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Fresenius' Journal of Analytical Chemistry

, Volume 368, Issue 5, pp 516–521 | Cite as

Determination of direct-bilirubin by a fluorimetric-enzymatic method based on bilirubin oxidase

  • Y. Andreu
  • J. Galbán
  • S. de Marcos
  • J. R. Castillo
Original Paper

Abstract

A method for the determination of direct bilirubin by reaction with bilirubin oxidase (BOx) is reported. The procedure is based on the changes in fluorescence which take place during the enzymatic reaction of BOx with any of the three forms of bilirubin (free, conjugated and with albumin) when the solution is excited at 240 nm and the emission is measured at 440 nm. The change in fluorescence was studied thoroughly. It seems mainly due to the fluorescence of one of the reaction products. A theoretical study was carried out to relate the changes in fluorescence observed to the species taking part in the reaction and to establish some of the enzymatic reaction constants. The optimum reaction conditions were studied for each of the three types of bilirubin together with their analytical characteristics (linear range and precision). Selective determination of direct bilirubin was carried out for various synthetic samples with good results. A linear response up to 7 mg L–1 of direct bilirubin was obtained. Using optimum conditions, the precision for free and conjugated bilirubin was 3.4% (n = 5) and 3.0% (n = 5), respectively.

Keywords

Albumin Optimum Condition Bilirubin Enzymatic Reaction Linear Range 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Springer-Verlag Berlin Heidelberg 2000

Authors and Affiliations

  • Y. Andreu
    • 1
  • J. Galbán
    • 1
  • S. de Marcos
    • 1
  • J. R. Castillo
    • 1
  1. 1.GEAS, Department of Analytical Chemistry, Science Faculty, University of Zaragoza, Zaragoza 50009, Spain e-mail: jgalban@posta.unizar.esES

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