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High-sensitivity analyses of metabolites in biological samples by capillary electrophoresis using dynamic pH junction-sweeping.
Chem Rec. 2002; 2(6):397-404.CR

Abstract

Emerging fields of biochemical research, such as metabolomics, present challenges to current separation technologies because of the large number of metabolites present in a cell and their often low (submicromolar) concentration. Although capillary electrophoresis (CE) holds great promise as the method of choice for high-resolution separations of biological samples, it suffers from poor concentration sensitivity, especially with the use of UV detection. In CE, sweeping and dynamic pH junction represent two complementary on-line focusing techniques that have been used for sensitivity enhancement of hydrophobic and weakly acidic analytes, respectively. However, the application of either the sweeping or dynamic pH junction technique alone might, in some cases, be less effective for the analysis of certain sample mixtures. Recent work in the development of a hyphenated dynamic pH junction-sweeping technique is presented as an effective on-line method of preconcentration suitable for both hydrophilic (anionic) and hydrophobic (neutral) analytes. Sensitive analyses of flavin metabolites by CE with laser-induced fluorescence (LIF) detection is demonstrated in various biological matrixes, including cell extracts of Bacillus subtilis, pooled human plasma, as well as heat-deproteinized flavoenzymes. Enhanced analyte band narrowing and improved sensitivity is achieved for flavins using dynamic pH junction-sweeping compared to either sweeping or dynamic pH junction alone. This results in over a 1200-fold improvement in sensitivity relative to conventional injection methods, giving a limit of detection (LOD, defined as S/N = 3) of about 4.0 x 10(-12) M. Strategies for sensitive and more comprehensive analyses of other cell metabolites, including nucleotides, coenzymes, and steroids, are also discussed when using on-line focusing techniques in conjunction with multiplexed CE and UV detection.

Authors+Show Affiliations

Britz-McKibbin P
Himeji Institute of Technology, Graduate School of Science, Department of Material Sciences, Kamigori, Hyogo 678-1297, Japan. philbmck@sci.himeji-tech.ac.jp
Terabe S
No affiliation info available

MeSH

Bacillus subtilisElectrophoresis, CapillaryFlavinsHumansHydrogen-Ion ConcentrationMetabolism

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Review

Language

eng

PubMed ID

12469351

Citation

Britz-McKibbin, Philip, and Shigeru Terabe. "High-sensitivity Analyses of Metabolites in Biological Samples By Capillary Electrophoresis Using Dynamic pH Junction-sweeping." Chemical Record (New York, N.Y.), vol. 2, no. 6, 2002, pp. 397-404.
Britz-McKibbin P, Terabe S. High-sensitivity analyses of metabolites in biological samples by capillary electrophoresis using dynamic pH junction-sweeping. Chem Rec. 2002;2(6):397-404.
Britz-McKibbin, P., & Terabe, S. (2002). High-sensitivity analyses of metabolites in biological samples by capillary electrophoresis using dynamic pH junction-sweeping. Chemical Record (New York, N.Y.), 2(6), 397-404.
Britz-McKibbin P, Terabe S. High-sensitivity Analyses of Metabolites in Biological Samples By Capillary Electrophoresis Using Dynamic pH Junction-sweeping. Chem Rec. 2002;2(6):397-404. PubMed PMID: 12469351.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - High-sensitivity analyses of metabolites in biological samples by capillary electrophoresis using dynamic pH junction-sweeping. AU - Britz-McKibbin,Philip, AU - Terabe,Shigeru, PY - 2002/12/7/pubmed PY - 2004/7/15/medline PY - 2002/12/7/entrez SP - 397 EP - 404 JF - Chemical record (New York, N.Y.) JO - Chem Rec VL - 2 IS - 6 N2 - Emerging fields of biochemical research, such as metabolomics, present challenges to current separation technologies because of the large number of metabolites present in a cell and their often low (submicromolar) concentration. Although capillary electrophoresis (CE) holds great promise as the method of choice for high-resolution separations of biological samples, it suffers from poor concentration sensitivity, especially with the use of UV detection. In CE, sweeping and dynamic pH junction represent two complementary on-line focusing techniques that have been used for sensitivity enhancement of hydrophobic and weakly acidic analytes, respectively. However, the application of either the sweeping or dynamic pH junction technique alone might, in some cases, be less effective for the analysis of certain sample mixtures. Recent work in the development of a hyphenated dynamic pH junction-sweeping technique is presented as an effective on-line method of preconcentration suitable for both hydrophilic (anionic) and hydrophobic (neutral) analytes. Sensitive analyses of flavin metabolites by CE with laser-induced fluorescence (LIF) detection is demonstrated in various biological matrixes, including cell extracts of Bacillus subtilis, pooled human plasma, as well as heat-deproteinized flavoenzymes. Enhanced analyte band narrowing and improved sensitivity is achieved for flavins using dynamic pH junction-sweeping compared to either sweeping or dynamic pH junction alone. This results in over a 1200-fold improvement in sensitivity relative to conventional injection methods, giving a limit of detection (LOD, defined as S/N = 3) of about 4.0 x 10(-12) M. Strategies for sensitive and more comprehensive analyses of other cell metabolites, including nucleotides, coenzymes, and steroids, are also discussed when using on-line focusing techniques in conjunction with multiplexed CE and UV detection. SN - 1527-8999 UR - https://www.unboundmedicine.com/medline/citation/12469351/High_sensitivity_analyses_of_metabolites_in_biological_samples_by_capillary_electrophoresis_using_dynamic_pH_junction_sweeping_ DB - PRIME DP - Unbound Medicine ER -