Optimizing separation conditions for riboflavin, flavin mononucleotide and flavin adenine dinucleotide in capillary zone electrophoresis with laser-induced fluorescence detection

J Chromatogr A. 2002 Aug 30;968(1-2):229-39. doi: 10.1016/s0021-9673(02)00953-6.

Abstract

A method was developed for the quantitative determination of riboflavin, flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), using free solution capillary zone electrophoresis in uncoated fused-silica capillaries with laser-induced fluorescence (LIF) detection. Various factors influencing the separation and detection of flavin vitamers were investigated, including pH (5.5-10.5), concentration and nature of the run buffer (phosphate, borate and carbonate), applied voltage (15-30 kV), temperature (15-30 degrees C) and injection time. Optimal resolution and detection were obtained with a pH 9.8, 30 mM aqueous phosphate buffer at 15 degrees C and 30 kV of applied voltage. LIF detection was obtained with a He-Cd laser source using an excitation wavelength at 442 nm and lambda(em) > or = 515 nm. Riboflavin could be determined in the concentration ranges 0.5-350 microg/l with a rather low detection limit (LOD) down to 50 amol. The LODs of FAD and FMN were slightly higher, 300 and 350 amol, respectively. Combined with a simple clean-up procedure, the practical utility of this method is illustrated by the measurements of flavin derivates in foods and beverages, such as wines, milk, yoghurt and raw eggs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Buffers
  • Electrophoresis, Capillary / methods*
  • Flavin Mononucleotide / isolation & purification*
  • Flavin-Adenine Dinucleotide / isolation & purification*
  • Food Analysis
  • Hydrogen-Ion Concentration
  • Lasers
  • Osmolar Concentration
  • Reference Standards
  • Riboflavin / isolation & purification*
  • Sensitivity and Specificity
  • Spectrometry, Fluorescence / methods*

Substances

  • Buffers
  • Flavin-Adenine Dinucleotide
  • Flavin Mononucleotide
  • Riboflavin