Quantification of monoamine neurotransmitters and melatonin in sea lamprey brain tissues by high performance liquid chromatography-electrospray ionization tandem mass spectrometry

Abstract

A rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for simultaneous quantification of three monoamine neurotransmitters (NTs) and melatonin in sea lamprey brain tissues. Separation was performed on a reversed-phase column with mobile phases of 1mM perfluoroheptanoic acid (PFHA) water solution/acetonitrile and mass spectra were acquired in positive electrospray ionization multiple reaction monitoring (MRM) mode. Solid-phase extraction (SPE) was employed to purify and extract the target compounds from the tissue samples. The matrix effects as well as the influence of two extraction methods, protein precipitation (PPT) and SPE, on matrix effects were examined for the first time on the quantification of NTs from brain tissue extracts. The matrix effects with SPE (2.4 to -14.9%) were about 30% lower on average than those with the ACN PPT method (-29 to -38%). The recoveries of three types of SPE cartridges were tested and Bond-Elut C18 cartridge was selected to process the samples because of its good extraction efficiencies (71.3-95.3%) and low matrix effects (-6.6 to -14.9%) for all four analytes. This method exhibited excellent linearity for all analytes with regression coefficients higher than 0.99. The limits of detection were between 0.03 ng/mL (melatonin) and 0.14 ng/mL (norepinephrine). The precisions, expressed as coefficients of variation (CV), ranged from 4.8 to 14.1% for intra-day analyses and from 6.1 to 16.2% for inter-day analyses. Brain tissues from 360 sea lampreys were analyzed by the developed method and the concentrations for four analytes were found to be at the level of nanogram per gram of brain tissues. To our knowledge, this is the first report on the quantification of NTs and melatonin in the sea lamprey using the LC-MS/MS method.