Determination of Folates in Rice

A method was developed to determined folic acid (FA), 5-methyltetrahydrofolate (5-MTHF), tetrahydrofolate (THF), 5-Formyltetrahydrofolate (5-CHOTHF) and 10-Formyltetrahydrofolate (10-CHOTHF) using Ultra Performance Liquid Chromatography with a Mass Spectrometer Detector (UPLC-MS). The method was developed according to the literature with different transitions. These transitions were selected for the most intense daughter ion. The parent ion, fragment ion and the conditions are described in table 1. The extraction method included 3-enzimes and SPE to clean the matrix effects. SPE involved several columns but was Oasis form Waters that provided better results. Labeled internal standard were used. 13C-FA was used for FA, 5-CHOTHF and 10-CHOTHF while 13C-5-MTHF was used for 5-MTHF and THF. Analytes were separated thought an Acquity HSS T3 from Waters with water: acetonitrile gradient for 9 minutes and analyzed by negative ion electrospray (ESI-) Multiple Reaction Monitoring (MRM). Three types of rice were tested and values were confirmed with SRM 1849a. This Certified Reference Material has values for FA and 5-MTHF determined by LC/MS/MS.Comparing the results obtained for the reference material analysed with the values present in the certificate we can conclude that the extraction method and chromatographic conditions are suitable for our samples. Brown rice presents the highest content in 5-MTHF and FA. The lowest content of all analysed folate species was found in the rice bran. Brown rice seems to be a better source of folate than whole grain rice, especially for folic acid and 5-methyltetrahydrofolate. However for 10-Formyltetrahydrofolate whole brain rice has a bigger content than brown rice.This work was done with the financial support from the Portuguese Innovation Agency (ADI) in the frame of project 23290 – ValCAP