A sensitive and accurate quantitative method to determine N-arachidonoyldopamine and N-oleoyldopamine in the mouse striatum using column-switching LC-MS-MS: Use of a surrogate matrix to quantify endogenous compounds

The transient receptor potential vanilloid 1 (TRPV1) channel, a nonselective Ca²⁺ and Na⁺ channel, is a molecular transducer of nociceptive stimuli. N-Arachidonoyl dopamine (NADA) and N-oleoyldopamine (OLDA), two unsaturated N-acyldopamines, are major activating endogenous TRPV1 ligands and their presence in mammalian brain tissue has been reported. However, the biological significance of NADA and OLDA remains unknown. To investigate their biological function in the nervous system, a sensitive and accurate quantitative method for determining endogenous NADA and OLDA in the brain is necessary. Thus, a column-switching liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed to quantify NADA and OLDA in mouse striatum. Mouse cerebellum tissue in which neither NADA nor OLDA were detected was used as a surrogate matrix to prepare calibrators. NADA and OLDA were extracted from mouse brain tissue by solid-phase extraction and then filtered and analyzed by LC-MS-MS with electrospray ionization in the positive ion mode. The selectivity results and comparison of calibration curves prepared with mouse cerebellum and striatum established that the former was acceptable as the surrogate matrix of the latter for analyzing NADA and OLDA. The validation results of the matrix effect, linearity, precision, accuracy, and stability were satisfactory. The limits of detection and limits of quantification were 0.125 pg mg^-1 for both analytes. This method was sensitive and accurate enough to determine endogenous concentrations of these compounds in mouse striatum and will be very useful for further study of the biological functions of NADA and OLDA and other related factors in vivo.