Free and total para-aminobenzoic acid analysis in plants with high-performance liquid chromatography/tandem mass spectrometry

para-Aminobenzoic acid (PABA), a precursor in the synthesis of folates in plants, is determined by liquid chromatography/tandem mass spectrometry (LC/MS/MS). In plants PABA can be converted into its beta-D-glucopyranosyl ester (PABA-Glc) and can also exist in its free form. In this work, we developed and validated a quantitative method to study free and total PABA in plants. The total PABA (free PABA plus PABA-Glc) can be evaluated after acid hydrolysis at 80 degrees C for 2 hours. The plant material is homogenized and the PABA content is quantified using the standard addition procedure. The validated method is selective, sensitive, simple, accurate, has a recovery between 99.6 to 102.5%, is reproducible (RSD between 1.4 and 4.4%), and is linear between 2.5 and 1538 ng/mL. Free and total PABA determinations in five vegetables showed that different plant species had different amounts of free and total PABA, and that the ratios of total versus free PABA were also variable. This new method could be valuable for studies of folate synthesis in plants.