Development of Liquid Chromatography methods coupled to Tandem Mass Spectrometry (LC-MS/MS) for monitoring the metabolites of new catecholamines

• Grant number: 23/15165-7
• Support Opportunities: Scholarships in Brazil - Support Program for Fixating Young Doctors
• Effective date (Start): November 01, 2023
• Effective date (End): October 31, 2024
• Field of knowledge: Biological Sciences - Pharmacology - Clinical Pharmacology
• Acordo de Cooperação: CNPq
• Principal Investigator: Gilberto de Nucci
• Grantee: Luiz Fernando Ribeiro
• Host Institution: Faculdade de Ciências Médicas (FCM). Universidade Estadual de Campinas (UNICAMP). Campinas , SP, Brazil
• Associated research grant: 23/01267-2 - An evolutionary perspective on endothelial cardiovascular control in vertebrates and its functions, AP.R

Abstract

We propose the detection and quantification of metabolites of 6-nitrodopamine, 6-cyanodopamine, 6-nitrodopa, 6-nitroadrenaline and 6-bromodopamine, new endogenous catecholamines recently discovered in tissues such as human umbilical cord, ventricles and atria of rabbits and rats, endothelium of snakes and aortic rings of turtles. 6-nitrodopamine was also considered the most potent endogenous inotropic agent ever discovered in rat hearts, being more potent than adrenaline and noradrenaline (a thousand times) and dopamine (ten thousand times). Substitution of the 6-position of the benzene ring of dopamine theoretically results in metabolites such as 6-nitrohomovanillic acid, 6-cyano homovanillic acid, 6-bromo homovanillic acid, analogues of homovanillic acid, which is the well-known metabolite of dopamine. However, there has not yet been a chromatographic study of these metabolites, which could clarify the fragmentation routes. For example, the chromatographic separation of 6-nitrohomovanillic acid from 5-nitrohomovanillic acid would elucidate the mechanism of formation of nitrodopamines, since 5-nitrohomovanillic acid can only be formed from 3-nitrotyrosine. The incubation of new endogenous catecholamines with enzymes such as monoamine oxidase A, monoamine oxidase B and renalase would also clarify which species is metabolized and which is its metabolite. In this project, it is expected to adopt chromatographic strategies to increase the separation power of the metabolites of these new catecholamines. One possibility to increase the separation power of metabolites of similar polarity would be to manipulate the pH of the mobile phase to promote the selective protonation of species with high pKa, keeping low pKa species deprotonated, which would lead to differentiation by RPLC and HILIC. With this, it is expected to develop an analytical methodology for quantifying new catecholamines in urine, plasma and human liquor.