Folate species and catabolites



Method(s): LC-MS/MS (1).
What is measured: 5-Methyl-tetrahydrofolate (mTHF), 5-formyl-tetrahydrofolate (fTHF), folic acid (FA), 4-alfa-hydroxy-5-methyl-tetrahydrofolate (hmTHF) and the folate catabolites, para-aminobenzoylglutamic acid (pABG), and acetyl-para-aminobenzoylglutamic acid (apABG).
The prevailing folate form in serum/plasma is mTHF.

What is measured on the same platform, click here.
Platform E: mTHF, fTHF, FA, hmTHF, pABG, apABG

Performance of the assay

Lower limit of detection (LOD) (mTHF/fTHF/FA/hmTHF/pABG/apABG): 0.07/0.52/0.27/0.20/0.08/0.13 nmol/L.
Within-day CV: 5-9/8-9/7-8/6-10/6-9/5-6 %; between-day CV: 4-5/6-11/6-12/6-7/3-10/5-9 %.

Indication(s)

Assessment of folate status.

Specimen, collection and processing

Patient/subject: 5-Methyl-tetrahydrofolate (mTHF, the main folate species in serum/plasma), increases after a folate rich meal, and the concentration reflects (in contrast to erythrocyte folate) short-term folate status. Folic acid (FA), the synthetic folate form, is normally not present in human serum/plasma, but becomes detectable immediately after intake of supplements containing FA.
Matrix: Serum or EDTA plasma.
Volume: Minimum volume is 60 µL, but 200 µL is optimal and allows reanalysis.

Stability and recovery of degraded folate

mTHF in serum/plasma is degraded at room temperature, but also in samples frozen at -20 °C.
At room temperature and in the absence of ascorbic acid, 5mTHF is oxidized to and can be recovered as hmTHF within days of storage (moderate degradation). The oxidation proceeds faster in EDTA plasma than in serum (2).
In samples stored at -20 °C, there seems to be a decline in concentration (to about 70 %) during the first year, and the concentration is relatively stable thereafter. In some samples stored frozen for years, folate is not detected. In such samples, folate is degraded beyond hmTHF, but can be almost completely recovered as pABG equivalents by an assay involving oxidation and mild acid hydrolysis (3, 4).

Transportation; for general instruction on transportation, click here.

Frozen, on dry ice.

Reported values, interpretation

Reported values (mTHF/fTHF/FA/hmTHF/pABG/apABG): >7.5/0-2/0-2/≤[mTHF in fresh samples]/0-0.5/0.01-1 nmol/L.
Intraclass correlation coefficients (ICC): na.

Literature

1. Hannisdal, R., Ueland, P.M., and Svardal, A. (2009). Liquid chromatography-tandem mass spectrometry analysis of folate and folate catabolites in human serum. Clin Chem 55, 1147-154
2. Hannisdal, R., Ueland, P.M., Eussen, S., Svardal, A., and Hustad, S. (2009). Analytical recovery of folate degradation products formed in human serum and plasma at room temperature. J Nutr 139, 1415-18.
3. Hannisdal, R., Svardal, A., and Ueland, P.M. (2008). Measurement of folate in fresh and archival serum samples as p-aminobenzoylglutamate equivalents. Clin Chem 54, 665-672.
4. Hannisdal, R., Gislefoss, R.E., Grimsrud, T.K., Hustad, S., Morkrid, L., and Ueland, P.M. (2010). Analytical recovery of folate and its degradation products in human serum stored at -25 degrees C for up to 29 years. J Nutr 140, 522-26.