Simultaneous determination of 5-aminosalicylic acid and 5-acetylaminosalicylic acid by high-performance liquid chromatography.

A high-performance liquid chromatographic method has been developed for the simultaneous determination of 5-aminosalicylic acid (5-ASA) and its main metabolite 5-acetylaminosalicylic acid (5-AcASA) in plasma, with 4-acetylaminosalicylic acid as an internal standard. Prior to extraction into ethyl acetate, 5-ASA is derivatized to 5-carbobenzyloxyaminosalicylic acid (5-CboASA). The calibration graphs for both 5-ASA and 5-CboASA are linear between 0.1 and 20.0 mumol/l. The limit of detection is 0.02 mumol/l for 5-ASA and 0.05 mumol/l for 5-AcASA. At 0.1 mumol/l, the coefficients of variation were 8.1 and 9.8% for 5-ASA and 5-AcASA, respectively. At 1.0 and 10.0 mumol/l, the coefficients of variation were 4.1% or less. The mean bias ranged from -6.2 to -2.0% for 5-ASA and from +6.0 to 0% for 5-AcASA.