Plasma and urinary concentrations of trimetoquinol by LC-MS-MS following intravenous and intra-tracheal administration to horses with heaves.

Trimetoquinol (TMQ) is a very potent and fast acting bronchodilator in horses with heaves. This study assessed the plasma and urinary concentrations of TMQ in horses with heaves following administration via the intravenous (IV, 0.2 microg/kg) and intra-tracheal (IT, 2 microg/kg) routes. TMQ was administered to six horses affected with heaves (RAO - Recurrent Airway Obstruction, used interchangeably) by the above routes and plasma and urine samples collected and stored at -20 degrees C until analyzed. Solid Phase Extraction (SPE) of TMQ was followed by highly sensitive ESI(+)-LC-MS-MS (ElectroSpray Ionization, positive mode - Liquid Chromatography - Mass Spectrometry - Mass Spectrometry); with a Limit of Detection (LOD) estimated at 1 pg/mL. Following IV administration, TMQ plasma levels peaked at 1 min at 707 pg/mL, and at 9 min at 306 pg/mL following IT administration. Our results show that TMQ plasma concentrations decline rapidly following IV administration, which is consistent with the fast onset and short duration of TMQ effect that was observed in our previous studies. On the other hand, IT administration showed a very unique plasma concentration pattern. From a regulatory standpoint, the current available TMQ ELISA kit was also used in an attempt to detect TMQ from the plasma and urine samples. We report that the ELISA kit was unable to detect TMQ from any of the samples generated in these studies.

[Determination of trimethoquinol in plasma by electrochemical detection after HPLC separation]. / Bestimmung von Trimethoquinol in Plasma mittels elektrochemischer Detektion nach HPLC-Trennung.

A high pressure liquid chromatographic method with internal analogue standardization for the determination of trimethoquinol (1-1-(3,4,5-trimethoxybenzyl)-6,7-dihydroxy-1,2,3,4-tetrahydroisoq uinoline hydrochloride) in plasma is described. The method comprises extraction from plasma at pH 9 with ethyl acetate, back-extraction into phosphoric acid and quantification after HPLC separation using a reverse phase by means of electrochemical (reductive) detection after electrochemical preoxidation. When using 5 ml plasma the lower limit of detection is 40 pg/ml. Under routine conditions the limit of determination is determined to be 0.1 ng/ml. The variation coefficients drop from about 7% in the low range to about 4% at 5 ng/ml. The determination of trimethoquinol is not impaired by its monomethylated metabolites.

Quantitative determination of trimetoquinol in plasma as its mixed pentafluoropropionyl-trimethylsilyl derivative by gas chromatography mass spectrometry.

A method is described for the determination of the bronchodilator trimetoquinol in plasma, based on selected ion monitoring gas chromatography mass spectrometry employing a deuterated trimetoquinol as internal standard. N-methyl-(trimethylsilyl)pentafluoropropionamide and N-methyl-bis-pentafluoropropionamide were prepared for derivatization of trimetoquinol into a mixed O-TMS-N-PFP derivative, which was found to be the most suitable form in terms of sensitivity, specificity and efficiency. The calibration curve was linear in the range of 2-50 ng ml-1, and the lower limit of detection was 0.1 ng ml-1 of rat plasma.