Abstract
Vitamin C (L-ascorbic acid, AA) is an essential cellular antioxidant and cofactor for several α-ketoglutarate-dependent dioxygenases. As an antioxidant, AA interacts with vitamin E to control oxidative stress. While several reports suggest an interaction of AA with folate (vitamin B9) in animals and humans, little is known about the nature of the interaction and the underlying molecular mechanisms at the cellular level. We used an untargeted metabolomics approach to study the impact of AA on the metabolome of C2C12 myoblast cells. Compared to untreated cells, treatment of C2C12 cells with AA at 100 µM resulted in enhanced concentrations of folic acid (2.5-fold) and 5-methyl-tetrahydrofolate (5-methyl-THF, 10-fold increase) whereas the relative concentrations of 10-formyl-tetrahydrofolate decreased by >90% upon AA pretreatment, indicative of increased utilization for the biosynthesis of active THF metabolites. The impact of AA on the folate-mediated one-carbon cycle further manifested itself as an increase in the levels of methionine, whose formation from homocysteine is 5-methyl-THF dependent, and an increase in thymidine, whose formation from deoxyuridine monophosphate (dUMP) is dependent on 5,10-methylene-THF. These findings shed new light on the interaction of AA with the folate-mediated one-carbon cycle and partially explain clinical findings that AA supplementation enhances erythrocyte folate status and that it may decrease serum levels of homocysteine, which is considered as a biomarker of cardiovascular disease risk.
Highlights
Vitamin C has important functions in animals and humans to maintain health and prevent disease [1]
Metabolomics analysis was performed on mouse skeletal muscle-derived C2C12 myoblasts that were AA deficient or AA supplemented
We exposed the cells to two isotopologue forms of AA, sodium ascorbate and 13 C6 -ascorbic acid, because the approach allowed us to distinguish between exposure compounds (AA and its metabolites or degradation products) and the effects of the exposure on the metabolome
Summary
Vitamin C (ascorbic acid, AA) has important functions in animals and humans to maintain health and prevent disease [1]. AA acts as a cofactor for several α-ketoglutarate-dependent dioxygenases, such as prolyl hydroxylase involved in collagen synthesis [2]. Its capacity to donate electrons gives AA antioxidant properties [1] and ability to scavenge electrophiles in Michael additions [3,4]. The antioxidant properties of AA can manifest in many ways and affect a multitude of metabolic pathways. AA supplementation in humans lowers the production and urinary excretion of lipid peroxidation-derived reactive aldehydes [5]. From metabolomics analyses of zebrafish fed a diet lacking sufficient AA, we established that AA deficiency, which causes oxidative stress, activates the purine nucleotide cycle to regenerate ATP [6].
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