Determination of caffeic acid in rabbit plasma by high-performance liquid chromatography

A simple and sensitive high-performance liquid chromatographic method involving UV detection was developed for determination of caffeic acid in rabbit plasma. A Lichrosphere CN column (250 mm x 4 mm I.D., 5 microns) was used as the stationary phase and the mobile phase consisted of 2% acetic acid solution at a flow-rate of 1.0 ml/min. The UV absorbance was monitored at 320 nm. The plasma sample was acidified by the addition of 0.01 parts of concentrated phosphoric acid (85%) to maintain caffeic acid stability. After a simple clean-up procedure, the limit of quantitation achieved was 0.1 micrograms/ml, and the standard curve was found to be linear over the concentration ranges of 0.1-2.0 micrograms/ml and 0.1-40 micrograms/ml. The coefficient of variation for within- and between-run precision and accuracy was less than 10%, and the recovery was 82.3%.