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Updated: Jun 9, 2026
Real-time Analyses of Retinol Transport by the Membrane Receptor of Plasma Retinol Binding Protein
Published on: January 28, 2013
Rapid determination of vitamin B₂ (riboflavin) in plasma by HPLC
Brian J Petteys1, Elizabeth L Frank
1ARUP Institute for Clinical and Experimental Pathology, Salt Lake City, UT 84108, USA.
Background:
Riboflavin (vitamin B₂), as the exclusive source for the coenzymes flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) in humans, is a water-soluble vitamin critical for metabolism and energy production. In its coenzyme forms, riboflavin is involved in essential oxidation-reduction reactions. Deficiency leads to skin and mucosal disorders. Measurement of plasma riboflavin can be used to assess vitamin B₂ status in at-risk individuals.
Methods:
Proteins are removed from plasma by acid precipitation. An aliquot of the resulting supernatant is analyzed by reversed-phase HPLC. Impurities are separated from riboflavin isocratically and the target material is detected fluorometrically (excitation 450 nm; emission 520 nm).
Results:
The method was validated for linearity, limit of quantification, accuracy, precision, and interference. The method was accurate and correlated well (R² = 0.993) to expected concentrations of spiked pooled plasma samples. Imprecision was < 10%. Riboflavin concentrations were determined in samples obtained from self-reported healthy adults who were not taking vitamin supplements. The reference interval established by nonparametric analysis was 6.7-50.1 nmol/l.
Conclusions:
This HPLC method allows separation and measurement of riboflavin in plasma in 7 min. Results from the assay may be used for clinical diagnosis of deficiency and to monitor therapeutic vitamin supplementation regimes.
