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Nucleotide isotachophoretic assay: Method and application for determination of ATP/ADP ratio in several rat tissues
Author(s) -
Pérez José A.,
Mateo Fátima,
MeléndezHevia Enrique
Publication year - 1982
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150030208
Subject(s) - isotachophoresis , creatine , chemistry , nucleotide , skeletal muscle , adenine nucleotide , biochemistry , chromatography , biology , anatomy , electrolyte , electrode , gene
Separation of ATP and ADP was achieved in isotachophoresis by using malonate, inorganic phosphate (Pi), creatine phosphate and lactate as spacers, which isolate these nucleotides from other UV‐absorbing products in rat tissues. By combining isotachophoresis with enzymatic end‐point analysis of several rat tissues, calibration for ATP and ADP was achieved obtaining an operative calibration curve that allows the use of isotachophoresis as the only analytical technique in further assays. ATP and ADP quantities of rat skeletal and smooth muscle, heart, liver, kidney and lung were determined by isotachophoresis. Individual variation was also calculated by analyzing 30 animals in the same control conditions, muscular tissues showing the greatest values for ATP and ADP, whereas kidney and lung show the smaller values. ATP/ADP ratio is discussed as a representative parameter for describing the energy content in biological tissues, there are also experimental reasons which imply that other product concentrations in this parameter not be included. This ATP/ADP ratio is calculated in the rat tissues studied, as well as its individual variation, obtaining the larger values for muscular tissues and the smaller for kidney. A great individual variation is found in kidney, followed by heart and skeletal muscle, and the most constant values are found in liver and smooth muscle. The results obtained in this calibration method for isotachophoresis assay suggest the use of a similar procedure in order to apply this technique for other nucleotide analysis.