Vitamin B1 in whole blood - HPLC

Vitamin B1 in whole blood - HPLC

Order no.: 35000/RUO
100 Tests
  • Vitamin B1

No post-column derivatisation
Easy handling
Highly stable calibration standards and controls

Description

The present method allows the rapid and simple determination of thiamine diphosphate, an analogue form of vitamin B1, in an isocratic HPLC system using fluorescent detection. The usual post-column derivatisation which requires extensive equipment is replaced here by a simple pre-column derivatisation. Sample preparation is restricted to a single precipitation step after addition of the extraction buffer and subsequent derivatisation in a reaction vial. A selected HPLC column and a mobile phase optimised for this specific separation permit high performance in HPLC quantification.

Chromatogram

Technical Data

Method of Analysis

HPLC

Number of Tests

100

Parameter

Vitamin B1

Analysis Time

approx. 5 min

Column temperature

ambient (~25 °C)

Flow rate

1 ml/min

Specimen

Whole Blood

Sample Preparation

  • To 200 μl whole blood/calibrator/control add 100 μl Extraction Buffer.
  • Mix briefly (vortex).
  • Add 300 μl Precipitation Reagent, mix 30 s (vortex).
  • Centrifuge 5 min at 9000 x g.
  • Pipette 200 μl of the derivatisation mix into a new vial.
  • Add 100 μl of the supernatant and mix briefly.
  • Add 100 µl Neutralisation Buffer.
  • Add 100 μl Stabilisation Buffer and mix briefly.
  • Let stand for 20 min, then inject 50 μl into the HPLC system.

Sample Stability

Research samples are stable for 24 hours at +2 to +8°C (light protected in a closed vial). For longer storage periods (up to 6 months) keep research samples frozen below -18°C.

Injection Volume

50 µl

Additional Info

For the analysis of vitamin B1 in whole blood every isocratic system with fluorescence detector is suitable.

Wavelength

EX 367 nm
EM 435 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

Mobile Phase


Order no.: 35021/RUO

Mobile Phase, 10 x 1000 ml


Order no.: 35022/RUO

Extraction Buffer


Order no.: 37003/RUO

Precipitation Reagent


Order no.: 37004/RUO

Derivatisation Reagent 1


Order no.: 35005/RUO

Derivatisation Reagent 2


Order no.: 35006/RUO

Stabilisation Buffer


Order no.: 35007/RUO

Neutralisation Buffer


Order no.: 35009/RUO

Reaction vials


Order no.: 33005/RUO

Calibrators

Calibrators available separately

Controls

Controls available separately

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