figure 4 Simplified presentation of the interaction between choline and one-carbon metabolism. The enzymes indicated are (1) choline dehydrogenase, (2) betaine aldehyde dehydrogenase, (3) betaine-homocysteine-s-methyltransferase, (4) dimethylglycine dehydrogenase, (5) sarcosine dehydrogenase, (6) methionine synthase, (7) phosphatidylethanolamine-N-methyltransferase.

DMG indicates dimethylglycine; mTHF, methyl tetrahydrofolate; PC, phosphatidylcholine; PE, phosphatidylethanolamine; SAH, S-adenosylhomocysteine; SAM, S-adenosylmethionine; THF, tetrahydrofolate.

Dietary sources, recommendations, and intake

Dietary sources

In 2004, the US Department of Agriculture (USDA) released the first Database for the Choline Content of Common Foods. This database included values for the individual choline forms free choline, GPC, phosphocholine, PC, and sphingomyelin as well as total choline and betaine in food items. The database was updated and expanded in 2008 (12). Choline is widely spread in food items and good sources are eggs, beef, chicken, fish, milk, and certain plant foods such as cruciferous vegetables and some beans. In general, animal-derived food items contain more choline per unit weight compared to plant foods. Foods can also contain betaine, even though it cannot be converted to choline, it can have a choline-sparing effect since it is a methyl donor. The majority of choline in food items is in the form of PC, as part of the cell membrane. Additionally, lecithin (i.e. PC) is often added to prepacked foods and can, therefore, be a hidden choline source. The information on choline content of food items outside North-America is very limited, complicating the accurate estimation of choline intake worldwide (5,13). Choline content is not included in the Norwegian food composition table (www.matvaretabellen.no).

Recommendations and intake

There is some uncertainty regarding the required amount of dietary choline. In 1998, NAM published the first adequate intake (AI) values for choline (Table 1). These are based on a depletion-repletion study where men were fed a choline-deficient diet with adequate methionine, folate, and vitamin B12 for three weeks. This resulted in decreased choline stores and liver damage (defined as elevated alanine aminotransferase). Additionally, individuals receiving total parenteral nutrition deficient in choline but containing adequate amounts of methionine and folate developed fatty liver and liver damage. This was resolved in most individuals after dietary choline was provided (3).

In 2016, the European Food Safety Authority (EFSA) issued the Dietary Reference Values for Choline (Table 1). Again, only an AI was set for choline due to a lack of data to determine an estimated average requirement. The recommended AI values are based on observed mean intakes in healthy populations from 12 national surveys in nine EU countries. Additionally, EFSA included a study that reported on the amount of choline needed to replete 70% of choline-depleted subjects with signs of organ dysfunction in their assessment (14). Until today, there are no dietary recommendations for choline in the Nordic countries (4).

Currently, information on dietary choline intake is mostly available from European and Northern American countries. EFSA reported the mean estimated total choline intakes ranging from 269 – 468 mg/d in adults. Reported intakes for men were slightly higher compared to women (332 – 468 mg/d and 269 – 404 mg/d respectively) (14). Studies conducted in the USA, Canada, and New Zealand reported total choline intakes ranging from 312 – 421 mg/d in men and 258 – 314 mg/d in women (5,15–18). Interestingly, it is a common finding that the mean total choline intakes are below the AIs of both NAM and EFSA. Given the definition of an AI, it is not possible to draw any conclusion on the adequacy of choline intake. Additionally, the recommended intakes from both institutions are based on very little data. This lack of data could be caused by the lack of food composition databases to estimate dietary choline intake.

Table 1. Adequate choline intake (mg/d) in USA and EU*.

USA

EU

Male

Female

Infant (0-6 months)

125

125

120

Infant (7-12 months)

150

1501

160

Children (1-14 years)

200-375

200-375

140-340

Adolescents and adults (≥ 15 years)

550

400-425

400

Pregnancy

450

480

Lactation

550

520

* USA recommendations obtained from the National Academies of Medicine (3). EU recommendations obtained from the European Food Safety Authority (14).

Choline intake in the Norwegian population

So far, very few studies have reported on choline intake in the Norwegian population which is most likely due to lack of food composition data. Øyen et al. (19) investigated choline intake in middle-aged (46-49 years) and elderly (71-74 years) women (n = 2649) and men (n = 1983) from the Hordaland Health Study (HUSK). The median energy-adjusted choline intake was 255 (interquartile range (IQR) 63) mg/d in middle-aged women compared to 259 (74) mg/d in middle-aged men and 265 (61) mg/d compared to 258 (61) mg/d in elderly women and men respectively. In the Western Norway Coronary Angiography Cohort, median energy-adjusted choline intake of patients with stable angina pectoris (n = 4164) totaled 247 (IQR 62.9) mg/d (20). Finally, the median energy-adjusted choline intake for 1981 participants with stable angina pectoris of the Western Norway B-vitamin Intervention Trial (WENBIT) was 288 (IQR 71) mg/d (21). Note that all these intakes are below the recommended AI set by both the NAM and EFSA.

In the HUSK study, dietary choline intake was positively correlated with the intake of meat, fish, eggs, milk, fiber, and protein and negatively correlated with intake of carbohydrates and fat (19). Additionally, positive correlations for intakes of fruit, berries, and vegetables with dietary choline were observed in the WENBIT cohort (21). According to the Developments in the Norwegian Diet (Utviklingen i norsk kosthold) report, intakes of both milk and fish have decreased drastically over the past years. Intakes of meat, fruit, berries, and vegetables also show a downward trend (22). These developments, together with estimated dietary choline intakes being below the recommended AI, indicate a possible insufficient choline intake in Norway. Norwegian food composition tables should be updated with choline values to allow an accurate estimation of the current choline intake in the Norwegian population and eventually, the composition of dietary recommendations.

