Fluorescent Derivatization Method of Proteins for Characterization by Capillary Electrophoresis- Sodium Dodecyl Sulfate with Laser-Induced Fluorescence Detection

2007 ◽  
Vol 79 (15) ◽  
pp. 5963-5971 ◽  
Author(s):  
David A. Michels ◽  
Lowell J. Brady ◽  
Amy Guo ◽  
Alain Balland
Keyword(s):  
Capillary Electrophoresis ◽  
Sodium Dodecyl Sulfate ◽  
Fluorescence Detection ◽  
Sodium Dodecyl ◽  
Laser Induced Fluorescence ◽  
Laser Induced Fluorescence Detection ◽  
Dodecyl Sulfate

scholarly journals Quantification of Riboflavin, Flavin Mononucleotide, and Flavin Adenine Dinucleotide in Human Plasma by Capillary Electrophoresis and Laser-induced Fluorescence Detection

1999 ◽  
Vol 45 (6) ◽  
pp. 862-868 ◽  
Author(s):  
Steinar Hustad ◽  
Per Magne Ueland ◽  
Jørn Schneede
Keyword(s):  
Capillary Electrophoresis ◽  
Human Plasma ◽  
Fluorescence Detection ◽  
Limit Of Detection ◽  
Sodium Dodecyl ◽  
Laser Induced Fluorescence ◽  
Fused Silica ◽  
Flavin Mononucleotide ◽  
Laser Induced Fluorescence Detection

Abstract Background: Riboflavin is the precursor of flavin mononucleotide (FMN) and FAD, which serve as cofactors for several redox enzymes. We have developed a capillary electrophoresis method for the determination of riboflavin and its two coenzyme forms in human plasma. Methods: Trichloroacetic acid-treated plasma was subjected to solid-phase extraction on reversed-phase columns. The analytes were separated by micellar electrokinetic capillary chromatography in uncoated fused- silica capillaries filled with borate buffer containing 50 mmol/L sodium dodecyl sulfate, methanol, and N-methylformamide. Native fluorescence was monitored at 530 nm, using an argon laser operating at 488 nm as excitation source. Results: The assay was linear over a concentration range of two orders of magnitude, and the limit of detection was far below physiological concentrations for all vitamers. The within-day and between-day coefficients of variation were 4–9% and 6–12%, respectively. The reference values (median, 5–95 percentiles) obtained by analyzing plasma from 63 healthy subjects were 8.6 nmol/L (2.7–42.5 nmol/L) for riboflavin, 7.0 nmol/L (3.5–13.3 nmol/L) for FMN, and 57.9 nmol/L (44.5–78.1 nmol/L) for FAD. Conclusions: Capillary electrophoresis with laser-induced fluorescence detection allows determination of all riboflavin vitamers far below physiological concentrations. The method may become a useful tool for the assessment of riboflavin status in humans.

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