37000 HPLC kit vitamin B2 whole blood

Vitamin B2 in Whole Blood - HPLC

Order no.: 37000
100 Tests
  • Flavin adenine dinucleotide (FAD)
  • Flavin mononucleotide (FMN)
  • Vitamin B2 (Riboflavin)

Safe results by stabilisation of FAD
Simultaneous determination of FMN
Highly stable calibrators and controls

Description

Clinical relevance

Vitamin B2 (riboflavin) belongs to the vitamin B complex, a heterogeneous group of water-soluble vitamins that are all precursors of coenzymes. The coenzymes flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN) are derived from riboflavin. In their capacity as a prosthetic group in flavoenzymes, they are responsible for the transfer of hydrogen in the respiratory chain. They also play a role in the metabolism of carbohydrates, fatty acids, and amino acids. In addition to its influence on other vitamins such as vitamin B6, riboflavin is essential for maintaining the glutathione reductase activity, thereby having an indirect antioxidative effect.

In Western Europe, a deficiency of riboflavin is observed particularly with vegans, smokers, diabetics and as a result of excessive consumption of alcohol. Typical signs of a vitamin B2 deficiency are poorly healing cracks at the corners of the mouth, inflammation of the oral mucosa and tongue, conjunctivitis and skin inflammation. A severe vitamin B2 deficiency may also promote the development of cataracts.

 

Product advantages

  • Reliable measurement results due to stabilisation of FAD
  • Simultaneous determination of riboflavin, FMN and FAD
  • Stable calibrators and controls

 

The present method developed by Chromsystems allows for the fast, simple and specific determination of riboflavin and its active metabolites FAD and FMN in whole blood using an isocratic HPLC system. Due to their native fluorescence, the analytes can be detected directly, without derivatisation. Sample preparation requires only one effective protein precipitation step and subsequent stabilisation of the analytes. A special HPLC column and an optimised mobile phase enable precise and reliable quantification of the analytes.

Chromatogram

Technical Data

Method of Analysis

HPLC

Number of Tests

100

Parameter

Flavin adenine dinucleotide (FAD), Flavin mononucleotide (FMN), Vitamin B2 (Riboflavin)

Analysis Time

aproxx. 9 min

Limit of quantification

FAD 3 µg/l
FMN 1 µg/l
Riboflavin 2 µg/l

Linearity

FAD up to 1000 µg/l
FMN up to 500 µg/l
Riboflavin up to 500 µg/l

Recovery

FAD 106 %, FMN 100 %, Rivoflavin 108 %

Intraassay

CV < 3 %

Interassay

CV < 6 %

Gradient

Isocratic

Information

For the HPLC analysis of vitamin B2 any isocratic HPLC system with fluorescence detector is suitable.

Column temperature

~20 °C

Flow rate

1.2 ml/min

Specimen

Whole Blood

Pre-analytic Treatment

Patient samples are stable up to 1 day stored in the dark at +2 to +8 °C. Samples not analysed immediately must be stored below -18 °C.

Sample Preparation

  • Pipette the following solutions into an amber reaction vial (light protected) and mix immediately for 30 s (vortex):
    200 μl whole blood (or reconstituted calibrator/control)
    + 200 μl Extraction Buffer
    + 400 μl Precipitation Reagent.

  • Centrifuge for 10 minutes at a minimum of 9000 x g.

  • Pipette 100 μl of Neutralisation Buffer into an amber autosampler vial (light protected), then add 500 μl supernatant and mix well (vortex).

  • Inject 50 μl of this mixture into the HPLC system.

Sample Stability

Protect samples from light. In a closed vial the samples are stable for up to 1 day at +2 to +8 °C. Samples not analysed immediately within 1 day must be kept deep-frozen below -18 °C.

Injection Volume

50 µl

Flow

1.2 ml/min

Additional Info

For the HPLC analysis of vitamin B2 any isocratic HPLC system with fluorescence detector is suitable.

Wavelength

EX 465 nm

EM 525 nm

Please note

The freely available information on this website, in particular on the sample preparation, are not sufficient to work with our products. Please read instructions and warning notices on products and/or instruction manuals.

Kit Components

Components available separately

Calibrators

Calibrators available separately

Controls

Controls available separately

Your Feedback is Appreciated

* Required Fields