Analysis of some folate monoglutamates by high-performance liquid chromatography-mass spectrometry. I

The use of high-performance liquid chromatography for the determination of folates is well documented. The methods used are based on reversed-phase ch romatography with UV and/or fluorescence detection. In many instances it is difficult to reach the required detection limits and many of the methods l ack specificity. High-performance liquid chromatography-mass spectrometry ( LC-MS) has become a well established analytical tool in: modern laboratorie s. It can offer superior specificity and often lower detection limits than conventional LC detection methods and thus is ideally suited to the analysi s of folates. A system capable of separating the four main folates [folic a cid (pteroylglutamic acid, PGA)I, 5-methyltetrahydrofolic acid, terahydrofo lic acid and 5- and/or 10-formyltetrahydrofolic acid [folinic acid (CHOTHF) ] using LC-MS with negative ion electrospray has been developed. After opti misation, a system using a 12.5 cmx3 mm, 3 mu m Hypersil ODS column and a m obile phase of 2.5 mM acetic acid-acetonitrile (88:12, v/v) was developed w hich was capable of separating the four folates in under 10 min without the use of a gradient system. Extraction of the folates is by heat treatment a nd sample clean-up is by solid-phase extraction (C-18). On column limits of confirmation are 0.6 ng for PGA and CHOTHF. (C) 1999 Elsevier Science B.V. All rights reserved.