Development and validation of a high-performance liquid chromatography-mass spectrometry assay for methylxanthines...

A procedure based on liquid chromatography-mass spectrometry (LC-MS) is described for determination of caffeine, theobromine, theophylline, taurine in different dietary supplements. After addition of tryptophan as internal standard, both solid and liquid specimens were extracted with 4 ml of hexane/isopropanol (9:1). Chromatography was performed on a C18 reversed-phase column using water/methanol/acetic acid (75:20:5, v/v/v) as a mobile phase. Analytes were determined in LC-MS single ion monitoring mode with atmospheric pressure ionization-electrospray (ESI) interface. The method was validated in the range 0.1-500 and 0.06-500 microg/ml or microg/g for taurine and caffeine, respectively; 0.06-100 microg/ml or microg/g for theobromine and theophylline. Mean recoveries ranged between 70.1 and 94.4% for different analytes. The quantification limits were 0.1 microg/ml or microg/g for taurine and 0.06 microg/ml or microg/g for methylxanthines either in liquid samples or in solid samples. The method was applied to the analysis of various dietary supplements containing methylxanthines and taurine. Energetic drinks contained amounts of taurine in the range of hundreds to thousands microg/ml and ten times lower amounts of caffeine. Conversely, herbal powders, tablets and capsules mainly contained mg amounts of caffeine per gram of product with the other two methylxanthines in the range of ten to hundred microg/g.