Unmetabolized folic acid is detected in nearly all serum samples from US children, adolescents, and adults

J Nutr. 2015 Mar;145(3):520-31. doi: 10.3945/jn.114.201210. Epub 2014 Dec 10.


Background: Serum total folate consists mainly of 5-methyltetrahydrofolate (5-methylTHF). Unmetabolized folic acid (UMFA) may occur in persons consuming folic acid-fortified foods or supplements.

Objectives: We describe serum 5-methylTHF and UMFA concentrations in the US population ≥1 y of age by demographic variables and fasting time, stratified by folic acid-containing dietary supplement use. We also evaluate factors associated with UMFA concentrations >1 nmol/L.

Methods: Serum samples from the cross-sectional NHANES 2007-2008 were measured for 5-methylTHF (n = 2734) and UMFA (n = 2707) by HPLC-tandem mass spectrometry.

Results: In supplement users compared with nonusers, we found significantly higher geometric mean concentrations of 5-methylTHF (48.4 and 30.7 nmol/L, respectively) and UMFA (1.54 and 0.794 nmol/L, respectively). UMFA concentrations were detectable (>0.3 nmol/L) in >95% of supplement users and nonusers, regardless of demographic or fasting characteristics; concentrations differed significantly by age and fasting time, but not by sex and race-ethnicity, both in supplement users and nonusers. The prevalence of UMFA concentrations >1 nmol/L was 33.2% overall and 21.0% in fasting (≥8 h) adults (≥20 y of age). Using multiple logistic regression analysis, UMFA concentrations >1 nmol/L were associated with being older, non-Hispanic black, nonfasting (<8 h), having smaller body surface area, higher total folic acid intake (diet and supplements), and higher red blood cell folate concentrations. In fasting adults, a decrease in the mean daily alcohol consumption was also associated with increased odds of UMFA concentrations >1 nmol/L.

Conclusions: UMFA detection was nearly ubiquitous, and concentrations >1 nmol/L were largely but not entirely explained by fasting status and by total folic acid intake from diet and supplements. These new UMFA data in US persons ≥1 y of age provide much-needed information on this vitamer in a fortified population with relatively high use of dietary supplements.

Keywords: 5-methyltetrahydrofolate; HPLC–tandem mass spectrometry; NHANES; serum folate; unmetabolized folic acid.

MeSH terms

  • Adolescent
  • Adult
  • Biomarkers / blood
  • Child
  • Child, Preschool
  • Cross-Sectional Studies
  • Dietary Supplements
  • Female
  • Folic Acid / administration & dosage
  • Folic Acid / blood*
  • Food, Fortified*
  • Humans
  • Infant
  • Logistic Models
  • Male
  • Middle Aged
  • Multivariate Analysis
  • Nutrition Surveys
  • Tandem Mass Spectrometry
  • Tetrahydrofolates / blood
  • United States
  • Young Adult


  • Biomarkers
  • Tetrahydrofolates
  • Folic Acid
  • 5-methyltetrahydrofolate