Quantification of 6-nitrodopamine in Krebs-Henseleit’s solution by LC-MS/MS for the assessment of its basal release from Chelonoidis carbonaria aortae in vitro

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Vol. 1173; p. 122668
• Main Authors: Campos, Rafael, Pinheiro, David Halen Araújo, Britto-Júnior, José, de Castro, Heleson Alves, Mendes, Gustavo Duarte, Moraes, Manoel Odorico, Moraes, Maria Elisabete A., Lopes-Martins, Rodrigo Álvaro Brandão, Antunes, Natalícia J., De Nucci, Gilberto
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 30.05.2021
• Subjects: 6-nitrodopamine; Endothelium; LC-MS/MS; Solid phase extraction; Tortoise; LC-MS/MS; Solid phase extraction; 6-nitrodopamine; Tortoise; Endothelium
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2021.122668

•A novel method for analysis of 6-nitrodopamine in Krebs-Henseleit’s solution is presented.•The method was linear in the range of 0.1–20.0 ng/mL.•Tortoise’s aortae display a basal endothelium-derived 6-nitrodopamine release. In this study, the development and validation of a method for quantification of 6-nitrodopamine in Krebs-Henseleit’s solution by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) with positive ion electrospray ionization is described. Aortic rings taken from tortoise were either denuded or left with endothelium intact (15 mm, N = 6) and were incubated for 30 min in 5 mL Krebs-Henseleit’s solution in an organ bath. Solid phase extraction (SPE) was performed for aliquots of 1 mL of the supernatant. The separation of 6-nitrodopamine was obtained on a 150 mm × 3 mm Shim-pack GIST-HP C18 column, using 75% of mobile phase A consisted of deionized water with 0.1% formic acid (v/v) and 25% of mobile phase B consisted of acetonitrile/deionized water (50/50, v/v) + 0.1% formic acid at a flow rate of 350 μL/min in an isocratic mode. The method was linear over the concentration range of 0.1–20 ng/mL. The method was sensitive, precise and accurate for the assessment of the basal release of 6-nitrodopamine from Chelonoidis carbonaria aortae in vitro. The mean ± SEM concentrations of 6-nitrodopamine released from endothelium-intact and endothelium-denuded aortae were 0.44 ± 0.06 ng/mL and 0.18 ± 0.05 ng/mL, respectively. These results indicate that tortoise’s aortae display a basal endothelium-derived 6-nitrodopamine release.