Identification and dynamic analysis of the purine alkaloids in rat plasma after oral administration of green tea by liquid chromatography hybrid ion trap time-of-flight mass spectrometry.

A liquid chromatography hybrid ion trap time-of-flight mass spectrometric (LC-IT-TOF-MS) method was developed and validated for identification and simultaneous determination of the potential bioactive components from green tea in rat plasma. The plasma samples were extracted by liquid-liquid extraction with ethyl acetate and separated on Shim-pack XR-ODS II column by a gradient elution within a runtime of 8.0 min. The mobile phase consisted of A (0.1% formic acid in acetonitrile) and B (0.1% formic acid in water) at a flow rate of 0.4 ml/min. Two prototype components and one metabolite were successfully identified as caffeine, theobromine and theophylline according to their retention times, accurate molecule weight, and major fragment ions. Then they were determined with the addition of two internal standards, hypoxanthine and paracetamol. The linear range was 10-10,000 ng/ml for caffeine, 2.0-2000 ng/ml for theobromine and 1.0-1000 ng/ml for theophylline, respectively. Intra-day and inter-day precision were within 6.0% and 10.9%, and accuracy was less than 4.8% and 6.5%, respectively. The validated method was successfully applied to investigate the dynamic change rules of caffeine, theobromine and theophylline in rat plasma after oral administration of caffeine, theobromine and green tea extract. The comparative analysis of the pharmacokinetic parameters indicated that there were obvious differences between green tea extract administration and single substances administration.