New screening method for basic compounds in urine by on-line extraction-high-performance liquid chromatography with photodiode-array detection

A fully automated, qualitative screening HPLC method for the identification of basic compounds in urine has been developed. A 1-ml volume of urine was extracted by on-line extraction and separated on two coupled strong cation-exchange (SCX) columns (2 × LunaSCX, 150 mm × 4.6 mm, 5 μm) under isocratic conditions. The mobile phase consisted of a mixture of potassium dihydrogenphosphate buffer (pH 2.3) and acetonitrile. The use of photodiode-array detection (DAD, λ = 190-800 nm) gave access to a library of approximately 2600 toxicologically relevant compounds. The validated method is reliable, simple and in addition successfully proven with the analysis of real biological specimen for the routine use.