Optimized high-performance liquid chromatographic procedure for the separation and determination of the main folacins...

Different methods for the extraction of folacins from biological materials and the hydrolysis of pteroylpolyglutamates prior to high-performance liquid chromatography were investigated. Acetone precipitation of proteins led to higher extraction rates of folates from biological materials as examined by using an endogenous labelling technique. It also caused less destruction of some folates but could not be combined with subsequent hydrolysis of pteroylpolyglutamates. Pteroylpolyglutamates were hydrolysed by a partially purified suspension of pteroylpolyglutamates hydrolase (PPH). Comparative studies on the folate content of rat liver revealed that complete hydrolysis of polyglutamates could also be achieved by incubating the homogenized tissue at 37 degrees C to allow endogenous PPH to act. This procedure causes interconversions of folates and is therefore not suitable for the analysis of biological materials.