Optimization of capillary electrophoretic method for determination of physiologically relevant nucleotides and coenzymes in cell extract
| Authors | |
|---|---|
| Year of publication | 2010 |
| Type | Article in Proceedings |
| Conference | CHEMICA 47S |
| MU Faculty or unit | |
| Citation | |
| Field | Biochemistry |
| Keywords | Nukleotid-Koenzym-Buněčný extrakt |
| Description | Due to importance of nucleotides and coenzymes for metabolism, the main aim of this work was to optimize capillary electrophoretic method for the selective and rapid determination of 17 energetically important metabolites (purine and pyrimidine nucleotides, adenine coenzymes and Acetyl CoA). For improvement of concentration sensitivity, the capillary zone electrophoresis (CZE) was combined with the on-line preconcentration technique -field enhanced sample stacking. The optimal separation was reached by using voltage 23 kV (positive polarity)and direct detection at 260, 280 and 340 nm. 50 mM concentration of phosphate buffer (pH 5.8) provided the best resolution of peaks. Resulting method showed RSD in the range from 0.65 to 1.40 % for migration time and from 2.37 to 5.99 % for relative peak area of metabolites (n=10). Optimized method was used for analyzing of cell-extract of bacterium Paracoccus denitrificans. |
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