Acta Medica Medianae
Bulevar Zorana Đinđića 81, 18000 Niš, Srbija
HPLC-FLD-DAD CHARACTERISTICS OF THE S-NITROSOGLUTATHIONE DERIVATIVES OBTAINED BY O-PHTHALALDEHYDE
Nataša Trutić1, Radmila Pavlović1, Goran Nikolić2, Maja Srećković1 i Tatjana Jovanović3
S-nitrosothiols (RSNO) are considered to be natural depo and a transport form of nitric oxide (NO); their physiological activity is realized by releasing NO, which stimulates vasodilatation, prevents platelet aggregation and damages originating from ischemia/reperfusion injury. The main reservoir of non-protein RSNO is S-nitrosoglutathione (GSNO) created by S-nitrosylation of glutathione (GSH), most widespread endogenous non-protein thiol. The aim of this study was to set up an efficient GSNO synthesis and to examine the possibility of creating fluorescent derivatives of GSNO with o-phthalaldehyde (OPA), perform their spectroscopic characterization and behavior in the system of high performance liquid chromatography (HPLC). Synthesized GSNO and its OPA derivative were analyzed by HPLC system by using fluorescent (FLD) and “diode array” (DAD) detection. We developed the synthetic procedure by which GSNO was obtained in a solid, anhydrous form with a good yield. After UV characterization of the obtained GSNO, derivation of GSH and GSNO with OPA reagent, with or without mercaptoethanol (ME), was performed, and the derived products were analyzed by HPLC-FLD-DAD system. We optimized the reaction conditions that had lead to the formation of a stable and fluorescent tricyclic isoindole derivative of GSNO with OPA-ME reagent, which is essential for the development of HPLC method that would be used for the quantification of GSNO in real biological systems. Acta Medica Medianae 2010;49(1):27-32.
Key words: S-nitosoglutathione, HPLC, DAD, FLD, o-phthalaldehyde