Factors influencing choline requirements

Choline requirement shows large interindividual variability since it is influenced by several factors. As mentioned earlier, folate, methionine, vitamin B, and choline metabolism are interrelated. A folate-deficient diet leads to increased choline requirements and vice versa. Additionally, vitamin B12 deficiency leads to less folate recycling, increasing choline requirements. Secondly, the promotor for the PEMT gene is estrogen-responsive causing higher choline requirements for men and postmenopausal women. Next, the individual choline requirements depend on the ability to synthesize choline de novo. Several single nucleotide polymorphisms (SNPs) have been identified in genes in the choline and folate metabolism (e.g. PEMT, 5,10 – methylenetetrahydrofolate dehydrogenase 1958A (MTHFD1), methyltetrahydrofolate reductase (MTHFR), etc.). This genetic variability may influence an individual’s susceptibility to choline deficiency. Finally, choline requirements increase in pregnant and lactating women. Adequate choline intake is essential for fetal brain development and maintenance of plasma homocysteine concentrations (1,14,23).

Choline intake and health outcomes

Choline deficiency

It has been shown that choline deficiency leads in both animals and humans to fat accumulation in the liver (non-alcoholic fatty liver disease (NAFLD)) and liver and muscle damage. As mentioned earlier, PC is necessary for both synthesis and secretion of VLDL in the liver. An insufficient dietary choline intake, therefore, leads to accumulation of hepatic triglycerides and eventually to NAFLD. The individual susceptibility to develop liver damage is related to the previously mentioned SNPs in genes of enzymes involved in choline and one-carbon metabolism (1,2).

Toxicity

The tolerable upper limit (UL) of dietary choline intake has been set to 3500 mg/d for adults by the NAM. This is based on a study performed on seven patients with Alzheimer’s disease where admission of 7.5 g choline per day resulted in hypotension, nausea, and diarrhea. Unfortunately, not enough data was available to determine a UL for children. EFSA has not defined a UL for adults nor children. Excess choline intake is associated with a fishy body odor, excessive sweating and salivation, hypotension, vomiting, and liver toxicity (2,14).

Cardiovascular disease

Dietary choline has been linked to CVD through various mechanisms. The first possible mechanism is through the conversion of choline to betaine and its role in one-carbon metabolism. Since betaine is a methyl donor in the remethylation of homocysteine to methionine, high choline levels have been associated with lower total homocysteine. This could be associated with decreased risk of CVD based on the assumption that total homocysteine causes CVD. However, this assumption is still a topic of discussion (9,24).

Secondly, choline, and more specifically PC, could influence lipid metabolism, which may be involved in the development of CVD. On one hand, PC is an essential component of VLDL and thus essential for cholesterol and triglyceride transport from the liver to the vessels. On the other hand, the betaine- homocysteine S-methyltransferase enzyme could increase VLDL secretion and betaine has been associated with increased plasma LDL and decreased HDL cholesterol. Current evidence is not sufficient to draw any firm conclusions and larger trials are needed (24).

Finally, choline-containing nutrients can be metabolized by the gut microbiota to TMA and finally to TMAO in the liver. High TMAO levels have been described to have proatherogenic and prothrombotic effects. Most, but not all, observational studies found that high plasma TMAO is associated with increased CVD risk (2,24). However, incident CVD was not associated with dietary choline or betaine in a recent review by Meyer et al (25). Additionally, reverse causality and/ or confounding have not been addressed adequately. TMAO could be a biomarker rather than a causative factor of CVD. It is also possible that bacteria producing TMAO additionally produce some atherogenic factor (1,2,24).

So far, most studies point in the direction of a higher risk of CVD with increased dietary choline intake which is in line with the TMAO and lipid metabolism hypothesis instead of the lower risk expected based on the homocysteine hypothesis (24). However, this is still a topic of discussion and possible underlying mechanisms remain unclear.

Neurological development and function

Choline plays a critical role in the development of the central nervous system and demand increases during pregnancy and lactation. Several rodent studies suggest a causal relationship between perinatal and early postnatal choline intake and cognitive function in offspring. However, these findings have yet to be confirmed in humans. It has however been shown in humans that higher periconceptional choline intake is associated with a large decrease in risk for neural tube defects (NTD). The potential underlying mechanisms could be the role of choline as a methyl donor. Increased methyl-donor supply and DNA methylation status have been suggested to be associated with NTD (1,9,24).

In animal models, high choline intake during gestation and early childhood may enhance cognitive function across the lifespan. However, this has yet to be confirmed in humans. In general, there are few and inconsistent results regarding choline intake and cognitive function. Nevertheless, might choline have some positive effects on cognitive function later in life. Phospholipids in neuronal membranes and thus structural integrity decrease with age. Choline, and more specifically PC, could serve as a phospholipid precursor and might, therefore, influence cognitive function in the elderly. Additionally, both choline and docosahexaenoic acid (DHA) promote neuronal membrane and synapse formation and have been shown to decrease the formation of amyloid plaque in the mice brain. More and larger trials are needed to conclude on the effect of dietary choline on neurological development and function (1,2,9,24).

Conclusions

Choline is an essential nutrient in humans and is associated with many health outcomes. However, findings on the effect of dietary choline on several of these outcomes are often too few and/ or inconsistent to draw any conclusions. Large prospective cohort and intervention studies should be conducted to gain insight into this association and the underlying mechanisms.

There is a need for the inclusion of choline in food composition tables both in Norway and globally. This will lead to better mapping and understanding of the intake of not only total choline but also of the individual choline forms. Such knowledge is crucial to establish updated, accurate, and much-needed dietary recommendations.

I have no conflict of interest to declare.

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