Quantification of folate metabolites in serum using ultraperformance liquid chromatography tandem mass spectrometry

• We report UPLC/MS/MS method to simultaneously detect folate and its metabolites.
• The method is simple, fast and sensitive.
• The method can help us to monitor folate metabolic pathways more comprehensively.

Folate deficiency is considered a risk factor for many diseases such as cancer, congenital heart disease and neural tube defects (NTDs). There is a pressing need for more methods of detecting folate and its main metabolites in the human body. Here, we developed a simple, fast and sensitive ultraperformance liquid chromatography tandem mass spectrometry (UPLC/MS/MS) method for the simultaneous quantifications of folate metabolites including folic acid, 5-methyltetrahydrofolate (5-MeTHF), 5-formyltetrahydrofolate (5-FoTHF), homocysteine (Hcy), S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH). The method was validated by determining the linearity (r2 > 0.998), sensitivity (limit of detection ranged from 0.05 to 0.200 ng/mL), intra- and inter-day precision (both CV < 6%) and recovery (each analyte was >90%). The total analysis time was 7 min. Serum samples of NTD-affected pregnancies and controls from a NTD high-risk area in China were analyzed by this method, the NTD serum samples showed lower concentrations of 5-MeTHF (P < 0.05) and 5-FoTHF (P < 0.05), and higher concentrations of Hcy (P < 0.05) and SAH (P < 0.05) compared with serum samples from controls, consistent with a previous study. These results showed that the method is sensitive and reliable for simultaneous determination of six metabolites, which might indicate potential risk factors for NTDs, aid early diagnosis and provide more insights into the pathogenesis of NTDs.