Determination of high-energy phosphate compounds and inorganic phosphate by reversed-phase high-performance liquid chromatography: evaluation of myocardial metabolic status in aerobically perfused and hypoxic mouse heart

• Published in: Journal of chromatography. B, Biomedical sciences and applications Vol. 751; no. 2; pp. 229 - 236
• Main Authors: Tomasetti, Marco, Graciotti, Laura, Fulgenzi, Gianluca, Granata, Anna Luisa
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 25-02-2001
• Subjects: Animals; Chromatography, High Pressure Liquid - methods; High-energy phosphate; Hypoxia - metabolism; Inorganic phosphate; Mice; Mice, Inbred C57BL; Myocardium - chemistry; Myocardium - enzymology; Organophosphorus Compounds - analysis; Phosphates - analysis; Reproducibility of Results; Thymidine Phosphorylase - metabolism; Inorganic phosphate; High-energy phosphate
• ISSN: 0378-4347; 1387-2273
• DOI: 10.1016/S0378-4347(00)00470-9

The present paper describes a simple HPLC method designed for measuring high-energy phosphate (HEP) compounds in a single run and inorganic phosphate (P
i) in an other short run under the same HPLC conditions. Inorganic phosphate was estimated by using thymidine phosphorylase (EC 2.4.2.4) which catalyzes a reaction involving inorganic phosphate to produce 2-deoxyribose 1-phosphate and thymine. The thymine/P
i stoichiometry was 1. The method provides a reproducible instrument for evaluating myocardial high-energy metabolism under physiological and pathological conditions.