Ascorbate supports folate?

Folate (vitamin B₉) is an essential carrier of 1-carbon (1C) units for DNA synthesis and methylation. More specifically, this involves reduced tetrahydrofolate (THF; H₄PteGlu) derivatives which are highly sensitive to oxidation; initially to dihydrofolate (DHF; H₂PteGlu), before eventually being destroyed by irreversible scission of the C₉–N₁₀ bond. It has long been known that the antioxidant ascorbic acid (vitamin C) can reduce DHF (to THF) and protect folate from degradation ¹. Further, in humans, dietary ascorbate and THF synergistically correlated RBC folate ², and ascorbate supplementation boosted the blood response to both natural 5-methyl-THF (over 8 hours) ³ and synthetic folic acid (after 45 days) ⁴, supporting physiological relevance.

Ascorbate may affect folate bioavailability at several levels. Dietary folate is naturally present as reduced THF derivatives, which have pH-dependent stability. Under moderately low pH conditions typical of the postprandial stomach, THF is relatively stable and DHF labile, while under neutral pH the opposite is true ¹. Ascorbate is present in the stomach against a plasma gradient (~5:1) where it may protect reduced folates during digestion ^1–3. On absorption, the gut efficiently converts reduced folates to 5-methyl-THF, the major circulating form ⁵; while a portion of demethylated tissue folate may be returned to the liver, remethylated and released into bile for enterohepatic recirculation ⁶. Conversely, the gut and liver seem to have slow and variable ability (i.e. DHFR activity) to reduce synthetic (oxidised) folic acid (i.e. PteGlu), which may underlie the appearance of unmetabolised folic acid in circulation ^5,7. In tissues, folates are susceptible to various factors. For instance, the skin epidermis is relatively low in folate, but proportionally high in 5-methyl-THF ⁸, which may protect from UV-induced photosensitization reactions (unlike folic acid), while being oxidised and degraded in the process, but preserved by ascorbate ⁹. Also, human cerebrospinal fluid is rich in 5-methyl-THF (2–4x plasma) which significantly, but mildly, correlated ascorbate (r=0.284) ¹⁰. This relationship was tested in a neuronal model, where mitochondrial dysfunction increased superoxide and depleted 5-methyl-THF, which was prevented by ascorbic acid ¹⁰. Consequently, under conditions of inadequate intake or oxidative stress, ascorbate may prolong the time to folate deficiency ¹¹. Notably, polyphenol antioxidants (co-nutrients of dietary ascorbate) may also protect folic acid from photo-degradation ^12,13 and oxidation-induced malabsorption ¹⁴.

Ascorbate may also affect folate metabolism. Accordingly, associations between dietary ascorbate and RBC folate were stronger in those with genetic variants in folate pathways ²; while others have reported negative and independent associations between dietary/plasma ascorbate and homocysteine ^15,16. Moreover, in myoblasts (in folic acid medium), untargeted metabolomics showed that ascorbate particularly stimulated folate-dependent 1C metabolism, increasing levels of both methionine (5-methyl-THF-dependent) and thymidine (5,10-methylene-THF-dependent), potentially by facilitating the reductive (NADPH-dependent) steps from 10-formyl-THF to 5-methyl-THF ¹⁷. On the other hand, a trial with 500mg vitamin C (w/wo 5mg folic acid) on Italian smokers increased RBC folate (~40%), but also homocysteine (~26%) ⁴ (as have other antioxidant trials ^18,19). It has been suggested that ascorbate-induced pro-oxidant effects may inactivate dietary cobalamin (vitamin B₁₂), a cofactor in folate-dependent homocysteine remethylation ²⁰. Also noteworthy, high-dose antioxidants can inhibit exercise adaptations mediated via ROS/Nrf2 ^21–23. Nrf2 is a redox-sensitive transcription factor which regulates 100s of genes and rewires metabolism to restore redox homeostasis ^24,25; and notably can induce transsulfuration enzymes (i.e. homocysteine catabolism to cysteine) ^26,27 and redox-dependent cobalamin processing (i.e. homocysteine remethylation to methionine) ^28,29. Accordingly, human gene mutations inducing over-activation of Nrf2 are associated with low plasma homocysteine, which was presumed to result from upregulated transsulfuration, and ascorbate was considered as a Nrf2-inhibiting treatment ³⁰. Thus unbalanced antioxidant supplementation may disrupt redox signalling and potentially homocysteine metabolism.

Importantly, the colonic microbiome is also a source of folate which resists dietary deficiency ³¹, and colon-delivered 5-formyl-THF was absorbed into systemic circulation in humans ³². Colonic folate may serve important functions both locally (e.g. immune tolerance ³¹ and colorectal cancer ³³) and systemically (e.g. autoimmunity ³⁴, fatty liver ^35,36 and hyperhomocysteinemia ³⁷). While some gut bacteria produce folate, others are auxotrophic, including important butyrate-producing bacteria (e.g. Fig. 2) ³⁸, potentially linking colonic folate to butyrate production. Notably, trials with ascorbate have modulated the gut microbiome ³⁹ and increased SCFA/butyrate production ⁴⁰. In vitro, antioxidants can even maintain the viability of highly oxygen-sensitive anaerobic bacteria in ambient air ^41,42 and enhance butyrate production ⁴³. So could unabsorbed antioxidants/ascorbate support a favourable colonic redox environment and bioavailability of microbial folate?

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