| Popis: |
Although lizard brains are simpler than mammalian brains, they share common neural circuits in the brainstem responsible for motivational behavior, including aggression. That allows the study of basic mechanisms of behavioral control, the results of which could be translated to other species in order to elucidate the axis linking neurotransmitter levels to the regulation of aggressive behavior. The aim of this work was to transfer a previously developed LC-DAD/FLD method for analysis of neurotransmitters in lizard brain to mass spectrometry detection that provided the required analytical sensitivity for our samples. Reverse‐phase chromatography with gradient elution was performed to separate monoamines from the endogenous molecules in the matrix, followed by MS detection using positive and negative ion multiple reaction monitoring mode. A reliable LC-MS/MS method was established for simultaneous determination and quantification of eleven analytes - serotonin (5-HT), dopamine (DA), L-3, 4- dihydroxyphenylalanine (L-DOPA), noradrenaline (NA), adrenaline (A), tryptophan (TRP), tyrosine (TYR), 5-hydroxytryptophan (5-HTP), 5- hydroxyindole-3-acetic acid (5-HIAA), vanillylmandelic acid (VMA) and homovanilic acid (HVA) in a single run. The mobile phase consisting of 0.1% formic acid in 5 mM ammonium formate in water (A) and 0.1% formic acid in 5 mM ammonium formate in acetonitrile/water (95:5) was performed on a Kromasil 100-5-C18 column with guard column. The analytical range of these monoamines was between 0.005-1, 00 ng/mL. The monoamine contents in the brains of two coexisting lizard species Podarcis siculus and Podarcis melisellensis (female and male) are presented. |
