Separation of ribonucleotides by capillary electrophoresis with multifunctional electrophoretic media of phosphate ammonium salts
Department of Chemistry, National University of Singapore, Singapore 119260, SingaporeJournal of Chromatography A (Impact Factor: 4.17). 08/1997; 779(1-2):215-225. DOI: 10.1016/S0021-9673(97)00445-7
Baseline separation of all common ribonucleotides by capillary electrophoresis (CE) was accomplished with bare fused-silica capillaries of 30 cm effective length and multifunctional electrophoretic media of simple composition, i.e., only phosphate ammonium salts, of moderate pH values in less than 10 min. A separation efficiency of 8·105/m in terms of theoretical plate number and a resolution between all adjacent peaks greater than 2 were obtained in capillaries of 75.0 cm effective length. Buffers containing phosphate anions resulted in good peak shape in the CE separation of the ribonucleotides. Among the phosphate buffers with different cations, potassium phosphate buffer showed better resolution than sodium phosphate buffer, but ammonium phosphate buffer gave the best resolution for the ribonucleotides. Inorganic ammonium cations affected the mobilities of ribonucleotides and enhanced their resolution during separation by ionic interaction and hydrogen-bonding. The migration behavior of the ribonucleotides was examined over the pH range from 5.5 to 9.0. In addition, the effects of organic solvents, such as acetonitrile,on the migration order of the ribonucleotides were studied. The CE method developed has proven to be a fast and simple means for the separation of the ribonucleotides with high efficiency and resolution. The results obtained demonstrated that buffers of phosphate ammonium salts have unique characteristics and may be excellent electrophoretic media for the separation of the ribonucleotides and other biochemical compounds by CE.
- [Show abstract] [Hide abstract]
ABSTRACT: The CE separation of twelve nucleotides (5′-mono-, di-, triphosphates of adenosine, guanosine, cytidine and uridine) was improved by adding cadmium ion to the ammonium citrate/citric acid buffer (pH 5, ionic strength 100 mM). Cadmium ion acts as a complexing agent for some nucleotides (ATP, CTP, GTP, UTP, GDP). In order to accelerate the separation, the electroosmotic flow was reversed by flushing the fused-silica capillary with 0.2 % aqueous solution of the polycationic surfactant hexadimethrine bromide. A good separation of the twelve nucleotides studied was then achieved on a dynamically coated capillary in less than 5 min by using an ammonium citrate/citric acid buffer (pH 5, ionic strength 100 mM) to which 2 mM cadmium ion has been added. High peak efficiencies were obtained (210 000 theoretical plates) and the resolution between two adjacent peaks was always greater than 1.5.Chromatographia 04/1999; 49(7):379-384. DOI:10.1007/BF02467610 · 1.41 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: An open-tubular wall-coated macrocyclic polyamine capillary column (70 cm x 75 microm ID) with 50 cm effective length for the separation of nucleoside monophosphates is described. Some parameters with respect to concentration, pH, composition of the buffer, and voltage in order to optimize the separation were studied. The coated capillary showed reversed electroosmotic flow (EOF), allowing anions to be separated in the co-EOF mode. Baseline separations were achieved for the eight nucleotides in less than 26 min using a background electrolyte consisting of H(3)PO(4)-NaH(2)PO(4) (30 mM, pH 3.10), an applied voltage of -15 kV, and detection at 254 nm. The macrocyclic polyamine on the capillary wall introduced anion coordination for the interaction with the analytes, the strength of which could be moderated by the type and concentration of the competing ion used in the background electrolyte (BGE). With a low concentration of the competing ion (phosphate ion), the migration behavior followed that obtained in the electrophoretic system. Increasing the concentration of the competing ion resulted in a faster migration and more complete elution of the analyte. The method established was also employed for the analysis of nucleotides in mushrooms. Aqueous extracts of mushrooms from different species and various extraction methods were injected directly for the analysis. Uridine 5'-monophosphate, guanosine 5'-monophosphate, adenosine 5'-monophosphate, and cytidine 5'-monophosphate, were found in the sample tested.Electrophoresis 05/2002; 23(9):1230-8. DOI:10.1002/1522-2683(200205)23:9<1230::AID-ELPS1230>3.0.CO;2-D · 3.03 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: This review is focused on the capillary electrophoresis-mass spectrometric (CE-MS) analysis of nucleic acid constituents in the broadest sense, going from nucleotides and adducted nucleotides over nucleoside analogues to oligonucleotides. These nucleic acid constituents play an important role in a variety of biochemical processes. Hence, their isolation, identification, and quantification will undoubtedly help reveal the process of life and disease mechanisms, such as carcinogenesis, and can also be useful for antitumor and antiviral drug research to provide valuable information about mechanism of action, pharmacokinetics, pharmacodynamics, toxicity, therapeutic drug level monitoring, and quality control related to this substance class. Fundamental investigations into their structure, the search for modifications, the occurrence and biochemical impact of structural variation amongst others, are therefore of great value. In view of the related bioanalytical procedures, the coupling of CE to MS has emerged as a powerful tool for the analysis of the complex mixtures of nucleic acid constituents: CE confers rapid analysis and efficient resolution, while MS provides high selectivity and sensitivity with structural characterization of minute amounts of compound. After an introduction about the biochemical and analytical perspectives on the nucleic acid constituents, the different modes of CE used in this field of research as well as the relevant CE-MS interfaces and the difficulties associated with quantitative CE-MS are briefly discussed. A large section is finally devoted to field-oriented applications.Electrophoresis 04/2005; 26(7-8):1221-53. DOI:10.1002/elps.200410278 · 3.03 Impact Factor
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.