5-Methyltetrahydrofolate
(dl-5-methyl-5,6,7,8-tetrahydropteroyl-tmonoglutamic acid) has been prepared directly from folic acid without intermediate isolation of tetrahydrofolic acid (1). We now report the preparation of the ethyl, n-propyl, and n-butyltetrahydrofolates as their calcium salts. EXPERIMENTAL General Chemicals were obtained from the following sources: folic acid, Koch Light Ltd., Colnbrook, Bucks; sodium borohydride, acetaldehyde, butyraldehyde, propionaldehyde, Tris buffer, Hopkin and Williams Ltd.; and DEAE-cellulose (D.E. 52), W. and R. Balston Ltd., Maidstone, Kent.
Ultraviolet spectra were determined in aqueous solution at pH 1.0, 7.0, and 13.0. Thin-layer chromatography was performed with 0.25 mm depth cellulose powder MN300, F254 (Macherey, Nagcl and Co., Duren, Germany) and developed at room temperature in the dark.
The following solvent systems were used;
(a) 0.1 M phosphate buffer, pH 7.0, containing mcrcaptoethanol (0.5% v/v).
(bl n-Propanol/water/ammonia (200: 100: 10 v/v) containing mercaptoethanol (0.5% v/v).
(c) The organic phase of 1-butanol/acetic acid/water (4:1:5 V/V) containing mercaptoethanol (0.5% v/v). Compounds were detected as quenching or fluorescent spots when viewed under ultraviolet light c~niittinq at cither 2.54 or 365 nip.. Proton magnetic resonance spectra were recorded with a Perkin-Elmer RlO spectrometer using tetramethylsilane (TMS) as internal reference.