title: Rapid and accurate determination of deoxyribonucleoside monophosphates from DNA using micellar electrokinetic chromatography with a cationic surfactant additive.
abstract: A micellar electrokinetic chromatography (MEKC) method for rapid and accurate determination of 2'-deoxyribonucleoside 5'-monophosphates (dNMPs), four structural elements of DNA, is described. MEKC separation at an optimized pH enabled complete separation of four dNMPs. The use of a cationic surfactant additive for MEKC led to the reversal of EOF, which enhanced the migration velocities of the negatively charged dNMPs. Under the optimized condition, full-baseline separation of the four dNMPs assuring accurate peak integration was obtained within 5 min. For the given separation condition, pH-mediated on-column sample stacking was optimized and applied to enhance sensitivity up to 6-fold. Analytical precision was improved by spiking iothalamate as an internal standard. The accuracy of dNMP quantitation was ensured with dNMP standard solutions determined by inductively coupled plasma-optical emission spectroscopy that measured phosphorous quantity. Performance of the proposed method was ultimately proven by accurate quantitation of a DNA oligonucleotide that was enzymatically hydrolyzed prior to dNMP analysis. The proposed MEKC method turned out to be a reliable analytical method for dNMPs that features high speed, high sensitivity, and high precision, and could be utilized for high-accuracy determination of the amount of DNA as well as the base composition of DNA.
authors: Center for Bioanalysis, Korea Research Institute of Standards and Science, Daejeon, 1 Doryong-dong, Yusung-gu, Daejeon, 305-340, South Korea.
authors: Nan-Sook
authors: NS
authors: Hong
authors: Lian Hua
authors: LH
authors: Shi
authors: Ji Seon
authors: JS
authors: Jeong
authors: Inchul
authors: I
authors: Yang
authors: Sook-Kyung
authors: SK
authors: Kim
authors: Sang-Ryoul
authors: SR
authors: Park
mesh_terms: D002138:Calibration; D002412:Cations; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods*; D004247:DNA / Q000737:chemistry*; D003853:Deoxyribonucleosides / Q000032:analysis*; D013501:Surface-Active Agents / Q000737:chemistry*
chemical_list: D002412:Cations; D003853:Deoxyribonucleosides; D013501:Surface-Active Agents; D004247:DNA