Tetrahydrobiopterin: Ubiquitous Metabolite

The Durk Pearson & Sandy Shaw®
Life Extension NewsTM
Volume 6 No. 2 • April–May 2003

Tetrahydrobiopterin: Ubiquitous Metabolite that Regulates the Superoxide Release by Endothelial Nitric Oxide Synthase

We have long wished that we could take tetrahydrobiopterin (BH4) as a dietary supplement, because it is a cofactor in many important metabolic processes. For example, it has long been known that pteridine in a reduced state is required as a cofactor in the conversion of phenylalanine to tyrosine, of tyrosine to dopamine and noradrenaline, and of tryptophan to serotonin.1 [The closest we could get to adding tetrahydrobiopterin to our phenylalanine-plus-cofactors formulations was to add folic acid, because our intermediary pathways charts suggested to us over 15 years ago that folic acid might increase the amount of tetrahydrobiopterin. As it turns out, we were pretty close.]

Recent studies have revealed that tetrahydrobiopterin (BH4) is also an essential cofactor required for the activation of nitric oxide synthase (NOS, the enzyme that makes nitric oxide from L-arginine). BH4 transfers electrons to arginine bound to the enzyme as well as stabilizing the active form of NOS and increasing the affinity of arginine for the enzyme. In addition—and this is very important—in the presence of inadequate levels of BH4, NOS becomes “uncoupled” from the production of nitric oxide, instead producing superoxide radicals but no nitric oxide! In animal and human studies, acute and chronic supplementation with BH4 has been shown to improve endothelium-dependent vasodilation.2

It has been suggested that in diseases such as diabetes, hypertension, and atherosclerosis, there is a BH4 deficiency that correlates with decreased NO synthesis and with endothelial dysfunction.3,7 Since, at low levels of BH4, more superoxide radicals are released by NOS, this in itself can explain, at least in part, the deficiency of NO, since NO reacts with superoxide to form peroxynitrite, a powerful oxidant. Thus, low levels of BH4 produce a state that is functionally like that of having almost no endothelial NOS at all, since superoxide radicals are produced, but little or no nitric oxide. Knockout mice lacking endothelial NOS develop hypertension, insulin resistance, hyperlipidemia, and augmented ischemia-reperfusion injury.3

Folate may improve vascular function by mimicking BH4 activity3

A recent paper reports evidence that 5-methyltetrahydrofolate (5-MTHF, the physiological form of folate) directly interacts with NOS to promote NO versus superoxide formation, thus improving endothelial function.4 The authors show that the 5-MTHF binds the active site of NOS and mimics the orientation of tetrahydrobiopterin. (Amazingly, however, the paper did not provide chemical structures for the two substances; hence, if you want to compare the structures yourself, you’ll have to get them somewhere else, such as The Merck Index.) They demonstrated that 5-MTHF supplementation in BH4-deficient fructose-fed rats restored NO-dependent endothelial function.

While folate is converted to 5-MTHF in healthy persons, there is a common mutation of 5,10-methylenetetrahydrofolate reductase, the enzyme that converts 5,10-methylenetetrahydrofolate to 5-MTHF, that has been reported to be a risk factor for vascular disease.5

These results suggest that folic acid supplementation may provide the protective effects of BH4 in the endothelial NOS system. We are currently recommending 800 mcg of folic acid a day. However, if you want to take more (even pregnant women have been given 5 mg/day for months in clinical studies to prevent neural-tube-defect births), just make sure that you are taking B6 and B12 along with it.

Vitamin C increases tetrahydrobiopterin levels in vitro and in vivo

Another paper6 notes that in cultured endothelial cells, L-ascorbic acid increases NOS activity, possibly via chemical stabilization of tetrahydrobiopterin (BH4). The authors conducted experiments in wild-type (C57BL/6J) and apolipoprotein E (apoE)-deficient mice and found that 26 to 28 weeks of dietary supplementation with vitamin C (1%/kg of chow) resulted in a significantly increased ratio of BH4 to 7,8-BH2/biopterin (an oxidized form of BH4) in aortas of both apoE-deficient and wild-type mice (the apoE-deficient mouse is a commonly used model of human atherosclerosis development). Vitamin C significantly decreased 7,8-BH2/biopterin levels in apoE-deficient mice, while the vitamin increased BH4 levels in wild-type mice without affecting 7,8-BH2/biopterin levels.

We have had vitamin C in our phenylalanine-plus-cofactors formulations for over 15 years because the aromatic amino acid hydroxylation requires vitamin C.


  1. Wurtman and Wurtman, eds. Nutrition and the Brain, Vol 3, p 268 (1979).
  2. Verma et al. Novel cardioprotective effects of tetrahydrobiopterin after anoxia and reoxygenation: identifying cellular targets for pharmacologic manipulation. J Thorac Cardiovasc Surg 123(6):1074-83 (2002).
  3. Vasquez-Vivar et al. The role of tetrahydrobiopterin in superoxide generation from eNOS: enzymology and physiological implications. Free Rad Res 37(2):121-7 (2003).
  4. Hyndman et al. Interaction of 5-methyltetrahydrofolate and tetrahydrobiopterin on endothelial function. Am J Physiol Heart Circ Physiol 282:H2167-72 (2002).
  5. Verhaar et al. Folates and cardiovascular disease. Arterioscler Throm Vasc Biol 22:6-13 (2002).
  6. d’Uscio et al. Long-term vitamin C treatment increases vascular tetrahydrobiopterin levels and nitric oxide synthase activity. Circ Res 92:88-95 (2003).
  7. Landmesser et al. Oxidation of tetrahydrobiopterin leads to uncoupling of endothelial cell nitric oxide synthase in hypertension. J Clin Invest 111(8):1201-9 (2003).

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