Molecular and Cellular PharmacologyMicrosomal formation of nitric oxide and cyanamides from non-physiological N-hydroxyguanidines: N-hydroxydebrisoquine as a model substrate
Section snippets
Chemicals and reagents
Catalase, SOD, and human hemoglobin were purchased from Sigma Chemical Co. Oxyhemoglobin was prepared from hemoglobin as described previously [6] and SIN-1 was supplied by Alexis Co. NADPH (tetrasodium salt) as well as all other chemicals and solvents were from Merck unless otherwise stated.
Synthesis
The cyanamide and urea derivatives of debrisoquine were obtained by reaction of tetrahydroisochinoline and bromocyanogen or potassium cyanate, respectively, as described earlier [7]. N-Hydroxydebrisoquine
Qualitative and quantitative analysis
The incubation of N-hydroxydebrisoquine with microsomes and NADPH resulted not only in the urea derivative [14], but also the cyanamide derivative (Fig. 1). The evidence for its formation was obtained unambiguously by HPLC. Addition of the reference substrate to the incubation mixture gave rise to an increase in the area of the metabolic peak, even when the eluent was varied (data not shown). A representative HPLC chromatogram recorded after the incubation of N-hydroxydebrisoquine with
Metabolism of N-hydroxydebrisoquine
The microsomal incubation of N-hydroxydebrisoquine produced not only the urea but also the cyanamide derivative as the main metabolites. The formation of a cyanamide was unexpected since during previous studies, also with amidoximes, similar metabolites were not identified [5]. Very recently, oxidation of various N-hydroxyguanidines and amidoximes by rat liver microsomes was found to lead to the corresponding cyanamides and nitriles, respectively∗. Moreover, investigations on the purely
Acknowledgements
The financial support of the “Fonds der Chemischen Industrie” is gratefully acknowledged.
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