Capillary electrophoretic method for nucleotide analysis in cells: application on inherited metabolic disorders
Language English Country Germany Media print
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Cetrimonium Compounds chemistry MeSH
- Cetrimonium MeSH
- CHO Cells MeSH
- Cricetulus MeSH
- Electrophoresis, Capillary methods MeSH
- Erythrocytes chemistry MeSH
- gamma-Aminobutyric Acid chemistry MeSH
- Hydrogen-Ion Concentration MeSH
- Cricetinae MeSH
- Citric Acid chemistry MeSH
- Humans MeSH
- Purine Nucleotides analysis MeSH
- Pyrimidine Nucleotides analysis MeSH
- Animals MeSH
- Check Tag
- Cricetinae MeSH
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Cetrimonium Compounds MeSH
- Cetrimonium MeSH
- gamma-Aminobutyric Acid MeSH
- Citric Acid MeSH
- Purine Nucleotides MeSH
- Pyrimidine Nucleotides MeSH
Purine and pyrimidine nucleotides influence many metabolic pathways and their analogs have been widely used in medicine. A capillary electrophoretic method was developed for measuring intracellular nucleotides. The final BGE consisted of 40 mM citric acid with addition of 0.8 mM CTAB titrated by gamma-aminobutyric acid to pH 4.4. The electrophoretic separations were carried out in an uncoated silica capillary (id/od - 75/375 microm; effective/total length - 90/97 cm). The method allows a complete separation of 21 nucleotides and deoxynucleotides within 15 min with separation efficiencies up to 400,000 theoretical plates per meter. Due to the use of an acidic separation medium, the method offers a high selectivity toward the studied analytes versus possible interferences from matrices. Sample preparation was optimized in order to shorten work-time and prevent analyte degradation. The method was applied for analyzing nucleotides in human erythrocytes and Chinese hamster ovary cells. Diagnostic potential for inherited metabolic disorders of nucleotide metabolism is presented.
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