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Improved high-performance liquid chromatographic assay for the determination of “high-energy” phosphates in mammalian skeletal muscle: Application to a single-fibre study in man |
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| Authors: | C. Karatzaferi A. De Haan C. Offringa A. J. Sargeant |
| Affiliation: | a Neuromuscular Biology Research Group, Manchester Metropolitan University, Alsager ST7 2HL, UK b Institute for Fundamental and Clinical Human Movement Sciences, Vrije University, Amsterdam, The Netherlands |
| Abstract: | A sensitive and reproducible method for the determination of adenine nucleotides (ATP, IMP) and creatine compounds [creatine (Cr), phosphocreatine (PCr)] in freeze–dried single human muscle fibre fragments is presented. The method uses isocratic reversed-phase high-performance liquid chromatography of methanol extracts. Average retention times (min) of ATP, IMP and PCr, Cr from standard solutions were 10.6±0.42, 2.11±0.06 (n=6) and 10.5±0.31 and 1.19±0.02 (n=9), respectively. Detection limits in extracts from muscle fibre fragments were 2.0, 1.0, 3.0 and 2.0 mmol/kg dm, respectively. The assay was found successful for analysis of fibre-fragments weighing ≥1 μg. |
| Keywords: | Phosphocreatine Creatine Adenosine triphosphate Inosine monophosphate |
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